Squirrels

Lecture 2

Functions of proteins

The chemical composition of proteins

Characterization of proteinogenic amino acids

Structure of proteins

Protein classification

Properties of proteins and research methods

Proteins are structural components of organs and tissues, exhibit enzymatic activity (enzymes), involved in the regulation of metabolism. Transport proteins that carry protons and electrons across membranes provide bioenergetics: light absorption, respiration, ATP production. Spare proteins (characteristic mainly of plants) are deposited in seeds and used to feed seedlings during germination. By burning ATP, proteins provide mechanical activities involved in the movement of the cytoplasm and other cell organelles. Important protective function of proteins: hydrolytic enzymes of lysosomes and vacuoles break down harmful substances that have entered the cell; glycoproteins are involved in plant defense against pathogens; proteins perform cryoprotective and antifreeze functions. One protein can perform two or more functions (some membrane proteins can perform structural and enzymatic functions).

The amazing variety of functions of proteins and the high prevalence are reflected in their name - proteins(from Greek " protos» - primary, most important). As a rule, the content of proteins in plants is lower than in animals: in vegetative organs, the amount of protein is usually 5-15% of the dry mass. Thus, timothy leaves contain 7% protein, while clover and wiki leaves contain 15%. More protein in seeds: in cereals, on average, 10-20%, in legumes and oilseeds - 25-35%. Soybean seeds are the richest in protein - up to 40%, and sometimes even higher.

In plant cells, proteins are usually associated with carbohydrates, lipids, and other compounds, as well as membranes, so they are difficult to extract and obtain pure preparations, especially from vegetative organs. In this regard, in plants, seed proteins are better studied, where they are more abundant and from where they are more easily extracted.

Proteins - organic compounds having the following elemental composition: carbon 51-55 %; oxygen 21-23 %; hydrogen 6,6-7,3 %; nitrogen 15-18 %; sulfur 0.3-2.4%. Some proteins also contain phosphorus (0,2-2 %), iron and other elements. A characteristic indicator of the elemental composition of proteins in all organisms is the presence of nitrogen, on average it is taken equal to 16 % . The relative constancy of this indicator makes it possible to use it for the quantitative determination of protein: the relative value of the content of protein nitrogen, in percent, is multiplied by the conversion factor - 6,25 (100:16=6.25). The chemical nature of proteins is heteropolymers built from leftovers amino acids. Amino acids (AA) organic compounds are called, in the molecules of which one or more hydrogen atoms are replaced amino groups(- NH 2).

The content of the article

PROTEINS (Article 1)- a class of biological polymers present in every living organism. With the participation of proteins, the main processes that ensure the vital activity of the body take place: respiration, digestion, muscle contraction, transmission of nerve impulses. Bone tissue, skin, hair, horn formations of living beings are composed of proteins. For most mammals, the growth and development of the organism occurs due to products containing proteins as a food component. The role of proteins in the body and, accordingly, their structure is very diverse.

The composition of proteins.

All proteins are polymers, the chains of which are assembled from fragments of amino acids. Amino acids are organic compounds containing in their composition (according to the name) an NH 2 amino group and an organic acid, i.e. carboxyl, COOH group. Of all the variety of existing amino acids (theoretically, the number of possible amino acids is unlimited), only those that have only one carbon atom between the amino group and the carboxyl group participate in the formation of proteins. In general, the amino acids involved in the formation of proteins can be represented by the formula: H 2 N–CH(R)–COOH. The R group attached to the carbon atom (the one between the amino and carboxyl groups) determines the difference between the amino acids that make up proteins. This group can consist only of carbon and hydrogen atoms, but more often contains, in addition to C and H, various functional (capable of further transformations) groups, for example, HO-, H 2 N-, etc. There is also an option when R \u003d H.

The organisms of living beings contain more than 100 different amino acids, however, not all are used in the construction of proteins, but only 20, the so-called "fundamental". In table. 1 shows their names (most of the names have developed historically), the structural formula, as well as the widely used abbreviation. All structural formulas are arranged in the table so that the main fragment of the amino acid is on the right.

Table 1. AMINO ACIDS INVOLVED IN THE CREATION OF PROTEINS

Name	Structure	Designation
GLYCINE		GLI
ALANIN		ALA
VALIN		SHAFT
LEUCINE		LEI
ISOLEUCINE		ILE
SERIN		SER
THREONINE		TRE
CYSTEINE		CIS
METIONINE		MET
LYSINE		LIZ
ARGININE		AWG
ASPARAGIC ACID		ACH
ASPARAGIN		ACH
GLUTAMIC ACID		GLU
GLUTAMINE		GLN
phenylalanine		hair dryer
TYROSINE		TIR
tryptophan		THREE
HISTIDINE		GIS
PROLINE		PRO
In international practice, the abbreviated designation of the listed amino acids using Latin three-letter or one-letter abbreviations is accepted, for example, glycine - Gly or G, alanine - Ala or A.

Among these twenty amino acids (Table 1), only proline contains an NH group (instead of NH 2) next to the COOH carboxyl group, since it is part of the cyclic fragment.

Eight amino acids (valine, leucine, isoleucine, threonine, methionine, lysine, phenylalanine and tryptophan), placed in the table on a gray background, are called essential, since the body must constantly receive them with protein food for normal growth and development.

A protein molecule is formed as a result of the sequential connection of amino acids, while the carboxyl group of one acid interacts with the amino group of the neighboring molecule, as a result, a –CO–NH– peptide bond is formed and a water molecule is released. On fig. 1 shows the serial connection of alanine, valine and glycine.

Rice. 1 SERIAL CONNECTION OF AMINO ACIDS during the formation of a protein molecule. The path from the terminal amino group H 2 N to the terminal carboxyl group COOH was chosen as the main direction of the polymer chain.

To compactly describe the structure of a protein molecule, the abbreviations for amino acids (Table 1, third column) involved in the formation of the polymer chain are used. The fragment of the molecule shown in Fig. 1 is written as follows: H 2 N-ALA-VAL-GLY-COOH.

Protein molecules contain from 50 to 1500 amino acid residues (shorter chains are called polypeptides). The individuality of a protein is determined by the set of amino acids that make up the polymer chain and, no less important, by the order of their alternation along the chain. For example, the insulin molecule consists of 51 amino acid residues (it is one of the shortest chain proteins) and consists of two interconnected parallel chains of unequal length. The sequence of amino acid fragments is shown in fig. 2.

Rice. 2 INSULIN MOLECULE, built from 51 amino acid residues, fragments of the same amino acids are marked with the corresponding background color. The amino acid residues of cysteine ​​(abbreviated designation CIS) contained in the chain form disulfide bridges -S-S-, which link two polymer molecules, or form jumpers within one chain.

Molecules of the amino acid cysteine ​​(Table 1) contain reactive sulfhydride groups -SH, which interact with each other, forming disulfide bridges -S-S-. The role of cysteine ​​in the world of proteins is special, with its participation, cross-links are formed between polymeric protein molecules.

The association of amino acids into a polymer chain occurs in a living organism under the control of nucleic acids, it is they that provide a strict assembly order and regulate the fixed length of the polymer molecule ( cm. NUCLEIC ACIDS).

The structure of proteins.

The composition of the protein molecule, presented in the form of alternating amino acid residues (Fig. 2), is called the primary structure of the protein. Hydrogen bonds arise between the imino groups HN present in the polymer chain and the carbonyl groups CO ( cm. HYDROGEN BOND), as a result, the protein molecule acquires a certain spatial shape, called the secondary structure. The most common are two types of secondary structure in proteins.

The first option, called the α-helix, is implemented using hydrogen bonds within one polymer molecule. The geometric parameters of the molecule, determined by the bond lengths and bond angles, are such that the formation of hydrogen bonds is possible for the H-N and C=O groups, between which there are two peptide fragments H-N-C=O (Fig. 3).

The composition of the polypeptide chain shown in fig. 3 is written in abbreviated form as follows:

H 2 N-ALA VAL-ALA-LEY-ALA-ALA-ALA-ALA-VAL-ALA-ALA-ALA-COOH.

As a result of the contracting action of hydrogen bonds, the molecule takes the form of a helix - the so-called α-helix, it is depicted as a curved helical ribbon passing through the atoms that form the polymer chain (Fig. 4)

Rice. 4 3D MODEL OF A PROTEIN MOLECULE in the form of an α-helix. Hydrogen bonds are shown as green dotted lines. The cylindrical shape of the spiral is visible at a certain angle of rotation (hydrogen atoms are not shown in the figure). The color of individual atoms is given in accordance with international rules, which recommend black for carbon atoms, blue for nitrogen, red for oxygen, and yellow for sulfur (white color is recommended for hydrogen atoms not shown in the figure, in this case the entire structure depicted on a dark background).

Another variant of the secondary structure, called the β-structure, is also formed with the participation of hydrogen bonds, the difference is that the H-N and C=O groups of two or more polymer chains located in parallel interact. Since the polypeptide chain has a direction (Fig. 1), variants are possible when the direction of the chains is the same (parallel β-structure, Fig. 5), or they are opposite (antiparallel β-structure, Fig. 6).

Polymer chains of various compositions can participate in the formation of the β-structure, while the organic groups framing the polymer chain (Ph, CH 2 OH, etc.) in most cases play a secondary role, the mutual arrangement of the H-N and C=O groups is decisive. Since the H-N and C=O groups are directed in different directions relative to the polymer chain (up and down in the figure), it becomes possible for three or more chains to interact simultaneously.

The composition of the first polypeptide chain in Fig. 5:

H 2 N-LEI-ALA-FEN-GLI-ALA-ALA-COOH

The composition of the second and third chain:

H 2 N-GLY-ALA-SER-GLY-TRE-ALA-COOH

The composition of the polypeptide chains shown in fig. 6, the same as in Fig. 5, the difference is that the second chain has the opposite (in comparison with Fig. 5) direction.

It is possible to form a β-structure within one molecule, when a chain fragment in a certain section turns out to be rotated by 180°, in this case, two branches of one molecule have the opposite direction, as a result, an antiparallel β-structure is formed (Fig. 7).

The structure shown in fig. 7 in a flat image, shown in fig. 8 in the form of a three-dimensional model. Sections of the β-structure are usually denoted in a simplified way by a flat wavy ribbon that passes through the atoms that form the polymer chain.

In the structure of many proteins, sections of the α-helix and ribbon-like β-structures alternate, as well as single polypeptide chains. Their mutual arrangement and alternation in the polymer chain is called the tertiary structure of the protein.

Methods for depicting the structure of proteins are shown below using the plant protein crambin as an example. Structural formulas of proteins, often containing up to hundreds of amino acid fragments, are complex, cumbersome and difficult to understand, therefore sometimes simplified structural formulas are used - without symbols of chemical elements (Fig. 9, option A), but at the same time they retain the color of valence strokes in accordance with international rules (Fig. 4). In this case, the formula is presented not in a flat, but in a spatial image, which corresponds to the real structure of the molecule. This method makes it possible, for example, to distinguish between disulfide bridges (similar to those found in insulin, Fig. 2), phenyl groups in the side frame of the chain, etc. The image of molecules in the form of three-dimensional models (balls connected by rods) is somewhat clearer (Fig. 9, option B). However, both methods do not allow showing the tertiary structure, so the American biophysicist Jane Richardson proposed to depict α-structures as spirally twisted ribbons (see Fig. 4), β-structures as flat wavy ribbons (Fig. 8), and connecting them single chains - in the form of thin bundles, each type of structure has its own color. This method of depicting the tertiary structure of a protein is now widely used (Fig. 9, variant B). Sometimes, for greater information content, the tertiary structure and the simplified structural formula are shown together (Fig. 9, variant D). There are also modifications of the method proposed by Richardson: α-helices are depicted as cylinders, and β-structures are in the form of flat arrows indicating the direction of the chain (Fig. 9, option E). Less common is the method in which the entire molecule is depicted as a bundle, where unequal structures are distinguished by different colors, and disulfide bridges are shown as yellow bridges (Fig. 9, variant E).

Option B is the most convenient for perception, when when depicting the tertiary structure, the structural features of the protein (amino acid fragments, their alternation order, hydrogen bonds) are not indicated, while it is assumed that all proteins contain “details” taken from a standard set of twenty amino acids ( Table 1). The main task in depicting a tertiary structure is to show the spatial arrangement and alternation of secondary structures.

Rice. 9 VARIOUS VERSIONS OF IMAGE OF THE STRUCTURE OF THE CRUMBIN PROTEIN.
A is a structural formula in a spatial image.
B - structure in the form of a three-dimensional model.
B is the tertiary structure of the molecule.
G - a combination of options A and B.
E - simplified image of the tertiary structure.
E - tertiary structure with disulfide bridges.

The most convenient for perception is a three-dimensional tertiary structure (option B), freed from the details of the structural formula.

A protein molecule that has a tertiary structure, as a rule, takes on a certain configuration, which is formed by polar (electrostatic) interactions and hydrogen bonds. As a result, the molecule takes the form of a compact coil - globular proteins (globules, lat. ball), or filamentous - fibrillar proteins (fibra, lat. fiber).

An example of a globular structure is the protein albumin, the protein of a chicken egg belongs to the class of albumins. The polymeric chain of albumin is assembled mainly from alanine, aspartic acid, glycine, and cysteine, alternating in a certain order. The tertiary structure contains α-helices connected by single chains (Fig. 10).

Rice. 10 GLOBULAR STRUCTURE OF ALBUMIN

An example of a fibrillar structure is the fibroin protein. They contain a large amount of glycine, alanine and serine residues (every second amino acid residue is glycine); cysteine ​​residues containing sulfhydride groups are absent. Fibroin, the main component of natural silk and cobwebs, contains β-structures connected by single chains (Fig. 11).

Rice. eleven FIBRILLARY PROTEIN FIBROIN

The possibility of forming a tertiary structure of a certain type is inherent in the primary structure of the protein, i.e. determined in advance by the order of alternation of amino acid residues. From certain sets of such residues, α-helices predominantly arise (there are quite a lot of such sets), another set leads to the appearance of β-structures, single chains are characterized by their composition.

Some protein molecules, while retaining a tertiary structure, are able to combine into large supramolecular aggregates, while they are held together by polar interactions, as well as hydrogen bonds. Such formations are called the quaternary structure of the protein. For example, the ferritin protein, which consists mainly of leucine, glutamic acid, aspartic acid and histidine (ferricin contains all 20 amino acid residues in varying amounts) forms a tertiary structure of four parallel-laid α-helices. When molecules are combined into a single ensemble (Fig. 12), a quaternary structure is formed, which can include up to 24 ferritin molecules.

Fig.12 FORMATION OF THE QUATERNARY STRUCTURE OF THE GLOBULAR PROTEIN FERRITIN

Another example of supramolecular formations is the structure of collagen. It is a fibrillar protein whose chains are built mainly of glycine alternating with proline and lysine. The structure contains single chains, triple α-helices, alternating with ribbon-like β-structures stacked in parallel bundles (Fig. 13).

Fig.13 SUPRAMOLECULAR STRUCTURE OF COLLAGEN FIBRILLARY PROTEIN

Chemical properties of proteins.

Under the action of organic solvents, waste products of some bacteria (lactic acid fermentation) or with an increase in temperature, secondary and tertiary structures are destroyed without damaging its primary structure, as a result, the protein loses solubility and loses biological activity, this process is called denaturation, that is, the loss of natural properties, for example, the curdling of sour milk, the coagulated protein of a boiled chicken egg. At elevated temperatures, the proteins of living organisms (in particular, microorganisms) quickly denature. Such proteins are not able to participate in biological processes, as a result, microorganisms die, so boiled (or pasteurized) milk can be stored longer.

Peptide bonds H-N-C=O, which form the polymer chain of the protein molecule, are hydrolyzed in the presence of acids or alkalis, and the polymer chain breaks, which, ultimately, can lead to the original amino acids. Peptide bonds that are part of α-helices or β-structures are more resistant to hydrolysis and various chemical influences (compared to the same bonds in single chains). A more delicate disassembly of the protein molecule into its constituent amino acids is carried out in an anhydrous medium using hydrazine H 2 N–NH 2, while all amino acid fragments, except for the last one, form the so-called carboxylic acid hydrazides containing the fragment C (O)–HN–NH 2 ( Fig. 14).

Rice. 14. POLYPEPTIDE CLEAVAGE

Such an analysis can provide information about the amino acid composition of a protein, but it is more important to know their sequence in a protein molecule. One of the methods widely used for this purpose is the action of phenylisothiocyanate (FITC) on the polypeptide chain, which in an alkaline medium attaches to the polypeptide (from the end that contains the amino group), and when the reaction of the medium changes to acidic, it detaches from the chain, taking with it fragment of one amino acid (Fig. 15).

Rice. 15 SEQUENTIAL POLYPEPTIDE Cleavage

Many special methods have been developed for such an analysis, including those that begin to “disassemble” a protein molecule into its constituent components, starting from the carboxyl end.

Cross disulfide bridges S-S (formed by the interaction of cysteine ​​residues, Fig. 2 and 9) are cleaved, turning them into HS-groups by the action of various reducing agents. The action of oxidizing agents (oxygen or hydrogen peroxide) again leads to the formation of disulfide bridges (Fig. 16).

Rice. 16. Cleavage of disulfide bridges

To create additional cross-links in proteins, the reactivity of amino and carboxyl groups is used. More accessible for various interactions are the amino groups that are in the side frame of the chain - fragments of lysine, asparagine, lysine, proline (Table 1). When such amino groups interact with formaldehyde, the process of condensation occurs and cross-bridges –NH–CH2–NH– appear (Fig. 17).

Rice. 17 CREATION OF ADDITIONAL TRANSVERSAL BRIDGES BETWEEN PROTEIN MOLECULES.

The terminal carboxyl groups of the protein are able to react with complex compounds of some polyvalent metals (chromium compounds are more often used), and cross-links also occur. Both processes are used in leather tanning.

The role of proteins in the body.

The role of proteins in the body is diverse.

Enzymes(fermentatio lat. - fermentation), their other name is enzymes (en zumh greek. - in yeast) - these are proteins with catalytic activity, they are able to increase the speed of biochemical processes by thousands of times. Under the action of enzymes, the constituent components of food: proteins, fats and carbohydrates are broken down into simpler compounds, from which new macromolecules are then synthesized, which are necessary for a certain type of body. Enzymes also take part in many biochemical processes of synthesis, for example, in the synthesis of proteins (some proteins help to synthesize others). Cm. ENZYMES

Enzymes are not only highly efficient catalysts, but also selective (direct the reaction strictly in the given direction). In their presence, the reaction proceeds with almost 100% yield without the formation of by-products and, at the same time, the flow conditions are mild: normal atmospheric pressure and temperature of a living organism. For comparison, the synthesis of ammonia from hydrogen and nitrogen in the presence of an activated iron catalyst is carried out at 400–500°C and a pressure of 30 MPa, the yield of ammonia is 15–25% per cycle. Enzymes are considered unsurpassed catalysts.

Intensive study of enzymes began in the middle of the 19th century; more than 2,000 different enzymes have now been studied; this is the most diverse class of proteins.

The names of enzymes are as follows: the name of the reagent with which the enzyme interacts, or the name of the catalyzed reaction, is added with the ending -aza, for example, arginase decomposes arginine (Table 1), decarboxylase catalyzes decarboxylation, i.e. elimination of CO 2 from the carboxyl group:

– COOH → – CH + CO 2

Often, to more accurately indicate the role of an enzyme, both the object and the type of reaction are indicated in its name, for example, alcohol dehydrogenase is an enzyme that dehydrogenates alcohols.

For some enzymes discovered quite a long time ago, the historical name (without the ending -aza) has been preserved, for example, pepsin (pepsis, Greek. digestion) and trypsin (thrypsis Greek. liquefaction), these enzymes break down proteins.

For systematization, enzymes are combined into large classes, the classification is based on the type of reaction, the classes are named according to the general principle - the name of the reaction and the ending - aza. Some of these classes are listed below.

Oxidoreductase are enzymes that catalyze redox reactions. The dehydrogenases included in this class carry out proton transfer, for example, alcohol dehydrogenase (ADH) oxidizes alcohols to aldehydes, the subsequent oxidation of aldehydes to carboxylic acids is catalyzed by aldehyde dehydrogenases (ALDH). Both processes occur in the body during the processing of ethanol into acetic acid (Fig. 18).

Rice. 18 TWO-STAGE OXIDATION OF ETHANOL to acetic acid

It is not ethanol that has a narcotic effect, but the intermediate product acetaldehyde, the lower the activity of the ALDH enzyme, the slower the second stage passes - the oxidation of acetaldehyde to acetic acid, and the longer and stronger the intoxicating effect from ingestion of ethanol. The analysis showed that more than 80% of the representatives of the yellow race have a relatively low activity of ALDH and therefore a markedly more severe alcohol tolerance. The reason for this innate reduced activity of ALDH is that part of the glutamic acid residues in the “attenuated” ALDH molecule is replaced by lysine fragments (Table 1).

Transferases- enzymes that catalyze the transfer of functional groups, for example, transiminase catalyzes the transfer of an amino group.

Hydrolases are enzymes that catalyze hydrolysis. The previously mentioned trypsin and pepsin hydrolyze peptide bonds, and lipases cleave the ester bond in fats:

–RC(O)OR 1 + H 2 O → –RC(O)OH + HOR 1

Liase- enzymes that catalyze reactions that take place in a non-hydrolytic way, as a result of such reactions, C-C, C-O, C-N bonds are broken and new bonds are formed. The enzyme decarboxylase belongs to this class

Isomerases- enzymes that catalyze isomerization, for example, the conversion of maleic acid to fumaric acid (Fig. 19), this is an example of cis-trans isomerization (see ISOMERIA).

Rice. 19. ISOMERIZATION OF MALEIC ACID into fumaric acid in the presence of the enzyme.

In the work of enzymes, the general principle is observed, according to which there is always a structural correspondence between the enzyme and the reagent of the accelerated reaction. According to the figurative expression of one of the founders of the doctrine of enzymes, E. Fisher, the reagent approaches the enzyme like a key to a lock. In this regard, each enzyme catalyzes a certain chemical reaction or a group of reactions of the same type. Sometimes an enzyme can act on a single compound, such as urease (uron Greek. - urine) catalyzes only the hydrolysis of urea:

(H 2 N) 2 C \u003d O + H 2 O \u003d CO 2 + 2NH 3

The finest selectivity is shown by enzymes that distinguish between optically active antipodes - left- and right-handed isomers. L-arginase acts only on levorotatory arginine and does not affect the dextrorotatory isomer. L-lactate dehydrogenase acts only on the levorotatory esters of lactic acid, the so-called lactates (lactis lat. milk), while D-lactate dehydrogenase only breaks down D-lactates.

Most of the enzymes act not on one, but on a group of related compounds, for example, trypsin "prefers" to cleave the peptide bonds formed by lysine and arginine (Table 1.)

The catalytic properties of some enzymes, such as hydrolases, are determined solely by the structure of the protein molecule itself, another class of enzymes - oxidoreductases (for example, alcohol dehydrogenase) can only be active in the presence of non-protein molecules associated with them - vitamins that activate Mg, Ca, Zn, Mn and fragments of nucleic acids (Fig. 20).

Rice. 20 ALCOHOLD DEHYDROGENASE MOLECULE

Transport proteins bind and transport various molecules or ions through cell membranes (both inside and outside the cell), as well as from one organ to another.

For example, hemoglobin binds oxygen as blood passes through the lungs and delivers it to various body tissues, where oxygen is released and then used to oxidize food components, this process serves as an energy source (sometimes the term “burning” of food in the body is used).

In addition to the protein part, hemoglobin contains a complex compound of iron with a cyclic porphyrin molecule (porphyros Greek. - purple), which determines the red color of the blood. It is this complex (Fig. 21, left) that plays the role of an oxygen carrier. In hemoglobin, the iron porphyrin complex is located inside the protein molecule and is retained by polar interactions, as well as by a coordination bond with nitrogen in histidine (Table 1), which is part of the protein. The O2 molecule, which is carried by hemoglobin, is attached via a coordination bond to the iron atom from the side opposite to that to which histidine is attached (Fig. 21, right).

Rice. 21 STRUCTURE OF THE IRON COMPLEX

The structure of the complex is shown on the right in the form of a three-dimensional model. The complex is held in the protein molecule by a coordination bond (dashed blue line) between the Fe atom and the N atom in histidine, which is part of the protein. The O 2 molecule, which is carried by hemoglobin, is coordinated (red dotted line) to the Fe atom from the opposite country of the planar complex.

Hemoglobin is one of the most studied proteins, it consists of a-helices connected by single chains and contains four iron complexes. Thus, hemoglobin is like a voluminous package for the transfer of four oxygen molecules at once. The form of hemoglobin corresponds to globular proteins (Fig. 22).

Rice. 22 GLOBULAR FORM OF HEMOGLOBIN

The main "advantage" of hemoglobin is that the addition of oxygen and its subsequent splitting off during transmission to various tissues and organs takes place quickly. Carbon monoxide, CO (carbon monoxide), binds to Fe in hemoglobin even faster, but, unlike O 2 , forms a complex that is difficult to break down. As a result, such hemoglobin is not able to bind O 2, which leads (when large amounts of carbon monoxide are inhaled) to the death of the body from suffocation.

The second function of hemoglobin is the transfer of exhaled CO 2, but not the iron atom, but the H 2 of the N-group of the protein is involved in the process of temporary binding of carbon dioxide.

The "performance" of proteins depends on their structure, for example, replacing the only amino acid residue of glutamic acid in the hemoglobin polypeptide chain with a valine residue (a rare congenital anomaly) leads to a disease called sickle cell anemia.

There are also transport proteins that can bind fats, glucose, amino acids and carry them both inside and outside the cells.

Transport proteins of a special type do not carry the substances themselves, but act as a “transport regulator”, passing certain substances through the membrane (the outer wall of the cell). Such proteins are often called membrane proteins. They have the shape of a hollow cylinder and, being embedded in the membrane wall, ensure the movement of some polar molecules or ions into the cell. An example of a membrane protein is porin (Fig. 23).

Rice. 23 PORIN PROTEIN

Food and storage proteins, as the name implies, serve as sources of internal nutrition, more often for the embryos of plants and animals, as well as in the early stages of development of young organisms. Dietary proteins include albumin (Fig. 10) - the main component of egg white, as well as casein - the main protein of milk. Under the action of the enzyme pepsin, casein curdles in the stomach, which ensures its retention in the digestive tract and efficient absorption. Casein contains fragments of all the amino acids needed by the body.

In ferritin (Fig. 12), which is contained in the tissues of animals, iron ions are stored.

Myoglobin is also a storage protein, which resembles hemoglobin in composition and structure. Myoglobin is concentrated mainly in the muscles, its main role is the storage of oxygen, which hemoglobin gives it. It is rapidly saturated with oxygen (much faster than hemoglobin), and then gradually transfers it to various tissues.

Structural proteins perform a protective function (skin) or support - they hold the body together and give it strength (cartilage and tendons). Their main component is the fibrillar protein collagen (Fig. 11), the most common protein of the animal world, in the body of mammals, it accounts for almost 30% of the total mass of proteins. Collagen has a high tensile strength (the strength of the skin is known), but due to the low content of cross-links in skin collagen, animal skins are not very suitable in their raw form for the manufacture of various products. To reduce the swelling of the skin in water, shrinkage during drying, as well as to increase the strength in the watered state and increase the elasticity in collagen, additional cross-links are created (Fig. 15a), this is the so-called leather tanning process.

In living organisms, collagen molecules that have arisen in the process of growth and development of the organism are not updated and are not replaced by newly synthesized ones. As the body ages, the number of cross-links in collagen increases, which leads to a decrease in its elasticity, and since renewal does not occur, age-related changes appear - an increase in the fragility of cartilage and tendons, the appearance of wrinkles on the skin.

Articular ligaments contain elastin, a structural protein that easily stretches in two dimensions. The resilin protein, which is located at the points of hinge attachment of the wings in some insects, has the greatest elasticity.

Horn formations - hair, nails, feathers, consisting mainly of keratin protein (Fig. 24). Its main difference is the noticeable content of cysteine ​​​​residues, which form disulfide bridges, which gives high elasticity (the ability to restore its original shape after deformation) to hair, as well as woolen fabrics.

Rice. 24. FRAGMENT OF FIBRILLAR PROTEIN KERATIN

For an irreversible change in the shape of a keratin object, you must first destroy the disulfide bridges with the help of a reducing agent, give it a new shape, and then re-create the disulfide bridges with the help of an oxidizing agent (Fig. 16), this is how, for example, perming hair is done.

With an increase in the content of cysteine ​​residues in keratin and, accordingly, an increase in the number of disulfide bridges, the ability to deform disappears, but high strength appears at the same time (horns of ungulates and turtle shells contain up to 18% of cysteine ​​fragments). Mammals have up to 30 different types of keratin.

The keratin-related fibrillar protein fibroin secreted by silkworm caterpillars during cocoon curling, as well as by spiders during web weaving, contains only β-structures connected by single chains (Fig. 11). Unlike keratin, fibroin does not have transverse disulfide bridges, it has a very strong tensile strength (strength per unit cross-section of some web samples is higher than that of steel cables). Due to the absence of cross-links, fibroin is inelastic (it is known that woolen fabrics are almost indelible, and silk fabrics are easily wrinkled).

regulatory proteins.

Regulatory proteins, more commonly referred to as hormones, are involved in various physiological processes. For example, the hormone insulin (Fig. 25) consists of two α-chains connected by disulfide bridges. Insulin regulates metabolic processes involving glucose, its absence leads to diabetes.

Rice. 25 PROTEIN INSULIN

The pituitary gland of the brain synthesizes a hormone that regulates the growth of the body. There are regulatory proteins that control the biosynthesis of various enzymes in the body.

Contractile and motor proteins give the body the ability to contract, change shape and move, primarily, we are talking about muscles. 40% of the mass of all proteins contained in the muscles is myosin (mys, myos, Greek. - muscle). Its molecule contains both a fibrillar and a globular part (Fig. 26)

Rice. 26 MYOSIN MOLECULE

Such molecules combine into large aggregates containing 300–400 molecules.

When the concentration of calcium ions changes in the space surrounding the muscle fibers, a reversible change in the conformation of the molecules occurs - a change in the shape of the chain due to the rotation of individual fragments around the valence bonds. This leads to muscle contraction and relaxation, the signal to change the concentration of calcium ions comes from the nerve endings in the muscle fibers. Artificial muscle contraction can be caused by the action of electrical impulses, leading to a sharp change in the concentration of calcium ions, this is the basis for stimulating the heart muscle to restore the work of the heart.

Protective proteins allow you to protect the body from the invasion of attacking bacteria, viruses and from the penetration of foreign proteins (the generalized name of foreign bodies is antigens). The role of protective proteins is performed by immunoglobulins (their other name is antibodies), they recognize antigens that have penetrated the body and firmly bind to them. In the body of mammals, including humans, there are five classes of immunoglobulins: M, G, A, D and E, their structure, as the name implies, is globular, in addition, they are all built in a similar way. The molecular organization of antibodies is shown below using class G immunoglobulin as an example (Fig. 27). The molecule contains four polypeptide chains connected by three S-S disulfide bridges (in Fig. 27 they are shown with thickened valence bonds and large S symbols), in addition, each polymer chain contains intrachain disulfide bridges. Two large polymer chains (highlighted in blue) contain 400–600 amino acid residues. The other two chains (highlighted in green) are almost half as long, containing approximately 220 amino acid residues. All four chains are located in such a way that the terminal H 2 N-groups are directed in one direction.

Rice. 27 SCHEMATIC DRAWING OF THE STRUCTURE OF IMMUNOGLOBULIN

After the body comes into contact with a foreign protein (antigen), the cells of the immune system begin to produce immunoglobulins (antibodies), which accumulate in the blood serum. At the first stage, the main work is done by chain sections containing terminal H 2 N (in Fig. 27, the corresponding sections are marked in light blue and light green). These are antigen capture sites. In the process of immunoglobulin synthesis, these sites are formed in such a way that their structure and configuration correspond as much as possible to the structure of the approaching antigen (like a key to a lock, like enzymes, but the tasks in this case are different). Thus, for each antigen, a strictly individual antibody is created as an immune response. Not a single known protein can change its structure so “plastically” depending on external factors, in addition to immunoglobulins. Enzymes solve the problem of structural conformity to the reagent in a different way - with the help of a gigantic set of various enzymes for all possible cases, and immunoglobulins each time rebuild the "working tool". Moreover, the hinge region of the immunoglobulin (Fig. 27) provides the two capture regions with some independent mobility, as a result, the immunoglobulin molecule can immediately “find” the two most convenient regions for capture in the antigen in order to securely fix it, this resembles the actions of a crustacean creature.

Next, a chain of successive reactions of the body's immune system is turned on, immunoglobulins of other classes are connected, as a result, the foreign protein is deactivated, and then the antigen (foreign microorganism or toxin) is destroyed and removed.

After contact with the antigen, the maximum concentration of immunoglobulin is reached (depending on the nature of the antigen and the individual characteristics of the organism itself) within a few hours (sometimes several days). The body retains the memory of such contact, and when attacked again with the same antigen, immunoglobulins accumulate in the blood serum much faster and in greater quantities - acquired immunity occurs.

The above classification of proteins is to a certain extent conditional, for example, the thrombin protein, mentioned among protective proteins, is essentially an enzyme that catalyzes the hydrolysis of peptide bonds, that is, it belongs to the class of proteases.

Protective proteins are often referred to as snake venom proteins and the toxic proteins of some plants, since their task is to protect the body from damage.

There are proteins whose functions are so unique that it makes it difficult to classify them. For example, the protein monellin, found in an African plant, is very sweet tasting and has been the subject of study as a non-toxic substance that can be used in place of sugar to prevent obesity. The blood plasma of some Antarctic fish contains proteins with antifreeze properties that keep the blood of these fish from freezing.

Artificial synthesis of proteins.

The condensation of amino acids leading to a polypeptide chain is a well-studied process. It is possible to carry out, for example, the condensation of any one amino acid or a mixture of acids and obtain, respectively, a polymer containing the same units, or different units, alternating in random order. Such polymers bear little resemblance to natural polypeptides and do not possess biological activity. The main task is to connect amino acids in a strictly defined, pre-planned order in order to reproduce the sequence of amino acid residues in natural proteins. The American scientist Robert Merrifield proposed an original method that made it possible to solve such a problem. The essence of the method is that the first amino acid is attached to an insoluble polymer gel that contains reactive groups that can combine with –COOH – groups of the amino acid. Cross-linked polystyrene with chloromethyl groups introduced into it was taken as such a polymeric substrate. So that the amino acid taken for the reaction does not react with itself and so that it does not join the H 2 N-group to the substrate, the amino group of this acid is pre-blocked with a bulky substituent [(C 4 H 9) 3] 3 OS (O) -group. After the amino acid has attached to the polymeric support, the blocking group is removed and another amino acid is introduced into the reaction mixture, in which the H 2 N group is also previously blocked. In such a system, only the interaction of the H 2 N-group of the first amino acid and the –COOH group of the second acid is possible, which is carried out in the presence of catalysts (phosphonium salts). Then the whole scheme is repeated, introducing the third amino acid (Fig. 28).

Rice. 28. SYNTHESIS SCHEME OF POLYPEPTIDE CHAINS

In the last step, the resulting polypeptide chains are separated from the polystyrene support. Now the whole process is automated, there are automatic peptide synthesizers that operate according to the described scheme. Many peptides used in medicine and agriculture have been synthesized by this method. It was also possible to obtain improved analogues of natural peptides with selective and enhanced action. Some small proteins have been synthesized, such as the hormone insulin and some enzymes.

There are also methods of protein synthesis that replicate natural processes: they synthesize fragments of nucleic acids configured to produce certain proteins, then these fragments are inserted into a living organism (for example, into a bacterium), after which the body begins to produce the desired protein. In this way, significant amounts of hard-to-reach proteins and peptides, as well as their analogues, are now obtained.

Proteins as food sources.

Proteins in a living organism are constantly broken down into their original amino acids (with the indispensable participation of enzymes), some amino acids pass into others, then proteins are synthesized again (also with the participation of enzymes), i.e. the body is constantly renewing itself. Some proteins (collagen of the skin, hair) are not renewed, the body continuously loses them and instead synthesizes new ones. Proteins as food sources perform two main functions: they supply the body with building material for the synthesis of new protein molecules and, in addition, supply the body with energy (sources of calories).

Carnivorous mammals (including humans) get the necessary proteins from plant and animal foods. None of the proteins obtained from food is integrated into the body in an unchanged form. In the digestive tract, all absorbed proteins are broken down to amino acids, and proteins necessary for a particular organism are already built from them, while the remaining 12 can be synthesized from 8 essential acids (Table 1) in the body if they are not supplied in sufficient quantities with food, but essential acids must be supplied with food without fail. Sulfur atoms in cysteine ​​are obtained by the body with the essential amino acid methionine. Part of the proteins breaks down, releasing the energy necessary to maintain life, and the nitrogen contained in them is excreted from the body with urine. Usually the human body loses 25–30 g of protein per day, so protein foods must always be present in the right amount. The minimum daily requirement for protein is 37 g for men and 29 g for women, but the recommended intake is almost twice as high. When evaluating foods, it is important to consider protein quality. In the absence or low content of essential amino acids, the protein is considered of low value, so such proteins should be consumed in greater quantities. So, the proteins of legumes contain little methionine, and the proteins of wheat and corn are low in lysine (both amino acids are essential). Animal proteins (excluding collagens) are classified as complete foods. A complete set of all essential acids contains milk casein, as well as cottage cheese and cheese prepared from it, so a vegetarian diet, if it is very strict, i.e. “dairy-free”, requires increased consumption of legumes, nuts and mushrooms to supply the body with essential amino acids in the right amount.

Synthetic amino acids and proteins are also used as food products, adding them to feed, which contain essential amino acids in small quantities. There are bacteria that can process and assimilate oil hydrocarbons, in this case, for the full synthesis of proteins, they need to be fed with nitrogen-containing compounds (ammonia or nitrates). The protein obtained in this way is used as feed for livestock and poultry. A set of enzymes, carbohydrases, are often added to animal feed, which catalyze the hydrolysis of carbohydrate food components that are difficult to decompose (the cell walls of grain crops), as a result of which plant foods are more fully absorbed.

Mikhail Levitsky

PROTEINS (Article 2)

(proteins), a class of complex nitrogen-containing compounds, the most characteristic and important (along with nucleic acids) components of living matter. Proteins perform many and varied functions. Most proteins are enzymes that catalyze chemical reactions. Many hormones that regulate physiological processes are also proteins. Structural proteins such as collagen and keratin are the main components of bone tissue, hair and nails. The contractile proteins of muscles have the ability to change their length, using chemical energy to perform mechanical work. Proteins are antibodies that bind and neutralize toxic substances. Some proteins that can respond to external influences (light, smell) serve as receptors in the sense organs that perceive irritation. Many proteins located inside the cell and on the cell membrane perform regulatory functions.

In the first half of the 19th century many chemists, and among them primarily J. von Liebig, gradually came to the conclusion that proteins are a special class of nitrogenous compounds. The name "proteins" (from the Greek protos - the first) was proposed in 1840 by the Dutch chemist G. Mulder.

PHYSICAL PROPERTIES

Proteins are white in the solid state, but colorless in solution, unless they carry some chromophore (colored) group, such as hemoglobin. The solubility in water of different proteins varies greatly. It also varies with pH and with the concentration of salts in the solution, so that one can choose the conditions under which one protein will selectively precipitate in the presence of other proteins. This "salting out" method is widely used to isolate and purify proteins. The purified protein often precipitates out of solution as crystals.

In comparison with other compounds, the molecular weight of proteins is very large - from several thousand to many millions of daltons. Therefore, during ultracentrifugation, proteins are precipitated, and, moreover, at different rates. Due to the presence of positively and negatively charged groups in protein molecules, they move at different speeds in an electric field. This is the basis of electrophoresis, a method used to isolate individual proteins from complex mixtures. Purification of proteins is also carried out by chromatography.

CHEMICAL PROPERTIES

Structure.

Proteins are polymers, i.e. molecules built like chains from repeating monomer units, or subunits, whose role is played by alpha-amino acids. General formula of amino acids

where R is a hydrogen atom or some organic group.

A protein molecule (polypeptide chain) may consist of only a relatively small number of amino acids or several thousand monomer units. The connection of amino acids in a chain is possible because each of them has two different chemical groups: an amino group with basic properties, NH2, and an acidic carboxyl group, COOH. Both of these groups are attached to the a carbon atom. The carboxyl group of one amino acid can form an amide (peptide) bond with the amino group of another amino acid:

After two amino acids have been connected in this way, the chain can be extended by adding a third to the second amino acid, and so on. As can be seen from the above equation, when a peptide bond is formed, a water molecule is released. In the presence of acids, alkalis or proteolytic enzymes, the reaction proceeds in the opposite direction: the polypeptide chain is cleaved into amino acids with the addition of water. This reaction is called hydrolysis. Hydrolysis proceeds spontaneously, and energy is required to combine amino acids into a polypeptide chain.

A carboxyl group and an amide group (or an imide group similar to it - in the case of the amino acid proline) are present in all amino acids, while the differences between amino acids are determined by the nature of that group, or "side chain", which is indicated above by the letter R. The role of the side chain can be played by one a hydrogen atom, like the amino acid glycine, and some bulky grouping, like histidine and tryptophan. Some side chains are chemically inert, while others are highly reactive.

Many thousands of different amino acids can be synthesized, and many different amino acids occur in nature, but only 20 types of amino acids are used for protein synthesis: alanine, arginine, asparagine, aspartic acid, valine, histidine, glycine, glutamine, glutamic acid, isoleucine, leucine, lysine , methionine, proline, serine, tyrosine, threonine, tryptophan, phenylalanine and cysteine ​​(in proteins, cysteine ​​may be present as a dimer - cystine). True, in some proteins there are other amino acids in addition to the regularly occurring twenty, but they are formed as a result of modification of any of the twenty listed after it has been included in the protein.

optical activity.

All amino acids, with the exception of glycine, have four different groups attached to the α-carbon atom. In terms of geometry, four different groups can be attached in two ways, and accordingly there are two possible configurations, or two isomers, related to each other as an object to its mirror image, i.e. like left hand to right. One configuration is called left, or left-handed (L), and the other right-handed, or right-handed (D), because two such isomers differ in the direction of rotation of the plane of polarized light. Only L-amino acids occur in proteins (the exception is glycine; it can only be represented in one form, since two of its four groups are the same), and they all have optical activity (since there is only one isomer). D-amino acids are rare in nature; they are found in some antibiotics and the cell wall of bacteria.

The sequence of amino acids.

Amino acids in the polypeptide chain are not arranged randomly, but in a certain fixed order, and it is this order that determines the functions and properties of the protein. By varying the order of the 20 types of amino acids, you can get a huge number of different proteins, just like you can make up many different texts from the letters of the alphabet.

In the past, determining the amino acid sequence of a protein often took several years. Direct determination is still a rather laborious task, although devices have been created that allow it to be carried out automatically. It is usually easier to determine the nucleotide sequence of the corresponding gene and derive the amino acid sequence of the protein from it. To date, the amino acid sequences of many hundreds of proteins have already been determined. The functions of decoded proteins are usually known, and this helps to imagine the possible functions of similar proteins formed, for example, in malignant neoplasms.

Complex proteins.

Proteins consisting of only amino acids are called simple. Often, however, a metal atom or some chemical compound that is not an amino acid is attached to the polypeptide chain. Such proteins are called complex. An example is hemoglobin: it contains iron porphyrin, which gives it its red color and allows it to act as an oxygen carrier.

The names of most complex proteins contain an indication of the nature of the attached groups: sugars are present in glycoproteins, fats in lipoproteins. If the catalytic activity of the enzyme depends on the attached group, then it is called a prosthetic group. Often, some vitamin plays the role of a prosthetic group or is part of it. Vitamin A, for example, attached to one of the proteins of the retina, determines its sensitivity to light.

Tertiary structure.

What is important is not so much the amino acid sequence of the protein (primary structure), but the way it is laid in space. Along the entire length of the polypeptide chain, hydrogen ions form regular hydrogen bonds, which give it the shape of a spiral or layer (secondary structure). From the combination of such helices and layers, a compact form of the next order arises - the tertiary structure of the protein. Around the bonds that hold the monomeric links of the chain, rotations through small angles are possible. Therefore, from a purely geometric point of view, the number of possible configurations for any polypeptide chain is infinitely large. In reality, each protein normally exists in only one configuration, determined by its amino acid sequence. This structure is not rigid, it seems to "breathe" - it oscillates around a certain average configuration. The chain is folded into a configuration in which the free energy (the ability to do work) is minimal, just as a released spring is compressed only to a state corresponding to a minimum of free energy. Often, one part of the chain is rigidly linked to the other by disulfide (–S–S–) bonds between two cysteine ​​residues. This is partly why cysteine ​​among amino acids plays a particularly important role.

The complexity of the structure of proteins is so great that it is not yet possible to calculate the tertiary structure of a protein, even if its amino acid sequence is known. But if it is possible to obtain protein crystals, then its tertiary structure can be determined by X-ray diffraction.

In structural, contractile, and some other proteins, the chains are elongated and several slightly folded chains lying side by side form fibrils; fibrils, in turn, fold into larger formations - fibers. However, most proteins in solution are globular: the chains are coiled in a globule, like yarn in a ball. Free energy with this configuration is minimal, since hydrophobic ("water-repelling") amino acids are hidden inside the globule, and hydrophilic ("water-attracting") amino acids are on its surface.

Many proteins are complexes of several polypeptide chains. This structure is called the quaternary structure of the protein. The hemoglobin molecule, for example, is made up of four subunits, each of which is a globular protein.

Structural proteins due to their linear configuration form fibers in which the tensile strength is very high, while the globular configuration allows proteins to enter into specific interactions with other compounds. On the surface of the globule, with the correct laying of the chains, a certain form of cavity appears, in which reactive chemical groups are located. If this protein is an enzyme, then another, usually smaller, molecule of some substance enters such a cavity, just as a key enters a lock; in this case, the configuration of the electron cloud of the molecule changes under the influence of chemical groups located in the cavity, and this forces it to react in a certain way. In this way, the enzyme catalyzes the reaction. Antibody molecules also have cavities in which various foreign substances bind and are thereby rendered harmless. The "key and lock" model, which explains the interaction of proteins with other compounds, makes it possible to understand the specificity of enzymes and antibodies, i.e. their ability to react only with certain compounds.

Proteins in different types of organisms.

Proteins that perform the same function in different plant and animal species and therefore bear the same name also have a similar configuration. They, however, differ somewhat in their amino acid sequence. As species diverge from a common ancestor, some amino acids in certain positions are replaced by mutations with others. Harmful mutations that cause hereditary diseases are discarded by natural selection, but beneficial or at least neutral ones can be preserved. The closer two biological species are to each other, the less differences are found in their proteins.

Some proteins change relatively quickly, others are quite conservative. The latter include, for example, cytochrome c, a respiratory enzyme found in most living organisms. In humans and chimpanzees, its amino acid sequences are identical, while in cytochrome c of wheat, only 38% of the amino acids turned out to be different. Even when comparing humans and bacteria, the similarity of cytochromes with (the differences here affect 65% of amino acids) can still be seen, although the common ancestor of bacteria and humans lived on Earth about two billion years ago. Nowadays, comparison of amino acid sequences is often used to build a phylogenetic (genealogical) tree that reflects the evolutionary relationships between different organisms.

Denaturation.

The synthesized protein molecule, folding, acquires its own configuration. This configuration, however, can be destroyed by heating, by changing the pH, by the action of organic solvents, and even by simply agitating the solution until bubbles appear on its surface. A protein altered in this way is called denatured; it loses its biological activity and usually becomes insoluble. Well-known examples of denatured protein are boiled eggs or whipped cream. Small proteins, containing only about a hundred amino acids, are able to renature, i.e. reacquire the original configuration. But most of the proteins are simply transformed into a mass of tangled polypeptide chains and do not restore their previous configuration.

One of the main difficulties in isolating active proteins is their extreme sensitivity to denaturation. This property of proteins finds useful application in the preservation of food products: high temperature irreversibly denatures the enzymes of microorganisms, and the microorganisms die.

PROTEIN SYNTHESIS

For protein synthesis, a living organism must have a system of enzymes capable of attaching one amino acid to another. A source of information is also needed that would determine which amino acids should be connected. Since there are thousands of types of proteins in the body, and each of them consists of an average of several hundred amino acids, the information required must be truly enormous. It is stored (similarly to how a record is stored on a magnetic tape) in the nucleic acid molecules that make up genes.

Enzyme activation.

A polypeptide chain synthesized from amino acids is not always a protein in its final form. Many enzymes are first synthesized as inactive precursors and become active only after another enzyme removes a few amino acids at one end of the chain. Some of the digestive enzymes, such as trypsin, are synthesized in this inactive form; these enzymes are activated in the digestive tract as a result of the removal of the terminal fragment of the chain. The hormone insulin, whose molecule in its active form consists of two short chains, is synthesized in the form of a single chain, the so-called. proinsulin. Then the middle part of this chain is removed, and the remaining fragments bind to each other, forming the active hormone molecule. Complex proteins are formed only after a certain chemical group is attached to the protein, and this attachment often also requires an enzyme.

Metabolic circulation.

After feeding an animal with amino acids labeled with radioactive isotopes of carbon, nitrogen or hydrogen, the label is quickly incorporated into its proteins. If labeled amino acids cease to enter the body, then the amount of label in proteins begins to decrease. These experiments show that the resulting proteins are not stored in the body until the end of life. All of them, with a few exceptions, are in a dynamic state, constantly decomposing to amino acids, and then re-synthesized.

Some proteins break down when cells die and are destroyed. This happens all the time, for example, with red blood cells and epithelial cells lining the inner surface of the intestine. In addition, the breakdown and resynthesis of proteins also occur in living cells. Oddly enough, less is known about the breakdown of proteins than about their synthesis. What is clear, however, is that proteolytic enzymes are involved in the breakdown, similar to those that break down proteins into amino acids in the digestive tract.

The half-life of different proteins is different - from several hours to many months. The only exception is collagen molecules. Once formed, they remain stable and are not renewed or replaced. Over time, however, some of their properties, in particular elasticity, change, and since they are not renewed, certain age-related changes are the result of this, for example, the appearance of wrinkles on the skin.

synthetic proteins.

Chemists have long since learned how to polymerize amino acids, but the amino acids are combined in a disorderly manner, so that the products of such polymerization bear little resemblance to natural ones. True, it is possible to combine amino acids in a given order, which makes it possible to obtain some biologically active proteins, in particular insulin. The process is quite complicated, and in this way it is possible to obtain only those proteins whose molecules contain about a hundred amino acids. It is preferable instead to synthesize or isolate the nucleotide sequence of a gene corresponding to the desired amino acid sequence, and then introduce this gene into a bacterium, which will produce by replication a large amount of the desired product. This method, however, also has its drawbacks.

PROTEINS AND NUTRITION

When proteins in the body are broken down into amino acids, these amino acids can be reused for protein synthesis. At the same time, the amino acids themselves are subject to decay, so that they are not fully utilized. It is also clear that during growth, pregnancy, and wound healing, protein synthesis must exceed degradation. The body continuously loses some proteins; these are the proteins of hair, nails and the surface layer of the skin. Therefore, for the synthesis of proteins, each organism must receive amino acids from food.

Sources of amino acids.

Green plants synthesize all 20 amino acids found in proteins from CO2, water and ammonia or nitrates. Many bacteria are also able to synthesize amino acids in the presence of sugar (or some equivalent) and fixed nitrogen, but sugar is ultimately supplied by green plants. In animals, the ability to synthesize amino acids is limited; they obtain amino acids by eating green plants or other animals. In the digestive tract, the absorbed proteins are broken down into amino acids, the latter are absorbed, and the proteins characteristic of the given organism are built from them. None of the absorbed protein is incorporated into body structures as such. The only exception is that in many mammals, part of maternal antibodies can pass intact through the placenta into the fetal circulation, and through maternal milk (especially in ruminants) be transferred to the newborn immediately after birth.

Need for proteins.

It is clear that in order to maintain life, the body must receive a certain amount of protein from food. However, the size of this need depends on a number of factors. The body needs food both as a source of energy (calories) and as a material for building its structures. In the first place is the need for energy. This means that when there are few carbohydrates and fats in the diet, dietary proteins are used not for the synthesis of their own proteins, but as a source of calories. With prolonged fasting, even your own proteins are spent to meet energy needs. If there are enough carbohydrates in the diet, then protein intake can be reduced.

nitrogen balance.

On average approx. 16% of the total protein mass is nitrogen. When the amino acids that make up proteins are broken down, the nitrogen contained in them is excreted from the body in the urine and (to a lesser extent) in the feces in the form of various nitrogenous compounds. Therefore, it is convenient to use such an indicator as nitrogen balance to assess the quality of protein nutrition, i.e. the difference (in grams) between the amount of nitrogen taken into the body and the amount of nitrogen excreted per day. With normal nutrition in an adult, these amounts are equal. In a growing organism, the amount of excreted nitrogen is less than the amount of incoming, i.e. the balance is positive. With a lack of protein in the diet, the balance is negative. If there are enough calories in the diet, but the proteins are completely absent in it, the body saves proteins. At the same time, protein metabolism slows down, and the re-utilization of amino acids in protein synthesis proceeds with the highest possible efficiency. However, losses are inevitable, and nitrogenous compounds are still excreted in the urine and partly in the feces. The amount of nitrogen excreted from the body per day during protein starvation can serve as a measure of the daily lack of protein. It is natural to assume that by introducing into the diet an amount of protein equivalent to this deficiency, it is possible to restore the nitrogen balance. However, it is not. Having received this amount of protein, the body begins to use amino acids less efficiently, so some additional protein is required to restore the nitrogen balance.

If the amount of protein in the diet exceeds what is necessary to maintain nitrogen balance, then there seems to be no harm from this. Excess amino acids are simply used as a source of energy. A particularly striking example is the Eskimo, who consume little carbohydrate and about ten times more protein than is required to maintain nitrogen balance. In most cases, however, using protein as an energy source is not beneficial, since you can get many more calories from a given amount of carbohydrates than from the same amount of protein. In poor countries, the population receives the necessary calories from carbohydrates and consumes a minimum amount of protein.

If the body receives the required number of calories in the form of non-protein products, then the minimum amount of protein that maintains the nitrogen balance is approx. 30 g per day. Approximately as much protein is contained in four slices of bread or 0.5 liters of milk. A slightly larger amount is usually considered optimal; recommended from 50 to 70 g.

Essential amino acids.

Until now, protein has been considered as a whole. Meanwhile, in order for protein synthesis to take place, all the necessary amino acids must be present in the body. Some of the amino acids the body of the animal itself is able to synthesize. They are called interchangeable, since they do not have to be present in the diet - it is only important that, in general, the intake of protein as a source of nitrogen is sufficient; then, with a shortage of non-essential amino acids, the body can synthesize them at the expense of those that are present in excess. The remaining "essential" amino acids cannot be synthesized and must be ingested with food. Essential for humans are valine, leucine, isoleucine, threonine, methionine, phenylalanine, tryptophan, histidine, lysine, and arginine. (Although arginine can be synthesized in the body, it is considered an essential amino acid because newborns and growing children produce insufficient amounts of it. On the other hand, for a person of mature age, the intake of some of these amino acids from food may become optional.)

This list of essential amino acids is approximately the same in other vertebrates and even in insects. The nutritional value of proteins is usually determined by feeding them to growing rats and monitoring the weight gain of the animals.

The nutritional value of proteins.

The nutritional value of a protein is determined by the essential amino acid that is most deficient. Let's illustrate this with an example. The proteins of our body contain an average of approx. 2% tryptophan (by weight). Let's say that the diet includes 10 g of protein containing 1% tryptophan, and that there are enough other essential amino acids in it. In our case, 10 g of this defective protein is essentially equivalent to 5 g of a complete one; the remaining 5 g can only serve as a source of energy. Note that since amino acids are practically not stored in the body, and in order for protein synthesis to take place, all amino acids must be present simultaneously, the effect of the intake of essential amino acids can be detected only if all of them enter the body at the same time.

The average composition of most animal proteins is close to the average composition of the proteins of the human body, so we are unlikely to face amino acid deficiency if our diet is rich in foods such as meat, eggs, milk and cheese. However, there are proteins, such as gelatin (a product of collagen denaturation), which contain very few essential amino acids. Vegetable proteins, although they are better than gelatin in this sense, are also poor in essential amino acids; especially little in them lysine and tryptophan. Nevertheless, a purely vegetarian diet is not at all harmful, unless it consumes a slightly larger amount of vegetable proteins, sufficient to provide the body with essential amino acids. Most protein is found in plants in the seeds, especially in the seeds of wheat and various legumes. Young shoots, such as asparagus, are also rich in protein.

Synthetic proteins in the diet.

By adding small amounts of synthetic essential amino acids or proteins rich in them to incomplete proteins, such as corn proteins, it is possible to significantly increase the nutritional value of the latter, i.e. thereby increasing the amount of protein consumed. Another possibility is to grow bacteria or yeasts on petroleum hydrocarbons with the addition of nitrates or ammonia as a source of nitrogen. The microbial protein obtained in this way can serve as feed for poultry or livestock, or can be directly consumed by humans. The third, widely used, method uses the physiology of ruminants. In ruminants, in the initial section of the stomach, the so-called. In the rumen, there are special forms of bacteria and protozoa that convert defective plant proteins into more complete microbial proteins, and these, in turn, after digestion and absorption, turn into animal proteins. Urea, a cheap synthetic nitrogen-containing compound, can be added to livestock feed. Microorganisms living in the rumen use urea nitrogen to convert carbohydrates (of which there is much more in the feed) into protein. About a third of all nitrogen in livestock feed can come in the form of urea, which in essence means some chemical protein synthesis.

Content:

What is protein and what functions does it perform in the body. What elements are included in its composition and what is the peculiarity of this substance.

Proteins are the main building material in the human body. Taken as a whole, these substances make up one fifth of our body. In nature, a group of subspecies is known - only in the human body there are five million different variants. With its participation, cells are formed, which are considered the main component of the living tissues of the body. What elements make up proteins and what is the peculiarity of the substance?

The subtleties of the composition

Protein molecules in the human body differ in structure and take on certain functions. So, the main contractile protein is myosin, which forms the muscles and guarantees the movement of the body. It ensures the work of the intestines and the movement of blood through the human vessels. An equally important substance in the body is creatine. The function of the substance is to protect the skin from negative effects - radiation, temperature, mechanical and others. Creatine also protects against the entry of microbes from the outside.

Proteins are made up of amino acids. At the same time, the first of them was discovered at the beginning of the 19th century, and the entire amino acid composition has been known to scientists since the 30s of the last century. Interestingly, of the two hundred amino acids that are discovered today, only two dozen form millions of proteins with different structures.

The main difference in the structure is the presence of radicals of different nature. In addition, amino acids are often classified according to electrical charge. Each of the components under consideration has common characteristics - the ability to react with alkalis and acids, solubility in water, and so on. Almost all representatives of the amino acid group are involved in metabolic processes.

Considering the composition of proteins, it is worth highlighting There are two categories of amino acids - non-essential and non-essential. They differ from each other in their ability to be synthesized in the body. The former are produced in the organs, which guarantees at least partial coverage of the current deficit, while the latter come only with food. If the amount of any of the amino acids decreases, then this leads to disturbances, and sometimes to death.

A protein that contains a complete set of amino acids is called "biologically complete". Such substances are part of animal food. Some plant representatives are also considered useful exceptions - for example, beans, peas and soybeans. The main parameter by which the benefits of the product are judged is the biological value. If we consider milk as the basis ( 100% ), then for fish or meat this parameter will be equals 95, for rice - 58 , bread (only rye) - 74 and so on.

The essential amino acids that make up the protein are involved in the synthesis of new cells and enzymes, that is, they cover plastic needs and are used as the main sources of energy. The composition of proteins includes elements that are capable of transformations, that is, the processes of decarboxylation and transamination. The reactions mentioned above involve two groups of amino acids (carboxyl and amine).

The most valuable and useful for the body is egg white, the structure and properties of which are perfectly balanced. That is why the percentage of amino acids in this product is almost always taken as a basis for comparison.

It was mentioned above that proteins are composed of amino acids, and independent representatives play a major role. Here are some of them:

• Histidine- an element that was obtained in 1911. Its function is aimed at normalizing conditioned reflex work. Histidine plays the role of a source for the formation of histamine, a key CNS mediator involved in the transmission of signals to different parts of the body. If the balance of this amino acid falls below the norm, then the production of hemoglobin in the human bone marrow is suppressed.
• Valine- a substance discovered in 1879, but finally deciphered only after 27 years. In case of its shortage, coordination is disturbed, the skin becomes sensitive to external stimuli.
• Tyrosine(1846). Proteins are made up of many amino acids, but this one plays a key role. It is tyrosine that is considered the main precursor of the following compounds - phenol, tyramine, thyroid gland and others.
• Methionine synthesized only by the end of the 20s of the last century. The substance helps in the synthesis of choline, protects the liver from excessive fat formation, and has a lipotropic effect. It has been proven that such elements play a key role in the fight against atherosclerosis and in the regulation of cholesterol levels. The chemical feature of methionine is that it is involved in the production of adrenaline, it interacts with vitamin B.
• cystine- a substance whose structure was established only by 1903. Its functions are aimed at participating in chemical reactions, metabolic processes of methionine. Cystine also reacts with sulfur-containing substances (enzymes).
• tryptophan is an essential amino acid found in proteins. It was successfully synthesized by 1907. The substance is involved in protein metabolism, guarantees an optimal nitrogen balance in the human body. Tryptophan is involved in the production of serum proteins and hemoglobin.
• Leucine- one of the most "early" amino acids, known since the beginning of the XIX century. Its action is aimed at helping the body grow. The lack of an element leads to disruption of the kidneys and thyroid gland.
• Isoleucine- a key element involved in the nitrogen balance. Scientists discovered the amino acid only in 1890.
• Phenylalanine synthesized in the early 90s of the XIX century. The substance is considered the basis for the formation of adrenal and thyroid hormones. Element deficiency is the main cause of hormonal disruptions.
• Lysine received only at the beginning of the 20th century. The lack of a substance leads to the accumulation of calcium in bone tissues, a decrease in the volume of muscles in the body, the development of anemia, and so on.

It is worth highlighting the chemical composition of proteins. This is not surprising, because the substances in question are chemical compounds.

• carbon - 50-55%;
• oxygen - 22-23%;
• nitrogen - 16-17%;
• hydrogen - 6-7%;
• sulfur - 0,4-2,5%.

In addition to those listed above, proteins include the following elements (depending on the type):

• copper;
• iron;
• phosphorus;
• micro and macro substances.

The chemical content of different proteins is different. The only exception is nitrogen, the content of which is always 16-17%. For this reason, the level of substance content is determined precisely by the percentage of nitrogen. The calculation process is as follows. Scientists know that 6.25 grams of protein contains one gram of nitrogen. To determine the protein volume, it is enough to multiply the current amount of nitrogen by 6.25.

The subtleties of the structure

When considering the question of what proteins are made of, it is worth studying the structure of this substance. Allocate:

• primary structure. The basis is the alternation of amino acids in the composition. If at least one element is switched on or "falls out", then a new molecule is formed. Thanks to this feature, the total number of the latter reaches an astronomical figure.
• secondary structure. The peculiarity of the molecules in the composition of the protein is that they are not in a stretched state, but have different (sometimes complex) configurations. Due to this, the vital activity of the cell is simplified. The secondary structure has the form of a spiral formed from uniform turns. At the same time, adjacent turns are distinguished by a close hydrogen bond. In the case of repeated repetition, stability increases.
• Tertiary structure is formed due to the ability of the mentioned spiral to fit into a ball. It is worth knowing that the composition and structure of proteins largely depends on the primary structure. The tertiary base, in turn, guarantees the retention of high-quality bonds between amino acids with different charges.
• Quaternary structure characteristic of some proteins (hemoglobin). The latter forms not one, but several chains that differ in their primary structure.

The secret of protein molecules is in the general pattern. The higher the structural level, the worse the formed chemical bonds are held together. So, secondary, tertiary and quaternary structures are exposed to radiation, high temperatures and other environmental conditions. The result is often a violation of the structure (denaturation). In this case, a simple protein in the event of a change in structure is capable of rapid recovery. If the substance has been subjected to a negative temperature effect or the influence of other factors, then the denaturation process is irreversible, and the substance itself cannot be restored.

Properties

Above, what are proteins, the definition of these elements, structure and other important issues. But the information will be incomplete if you do not highlight the main properties of the substance (physical and chemical).

The molecular weight of a protein is 10 thousand to one million(here much depends on the type). In addition, they are soluble in water.

Separately, it is worth highlighting the common features of a protein with colloidal solutions:

• Swelling ability. The greater the viscosity of the composition, the higher the molecular weight.
• slow diffusion.
• The ability to dialysis, that is, the division of amino acid groups into other elements using semipermeable type membranes. The main difference between the substances under consideration is their inability to pass through membranes.
• Two-factor stability. This means that the protein is hydrophilic in structure. The charge of a substance directly depends on what the protein consists of, the number of amino acids and their properties.
• The size of each particle is 1-100 nm.

Also, proteins have certain similarities with true solutions. The main thing is in the ability to form homogeneous systems. At the same time, the formation process is spontaneous and does not require an additional stabilizer. In addition, protein solutions are thermodynamically stable.

Scientists identify special amorphous properties of the substances in question. This is explained by the presence of an amino group. If the protein is presented in the form of an aqueous solution, then there are equally different mixtures in it - a cationic, bipolar ion, and also an anionic form.

Also The properties of a protein include:

• The ability to act as a buffer, that is, to react similarly to a weak acid or base. So, in the human body there are two types of buffer systems - protein and hemoglobin, involved in the normalization of the level of homeostasis.
• Movement in an electric field. Depending on the number of amino acids in the protein, their mass and charge, the speed of movement of molecules also changes. This function is used for separation by electrophoresis.
• Salting out (reverse precipitation). If you add ammonium ions, alkaline earth metals and alkaline salts to a protein solution, these molecules and ions compete with each other for water. Against this background, the hydration shell is removed, and the proteins cease to be stable. As a result, they fall out. If you add a certain amount of water, then it is possible to restore the hydration shell.
• Sensitivity to external influences. It should be noted that in the case of a negative external influence, proteins are destroyed, which leads to the loss of many chemical and physical properties. In addition, denaturation causes the breakage of the main bonds that stabilize all levels of the protein structure (except for the primary one).

There are many reasons for denaturation.- the negative effect of organic acids, the action of alkalis or heavy metal ions, the negative effect of urea and various reducing agents, leading to the destruction of disulfide-type bridges.

• The presence of color reactions with different chemical elements (depends on the amino acid composition). This property is used in the laboratory when it is required to determine the total amount of protein.

Results

Protein is a key element of the cell, which ensures the normal development and growth of a living organism. But, despite the fact that the substance has been studied by scientists, there are still many discoveries ahead that will allow us to better understand the secret of the human body and its structure. In the meantime, each of us should know where proteins are formed, what are their features and for what purposes they are needed.

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CHAPTER 1. INTRODUCTION

Reports of a revolution in biology have now become rather banal. It is also considered indisputable that these revolutionary changes were associated with the formation of a complex of sciences at the intersection of biology and chemistry, among which molecular biology and bioorganic chemistry occupied and continue to occupy a central position.

“Molecular biology is a science that aims to understand the nature of life phenomena by studying biological objects and systems at a level approaching the molecular one ... characteristic manifestations of life ... are due to the structure, properties and interaction of molecules of biologically important substances, primarily proteins and nucleic acids ”

“Bioorganic chemistry is a science that studies the substances that underlie life processes ... the main objects of bioorganic chemistry are biopolymers (proteins and peptides, nucleic acids and nucleotides, lipids, polysaccharides, etc.).

From this comparison it becomes obvious how important the study of proteins is for the development of modern biology.

biology protein biochemistry

CHAPTER 2. HISTORY OF PROTEIN RESEARCH

2.1 Early stages in protein chemistry

Protein was among the objects of chemical research 250 years ago. In 1728, the Italian scientist Jacopo Bartolomeo Beccari obtained the first protein preparation, gluten, from wheat flour. He subjected gluten to dry distillation and made sure that the products of this distillation were alkaline. This was the first proof of the unity of the nature of the substances of the plant and animal kingdoms. He published the results of his work in 1745, and this was the first paper on a protein.

In the XVIII - early XIX centuries, protein substances of plant and animal origin were repeatedly described. A feature of such descriptions was the convergence of these substances and their comparison with inorganic substances.

It is important to note that at that time, even before the advent of elemental analysis, there was an idea that proteins from various sources were a group of individual substances with similar properties.

In 1810, J. Gay-Lussac and L. Tenard first determined the elemental composition of protein substances. In 1833, J. Gay-Lussac proved that nitrogen is necessarily present in proteins, and soon it was shown that the nitrogen content in different proteins is approximately the same. At the same time, the English chemist D. Dalton tried to depict the first formulas of protein substances. He represented them as rather simply arranged substances, but in order to emphasize their individual difference with the same composition, he resorted to depicting molecules that would now be called isomeric. However, the concept of isomerism did not yet exist in Dalton's time.

Protein formulas by D. Dalton

The first empirical formulas of proteins were derived and the first hypotheses were put forward regarding the regularities of their composition. So, N. Lieberkün believed that albumin is described by the formula C 72 H 112 N 18 SO 22, and A. Danilevsky believed that the molecule of this protein is at least an order of magnitude larger: C 726 H 1171 N 194 S 3 O 214.

The German chemist J. Liebig in 1841 suggested that animal proteins have analogues among vegetable proteins: the assimilation of legumin protein in the animal body, according to Liebig, led to the accumulation of a similar protein - casein. One of the most widespread theories of prestructural organic chemistry was the theory of radicals, the invariable components of related substances. In 1836, the Dutchman G. Mulder suggested that all proteins contain the same radical, which he called protein (from the Greek word “I take the lead”, “I take the first place”). The protein, according to Mulder, had the composition Pr = C 40 H 62 N 10 O 12 . In 1838, G. Mulder published protein formulas based on protein theory. These were the so-called. dualistic formulas, where the protein radical served as a positive group, and sulfur or phosphorus atoms served as a negative group. Together they formed an electrically neutral molecule: blood serum protein Pr 10 S 2 P, fibrin Pr 10 SP. However, an analytical verification of G. Mulder's data, carried out by the Russian chemist Lyaskovskii, as well as Yu. Liebig, showed that "protein radicals" do not exist.

In 1833, the German scientist F. Rose discovered the biuret reaction for proteins - one of the main color reactions for protein substances and their derivatives at the present time (more on color reactions on page 53). It was also concluded that this was the most sensitive reaction for a protein, so it attracted the most attention from chemists at the time.

In the middle of the 19th century, numerous methods were developed for extracting proteins, purifying and isolating them in solutions of neutral salts. In 1847, K. Reichert discovered the ability of proteins to form crystals. In 1836, T. Schwann discovered pepsin, an enzyme that breaks down proteins. In 1856, L. Corvisar discovered another similar enzyme - trypsin. By studying the action of these enzymes on proteins, biochemists tried to unravel the mystery of digestion. However, the substances resulting from the action of protelytic enzymes (proteases, these include the above enzymes) on proteins attracted the most attention: some of them were fragments of the original protein molecules (they were called peptones ), while others were not subjected to further cleavage by proteases and belonged to the class of compounds known since the beginning of the century - amino acids (the first amino acid derivative, asparagine amide, was discovered in 1806, and the first amino acid, cystine, in 1810). Amino acids in the composition of proteins were first discovered in 1820 by the French chemist A. Braconno. He applied the acid hydrolysis of the protein and found a sweetish substance in the hydrolyzate, which he called glycine. In 1839, the existence of leucine in proteins was proven, and in 1849, F. Bopp isolated another amino acid from protein - tyrosine (see Appendix II for a complete list of the dates of discoveries of amino acids in proteins).

By the end of the 80s. In the 19th century, 19 amino acids were already isolated from protein hydrolysates, and the opinion slowly began to grow stronger that information about the products of protein hydrolysis carries important information about the structure of the protein molecule. However, amino acids were considered essential, but not the main component of the protein.

In connection with the discoveries of amino acids in the composition of proteins, the French scientist P. Schutzenberger in the 70s. XIX century proposed the so-called. ureide theory protein structures. According to it, a protein molecule consisted of a central core, the role of which was played by a tyrosine molecule, and complex groups attached to it (with the substitution of 4 hydrogen atoms), called Schutzenberger leucines . However, the hypothesis was very weakly supported experimentally, and further research proved to be inconsistent.

2.2 Theory of “carbon-nitrogen complexes” A.Ya. Danilevsky

The original theory about the structure of the protein was expressed in the 80s. XIX century Russian biochemist A. Ya. Danilevsky. He was the first chemist to draw attention to the possible polymeric nature of the structure of protein molecules. In the early 70s. he wrote to A.M. Butlerov that “albumin particles are a mixed polymeride”, that for the definition of protein he does not find “a term more suitable than the word polymer in the broad sense”. Studying the biuret reaction, he suggested that this reaction is associated with the structure of intermittent carbon and nitrogen atoms - N - C - N - C - N -, which are included in the so-called. carbonazo T complex R "- NH - CO - NH - CO - R". Based on this formula, Danilevsky believed that the protein molecule contains 40 such carbon-nitrogen complexes. Separate carbon-nitrogen amino acid complexes, according to Danilevsky, looked like this:

According to Danilevsky, carbon-nitrogen complexes could be connected by an ether or amide bond to form a high-molecular structure.

2.3 The theory of “kirins” A. Kossel

The German physiologist and biochemist A. Kossel, studying protamines and histones, relatively simple proteins, found that a large amount of arginine is formed during their hydrolysis. In addition, he discovered in the composition of the hydrolyzate the then unknown amino acid - histidine. Based on this, Kossel suggested that these protein substances can be considered as some simple models of more complex proteins, built, in his opinion, according to the following principle: arginine and histidine form a central core (“protamine core”), which is surrounded by complexes of other amino acids.

Kossel's theory was the most perfect example of the development of the hypothesis of the fragmented structure of proteins (first proposed, as mentioned above, by G. Mulder). This hypothesis was used by the German chemist M. Siegfried at the beginning of the 20th century. He believed that proteins are built from complexes of amino acids (arginine + lysine + glutamine acid), which he called kirinami (from the Greek "kyrios" basic). However, this hypothesis was put forward in 1903, when E. Fisher was actively developing his peptide theory , which gave the key to the mystery of the structure of proteins.

2.4 Peptide theory E. Fisher

The German chemist Emil Fischer, already famous throughout the world for his studies of purine compounds (alkaloids of the caffeine group) and deciphering the structure of sugars, created the peptide theory, which was largely confirmed in practice and received universal recognition during his lifetime, for which he was awarded the second Nobel Prize in the history of chemistry. prizes (the first was received by Ya.G. Van't Hoff).

It is important that Fisher built a research plan that differs sharply from what was done before, but takes into account all the facts known at that time. First of all, he accepted as the most likely hypothesis that proteins are built from amino acids connected by an amide bond:

Fisher called this type of bond (by analogy with peptones) peptide . He suggested that proteins are polymers of amino acids linked by peptide bonds . The idea of ​​the polymeric nature of the structure of proteins, as is well known, was expressed by Danilevsky and Hert, but they believed that “monomers” are very complex formations - peptones or “carbon-nitrogen complexes”.

Proving the peptide type of compound of amino acid residues. E. Fisher proceeded from the following observations. First, both during the hydrolysis of proteins and during their enzymatic decomposition, various amino acids were formed. Other compounds were extremely difficult to describe and even more difficult to obtain. In addition, Fischer knew that proteins do not have a predominance of either acidic or basic properties, which means, he argued, amino and carboxyl groups in the composition of amino acids in protein molecules are closed and, as it were, mask each other (the amphotericity of proteins, as they would say now ).

Fisher divided the solution to the problem of protein structure, reducing it to the following provisions:

Qualitative and quantitative determination of the products of complete hydrolysis of proteins.

Establishing the structure of these final products.

Synthesis of amino acid polymers with amide (peptide) type compounds.

Comparison of the compounds thus obtained with natural proteins.

From this plan it can be seen that Fisher used for the first time a new methodological approach - the synthesis of model compounds, as a way of proving by analogy.

2.5 Development of methods for the synthesis of amino acids

In order to proceed to the synthesis of derivatives of amino acids connected by a peptide bond, Fischer did a great deal of work on the study of the structure and synthesis of amino acids.

Before Fischer, the general method for the synthesis of amino acids was A. Strecker's cyanohydrin synthesis:

According to the Strecker reaction, it was possible to synthesize alanine, serine, and some other amino acids, and according to its modification (Zelinsky-Stadnikov reaction), both -amino acids and their N-substituted ones.

However, Fischer himself sought to develop methods for the synthesis of all the then known amino acids. He considered Strecker's method not universal enough. Therefore, E. Fischer had to look for a general method for the synthesis of amino acids, including amino acids with complex side radicals.

He proposed to aminate bromo-substituted in - position carboxylic acids. To obtain bromo derivatives, he used, for example, in the synthesis of leucine, arylated or alkylated malonic acid:

But E. Fisher failed to create an absolutely universal method. More reliable reactions have also been developed. For example, Fisher's student G. Lakes proposed the following modification to obtain serine:

Fisher also proved that proteins are composed of optically active amino acid residues (see p. 11). This forced him to develop a new nomenclature of optically active compounds, methods for the separation and synthesis of optical isomers of amino acids. Fisher also came to the conclusion that proteins contain residues of the L-forms of optically active amino acids, and he proved this by first using the principle of diastereoisomerism. This principle was as follows: an optically active alkaloid (brucine, strychnine, cinchonine, quinidine, quinine) was added to the N-acyl derivative of a racemic amino acid. As a result, two stereoisomeric forms of salts with different solubility were formed. After separation of these diastereoisomers, the alkaloid was recovered and the acyl group was removed by hydrolysis.

Fischer was able to develop a method for the complete determination of amino acids in the products of protein hydrolysis: he converted the hydrochloride esters of amino acids by treatment with concentrated alkali in the cold into free esters, which were not appreciably saponified. Then the mixture of these ethers was subjected to fractional distillation and individual amino acids were isolated from the resulting fractions by fractional crystallization.

The new method of analysis not only finally confirmed that proteins consist of amino acid residues, but made it possible to refine and supplement the list of amino acids found in proteins. But still, quantitative analyzes could not answer the main question: what are the principles of the structure of a protein molecule. And E. Fisher formulated one of the main tasks in the study of the structure and properties of proteins: the development experimental memethods for the synthesis of compounds whose main components would be amino acidsOyou connected by a peptide bond.

Thus, Fisher set a non-trivial task - to synthesize a new class of compounds in order to establish the principles of their structure.

Fisher solved this problem, and chemists received convincing evidence that proteins are polymers of amino acids connected by a peptide bond:

CO - CHR" - NH - CO - CHR"" - NH - CO CHR""" - NH -

This position was supported by biochemical evidence. Along the way, it turned out that proteases do not hydrolyze all bonds between amino acids at the same rate. Their ability to cleave the peptide bond was affected by the optical configuration of the amino acids, the substituents at the nitrogen of the amino group, the length of the peptide chain, and the set of residues included in it.

The main proof of the peptide theory was the synthesis of model peptides and their comparison with the peptones of the protein hydrolyzate. The results showed that peptides identical to those synthesized are isolated from protein hydrolysates.

In the course of these studies, E.Fischer and his student E.Abdergalden developed for the first time a method for determining the amino acid sequence in a protein. Its essence was to establish the nature of the amino acid residue of the polypeptide having a free amino group (N-terminal amino acid). To do this, they proposed blocking the amino terminus in the peptide with a naphthalene-sulfonyl group, which is not cleaved off during hydrolysis. By isolating the amino acid labeled with such a group from the hydrolyzate, it was possible to determine which of the amino acids was N-terminal.

After E. Fisher's research, it became clear that proteins are polypeptides. This was an important achievement, including for protein synthesis tasks: it became clear what exactly needed to be synthesized. Only after these works did the problem of protein synthesis acquire a certain direction and the necessary rigor.

Speaking about Fisher's work as a whole, it should be noted that the approach to research itself was rather typical of the coming 20th century - it operated with a wide range of theoretical positions and methodological techniques; his syntheses looked less and less like an art based on intuition than on exact knowledge, and approached the creation of a series of precise, almost technological devices.

2. 6 Crisis of peptide theory

In connection with the use of new physical and physico-chemical research methods in the early 20s. 20th century there were doubts that the protein molecule is a long polypeptide chain. The hypothesis of the possibility of compact folding of peptide chains was treated with skepticism. All this required a revision of the peptide theory of E. Fisher.

In the 20-30s. The diketopiperazine theory has been widely adopted. According to it, diketopiperase rings, which are formed during the cyclization of two amino acid residues, play a central role in the construction of the protein structure. It was also assumed that these structures constitute the central core of the molecule, to which short peptides or amino acids are attached (“fillers” of the cyclic skeleton of the main structure). The most convincing schemes for the participation of diketopiperazines in the construction of the protein structure were presented by N.D. Zelinsky and E. Fisher's students.

However, attempts to synthesize model compounds containing diketopiperazines did little for protein chemistry; subsequently, the peptide theory triumphed, but these works had a stimulating effect on the chemistry of piperazines in general.

After the peptide and diketopiperase theories, attempts continued to prove the existence of only peptide structures in the protein molecule. At the same time, they tried to imagine not only the type of molecule, but also its general outlines.

The original hypothesis was expressed by the Soviet chemist D.L. Talmud. He suggested that the peptide chains in the composition of protein molecules are folded into large rings, which in turn was a step towards creating his idea of ​​a protein globule.

At the same time, data appeared indicating a different set of amino acids in different proteins. But the patterns that govern the sequence of amino acids in the protein structure were not clear.

M. Bergman and K. Niemann were the first to try to answer this question in their hypothesis of “intermittent frequencies”. According to it, the sequence of amino acid residues in a protein molecule obeyed numerical patterns, the foundations of which were derived from the principles of the structure of the silk fibroin protein molecule. But this choice was unsuccessful, because. this protein is fibrillar, while the structure of globular proteins obeys completely different patterns.

According to M. Bergman and K. Nieman, each amino acid occurs in the polypeptide chain at a certain interval or, as M. Bergman said, has a certain “periodicity.” This periodicity is determined by the nature of amino acid residues.

They imagined the silk fibroin molecule as follows:

GlyAlaGlyTyr GlyAlaGlyArg GlyAlaGlyx GlyAlaGlyx

(GlyAlaGlyTyr GlyAlaGlyx GlyAlaGlyx GlyAlaGlyx) 12

GlyAlaGlyTyr GlyAlaGlyx GlyAlaGlyx GlyAlaGlyArg

(GlyAlaGlyTyr GlyAlaGlyx GlyAlaGlyx GlyAlaGlyx) 13

The Bergman-Niemann hypothesis had a significant impact on the development of amino acid chemistry, a large number of works were devoted to its verification.

In conclusion of this chapter, it should be noted that by the middle of the XX century. enough evidence of the validity of the peptide theory has been accumulated, its main provisions have been supplemented and refined. Therefore, the center for protein research in the 20th century. already lay the field of research and search for methods of protein synthesis by artificial means. This problem was successfully solved, reliable methods were developed for determining the primary structure of a protein - the sequence of amino acids in the peptide chain, methods for the chemical (abiogenic) synthesis of irregular polypeptides were developed (these methods are discussed in more detail in Chapter 8, p. 36), including methods for automatic synthesis of polypeptides. This made it possible already in 1962 for the largest English chemist F. Senger to decipher the structure and artificially synthesize the hormone insulin, which marked a new era in the synthesis of functional protein polypeptides.

CHAPTER 3. CHEMICAL COMPOSITION OF PROTEINS

3.1 Peptide bond

Proteins are irregular polymers built from α-amino acid residues, the general formula of which in an aqueous solution at pH values ​​close to neutral can be written as NH 3 + CHRCOO - . Amino acid residues in proteins are linked together by an amide bond between α-amino and α-carboxyl groups. Peptide bond between two-amino acid residues are commonly referred to as peptide bond , and polymers built from α-amino acid residues connected by peptide bonds are called polypeptides. A protein as a biologically significant structure can be either a single polypeptide or several polypeptides that form a single complex as a result of non-covalent interactions.

3.2 Elemental composition of proteins

Studying the chemical composition of proteins, it is necessary to find out, firstly, what chemical elements they consist of, and secondly, the structure of their monomers. To answer the first question, the quantitative and qualitative composition of the chemical elements of the protein is determined. Chemical analysis showed present in all proteins carbon (50-55%), oxygen (21-23%), nitrogen (15-17%), hydrogen (6-7%), sulfur (0.3-2.5%). Phosphorus, iodine, iron, copper and some other macro- and microelements were also found in the composition of individual proteins, in various, often very small amounts.

The content of the main chemical elements in proteins can vary, with the exception of nitrogen, the concentration of which is characterized by the greatest constancy and averages 16%. In addition, the content of nitrogen in other organic substances is low. In accordance with this, it was proposed to determine the amount of protein by its constituent nitrogen. Knowing that 1 g of nitrogen is contained in 6.25 g of protein, the found amount of nitrogen is multiplied by a factor of 6.25 and the amount of protein is obtained.

To determine the chemical nature of protein monomers, it is necessary to solve two problems: to separate the protein into monomers and to find out their chemical composition. The breakdown of a protein into its constituent parts is achieved by hydrolysis - prolonged boiling of the protein with strong mineral acids. (acid hydrolysis) or grounds (alkaline hydrolysis). Boiling at 110 C with HCl for 24 hours is most commonly used. At the next stage, the substances that make up the hydrolyzate are separated. For this purpose, various methods are used, most often chromatography (for more details, see the chapter “Research methods ...”). Amino acids are the main part of the separated hydrolysates.

3.3. Amino acids

Currently, up to 200 different amino acids have been found in various objects of wildlife. In the human body, for example, there are about 60 of them. However, proteins contain only 20 amino acids, sometimes called natural ones.

Amino acids are organic acids in which the hydrogen atom - carbon atom is replaced by an amino group - NH 2. Therefore, by chemical nature, these are amino acids with the general formula:

From this formula it can be seen that the composition of all amino acids includes the following general groups: - CH 2 - NH 2 - COOH. Side chains (radicals - R) amino acids differ. As can be seen from Appendix I, the chemical nature of radicals is diverse: from a hydrogen atom to cyclic compounds. It is the radicals that determine the structural and functional features of amino acids.

All amino acids, except for the simplest aminoacetic to-you glycine (NH 3 + CH 2 COO) have a chiral atom C and can exist in the form of two enantiomers (optical isomers):

All currently studied proteins contain only amino acids of the L-series, in which, if we consider the chiral atom from the side of the H atom, the NH 3 + , COO groups and the R radical are located clockwise. The need to build a biologically significant polymer molecule from a strictly defined enantiomer is obvious - from a racemic mixture of two enantiomers, an unimaginably complex mixture of diastereoisomers would be obtained. The question why life on Earth is based on proteins built precisely from L-, and not D-amino acids, still remains an intriguing mystery. It should be noted that D-amino acids are fairly widespread in nature and, moreover, are part of biologically significant oligopeptides.

Proteins are built from the twenty basic α-amino acids, but the rest, quite diverse amino acids, are formed from these 20 amino acid residues already in the composition of the protein molecule. Among these transformations, one should first of all note the formation disulfide bridges during the oxidation of two cysteine ​​residues in the composition of already formed peptide chains. As a result, a diaminodicarboxylic acid residue is formed from two cysteine ​​residues cystine (See Appendix I). In this case, cross-linking occurs either within one polypeptide chain or between two different chains. As a small protein that has two polypeptide chains connected by disulfide bridges, as well as crosslinks within one of the polypeptide chains:

An important example of modification of amino acid residues is the conversion of proline residues into residues hydroxyproline :

This transformation occurs, and on a significant scale, during the formation of an important protein component of the connective tissue - collagen .

Another very important type of protein modification is the phosphorylation of hydroxo groups of serine, threonine and tyrosine residues, for example:

Amino acids in an aqueous solution are in an ionized state due to the dissociation of amino and carboxyl groups that make up the radicals. In other words, they are amphoteric compounds and can exist either as acids (proton donors) or as bases (donor acceptors).

All amino acids, depending on the structure, are divided into several groups:

Acyclic. Monoaminomonocarboxylic amino acids have in their composition one amine and one carboxyl group, in an aqueous solution they are neutral. Some of them have common structural features, which allows them to be considered together:

Glycine and alanine. Glycine (glycocol or aminoacetic acid) is optically inactive - it is the only amino acid that does not have enantiomers. Glycine is involved in the formation of nucleic and bile to - t, heme, is necessary for the neutralization of toxic products in the liver. Alanine is used by the body in various carbohydrate and energy metabolism processes. Its isomer - alanine is an integral part of the pantothenic vitamin to - you, coenzyme A (CoA), extractive substances of muscles.

Serine and threonine. They belong to the group of hydrohydroxy acids, because. have a hydroxyl group. Serine is a part of various enzymes, the main protein of milk - casein, and also a part of many lipoproteins. Threonine is involved in protein biosynthesis, being an essential amino acid.

cysteine ​​and methionine. Amino acids containing a sulfur atom. The value of cysteine ​​is determined by the presence of a sulfhydryl (-SH) group in its composition, which gives it the ability to easily oxidize and protect the body from substances with a high oxidizing ability (in case of radiation injury, phosphorus poisoning). Methionine is characterized by the presence of an easily mobile methyl group, which is used for the synthesis of important compounds in the body (choline, creatine, thymine, adrenaline, etc.)

Valine, leucine and isoleucine. They are branched amino acids that are actively involved in metabolism and are not synthesized in the body.

Monoaminodicarboxylic amino acids have one amino and two carboxyl groups and give an acidic reaction in aqueous solution. These include aspartic and glutamine to-you, asparagine and glutamine. They are part of the inhibitory mediators of the nervous system.

Diaminomonocarboxylic amino acids in aqueous solution have an alkaline reaction due to the presence of two amine groups. Related to them, lysine is necessary for the synthesis of histones and also in a number of enzymes. Arginine is involved in the synthesis of urea, creatine.

Cyclic. These amino acids have an aromatic or heterocyclic nucleus in their composition and, as a rule, are not synthesized in the human body and must be supplied with food. They are actively involved in a variety of metabolic processes. So phenyl-alanine serves as the main source for the synthesis of tyrosine - the precursor of a number of biologically important substances: hormones (thyroxine, adrenaline), some pigments. Tryptophan, in addition to participating in protein synthesis, is a component of vitamin PP, serotonin, tryptamine, and a number of pigments. Histidine is necessary for the synthesis of proteins, is a precursor of histamine, which affects blood pressure and secretion of gastric juice.

CHAPTER 4. STRUCTURE

When studying the composition of proteins, it was found that they are all built according to a single principle and have four levels of organization: primary, secondary, tertiary, and some of them Quaternary structures.

4.1 Primary structure

It is a linear chain of amino acids arranged in a certain sequence and interconnected by peptide bonds. Peptide bond formed by the -carboxyl group of one amino acid and the -amine group of another:

The peptide bond due to the p, -conjugation -bond of the carbonyl group and the p-orbital of the N atom, on which the unshared pair of electrons is located, cannot be considered as a single one and there is practically no rotation around it. For the same reason, the chiral atom C and carbonyl atom Ck of any i-th amino acid residue of the peptide chain and the N and C atoms of the (i+1)-th residue are in the same plane. The carbonyl O atom and the amide H atom are located in the same plane (however, the material accumulated in the study of the structure of proteins shows that this statement is not entirely rigorous: the atoms associated with the peptide nitrogen atom are not in the same plane with it, but form a trihedral pyramid with angles between bonds very close to 120. Therefore, between the planes formed by the atoms C i , C i k , O i and N i +1 , H i +1 , C i +1 , there is some angle that differs from 0. But, as as a rule, it does not exceed 1 and does not play a special role). Therefore, geometrically, the polypeptide chain can be considered as formed by such flat fragments containing six atoms each. The mutual arrangement of these fragments, like any mutual arrangement of two planes, must be determined by two angles. As such, it is customary to take torsion angles that characterize rotations around N C and C C k -bonds.

The geometry of any molecule is determined by three groups of geometric characteristics of its chemical bonds - bond lengths, bond angles and torsion angles between bonds adjacent to neighboring atoms. The first two groups are determined to a decisive extent by the nature of the participating atoms and the bonds formed. Therefore, the spatial structure of polymers is mainly determined by the torsion angles between the links of the polymer backbone of the molecules, i.e. polymer chain conformation. That R sion angle , i.e. angle of rotation of the A-B connection around the B-C connection relative to the C- connectionD, is defined as the angle between the planes containing atoms A, B, C and atomsB, C, D.

In such a system, it is possible that the A-B and C-D bonds are located in parallel and are on the same side of the B-C bond. If we consider this system along theIzi B-C, then the connection A-B, as it were, obscures the connectionC- D, so this conformation is calledsvaetsyaobscured. According to the recommendations of the international unions of chemistry IUPAC (International Union of Pure and Applied Chemistry) and IUB (International Union of Biochemistry), the angle between the ABC and BCD planes is considered positive if, in order to bring the conformation into a eclipsed state by turning through an angle of no more than 180, closest to the observer connection must be rotated clockwise. If this bond must be rotated counterclockwise to obtain a eclipsed conformation, then the angle is considered negative. It can be seen that this definition does not depend on which of the bonds is closer to the observer.

In this case, as can be seen from the figure, the orientation of the fragment containing the atoms C i -1 and C i [(i-1)-th fragment], and the fragment containing the atoms C i and C i +1 (i-th fragment), is determined by the torsion angles corresponding to the rotation around the bond N i C i and the bond C i C i k . These angles are usually denoted as and, in the given case, respectively i and i. Their values ​​for all monomer units of the polypeptide chain mainly determine the geometry of this chain. There are no unambiguous values ​​either for the value of each of these angles or for their combinations, although restrictions are imposed on both of them, determined both by the properties of the peptide fragments themselves and by the nature of side radicals, i.e. the nature of the amino acid residues.

To date, amino acid sequences have been established for several thousand different proteins. Recording the structure of proteins in the form of detailed structural formulas is cumbersome and not visual. Therefore, an abbreviated form of writing is used - three-letter or one-letter (vasopressin molecule):

When writing an amino acid sequence in polypeptide or oligopeptide chains using abbreviated symbols, it is assumed, unless otherwise noted, that the α-amino group is on the left and the α-carboxyl group is on the right. The corresponding sections of the polypeptide chain are called the N-terminus (amine end) and the C-terminus (carboxyl end), and the amino acid residues are called the N-terminal and C-terminal residues, respectively.

4.2 Secondary structure

Fragments of the spatial structure of a biopolymer having a periodic structure of the polymer backbone are considered as elements of the secondary structure.

If over a certain section of the chain the angles of the same type, which were mentioned on page 15, are approximately the same, then the structure of the polypeptide chain acquires a periodic character. There are two classes of such structures - spiral and stretched (flat or folded).

Spiral a structure is considered in which all atoms of the same type lie on the same helix. In this case, the spiral is considered right if, when observed along the axis of the spiral, it moves away from the observer in a clockwise direction, and left - if it moves away counterclockwise. The polypeptide chain has a helical conformation if all C atoms are on one helix, all carbonyl atoms C k - on the other, all N atoms - on the third, and the helix pitch for all three groups of atoms should be the same. The number of atoms per one turn of the helix should also be the same, regardless of whether we are talking about atoms C k , C or N. The distance to the common helix for each of these three types of atoms is different.

The main elements of the secondary structure of proteins are -helices and -folds.

Helical protein structures. Several different types of helices are known for polypeptide chains. Among them, the right-handed helix is ​​the most common. The ideal -helix has a pitch of 0.54 nm and the number of atoms of the same type per turn of the helix is ​​3.6, which means a complete periodicity on five turns of the helix every 18 amino acid residues. The values ​​of torsion angles for an ideal α-helix = - 57 = - 47 , and the distances from the atoms forming the polypeptide chain to the axis of the helix are 0.15 nm for N, 0.23 nm for C, and 0.17 nm for C k . Any conformation exists provided that there are factors stabilizing it. In the case of a helix, such factors are the hydrogen bonds formed by each carbonyl atom of the (i + 4)th fragment. An important factor in the stabilization of the α-helix is ​​also the parallel orientation of the dipole moments of peptide bonds.

Folded protein structures. One of the common examples of the folded periodic structure of a protein is the so-called. -folds, consisting of two fragments, each of which is represented by a polypeptide.

Folds are also stabilized by hydrogen bonds between the hydrogen atom of the amine group of one fragment and the oxygen atom of the carboxyl group of another fragment. In this case, the fragments can have both parallel and antiparallel orientation relative to each other.

The structure resulting from such interactions is a corrugated structure. This affects the values ​​of torsion angles and. If in a flat, fully stretched structure they should be 180, then in real β-layers they have the values ​​= - 119 and = + 113. a section that has a structure that differs sharply from a periodic one.

4.2.1 Factors affecting secondary structure formation

The structure of a certain section of the polypeptide chain essentially depends on the structure of the molecule as a whole. The factors influencing the formation of areas with a certain secondary structure are very diverse and by no means have been fully identified in all cases. It is known that a number of amino acid residues preferentially occur in α-helical fragments, a number of others - in α-folds, some amino acids - mainly in regions devoid of a periodic structure. The secondary structure is largely determined by the primary structure. In some cases, the physical meaning of such a dependence can be understood from a stereochemical analysis of the spatial structure. For example, as can be seen from the figure, not only side radicals of amino acid residues adjacent along the chain are brought together in the -helix, but also some pairs of residues located on adjacent turns of the helix, first of all, each (i + 1)th residue with (i + 4) -th and with (i+5)-th. Therefore, in positions (i + 1) and (i + 2), (i + 1) and (i + 4), (i + 1) and (i + 5) -helices, two bulky radicals rarely occur simultaneously, such as, for example , as side radicals of tyrosine, tryptophan, isoleucine. Even less compatible with the helix structure is the simultaneous presence of three bulk residues in positions (i+1), (i+2) and (i+5) or (i+1), (i+4) and (i+5). Therefore, such combinations of amino acids in α-helical fragments are rare exceptions.

4.3 Tertiary structure

This term refers to the complete folding in space of the entire polypeptide chain, including the folding of side radicals. A complete picture of the tertiary structure is given by the coordinates of all atoms of the protein. Thanks to the enormous success of X-ray diffraction analysis, such data, with the exception of the coordinates of hydrogen atoms, have been obtained for a significant number of proteins. These are huge amounts of information stored in special data banks on machine-readable media, and their processing is unthinkable without the use of high-speed computers. Atomic coordinates obtained on computers provide complete information about the geometry of the polypeptide chain, including the values ​​of torsion angles, which makes it possible to reveal a helical structure, folds, or irregular fragments. An example of such a research approach is the following spatial model of the structure of the phosphoglycerate kinase enzyme:

General scheme of the structure of phosphoglycerate kinase. For clarity, the α-helical sections are presented as cylinders, and the α-folds are presented as ribbons with an arrow indicating the direction of the chain from the N-terminus to the C-terminus. Lines are irregular sections connecting structured fragments.

The image of the complete structure of even a small protein molecule on a plane, whether it is a page of a book or a display screen, is not very informative due to the extremely complex structure of the object. In order for the researcher to be able to visualize the spatial structure of the molecules of complex substances, they use the methods of three-dimensional computer graphics, which allow displaying individual parts of the molecules and manipulating them, in particular, turning them in the right angles.

The tertiary structure is formed as a result of non-covalent interactions (electrostatic, ionic, van der Waals forces, etc.) of side radicals framing α-helices and folds and non-periodic fragments of the polypeptide chain. Among the bonds holding the tertiary structure, it should be noted:

a) disulfide bridge (- S - S -)

b) ester bridge (between carboxyl group and hydroxyl group)

c) salt bridge (between carboxyl group and amino group)

d) hydrogen bonds.

In accordance with the shape of the protein molecule due to the tertiary structure, the following groups of proteins are distinguished:

globular proteins. The spatial structure of these proteins in a rough approximation can be represented as a ball or a not too elongated ellipsoid - globatly. As a rule, a significant part of the polypeptide chain of such proteins forms β-helices and β-folds. The ratio between them can be very different. For example, at myoglobin(more about it on page 28) there are 5 helical segments and not a single fold. In immunoglobulins (more details on p. 42), on the contrary, the main elements of the secondary structure are -folds, and -helices are absent altogether. In the above structure of phosphoglycerate kinase, both types of structures are represented approximately the same. In some cases, as can be seen in the example of phosphoglycerate kinase, two or more clearly separated in space (but nevertheless, of course, connected by peptide bridges) parts are clearly visible - domains. Often, different functional regions of a protein are separated into different domains.

fibrillar proteins. These proteins have an elongated filamentous shape; they perform a structural function in the body. In the primary structure, they have repeating sections and form a fairly uniform secondary structure for the entire polypeptide chain. Thus, the protein - creatine (the main protein component of nails, hair, skin) is built from extended - spirals. Silk fibroin consists of periodically repeating fragments Gly - Ala - Gly - Ser, forming folds. There are less common elements of the secondary structure, for example, collagen polypeptide chains that form left spirals with parameters sharply different from those of -helices. In collagen fibers, three helical polypeptide chains are twisted into a single right supercoil:

4.4 Quaternary structure

In most cases, for the functioning of proteins, it is necessary that several polymer chains be combined into a single complex. Such a complex is also considered as a protein consisting of several subunits. The subunit structure often appears in the scientific literature as a quaternary structure.

Proteins consisting of several subunits are widely distributed in nature. A classic example is the quaternary structure of hemoglobin (more details - p. 26). subunits are usually denoted by Greek letters. Hemoglobin has two and two subunits. The presence of several subunits is functionally important - it increases the degree of oxygen saturation. The quaternary structure of hemoglobin is designated as 2 2 .

The subunit structure is characteristic of many enzymes, primarily those that perform complex functions. For example, RNA polymerase from E. coli has a subunit structure 2 ", i.e. it is built from four different types of subunits, and the -subunit is duplicated. This protein performs complex and diverse functions - initiates DNA, binds substrates - ribonucleoside triphosphates, and also transfers nucleotide residues to a growing polyribonucleotide chain and some other functions .

The work of many proteins is subject to the so-called. allosteric regulation- special compounds (effectors) “switch off” or “switch on” the work of the active center of the enzyme. Such enzymes have special effector recognition sites. And there are even special regulatory subunits, which include, among other things, the indicated sections. A classic example is protein kinase enzymes that catalyze the transfer of a phosphoric acid residue from an ATP molecule to substrate proteins.

CHAPTER 5. PROPERTIES

Proteins have a high molecular weight, some are soluble in water, capable of swelling, are characterized by optical activity, mobility in an electric field, and some other properties.

Proteins are actively involved in chemical reactions. This property is due to the fact that the amino acids that make up proteins contain different functional groups that can react with other substances. It is important that such interactions also occur inside the protein molecule, resulting in the formation of peptide, hydrogen disulfide, and other types of bonds. Various compounds and ions can attach to the radicals of amino acids, and hence proteins, which ensures their transport through the blood.

Proteins are macromolecular compounds. These are polymers consisting of hundreds and thousands of amino acid residues - monomers. Accordingly and molecular mass proteins is in the range of 10,000 - 1,000,000. So, ribonuclease (an enzyme that breaks down RNA) contains 124 amino acid residues and its molecular weight is approximately 14,000. Myoglobin (muscle protein), consisting of 153 amino acid residues, has a molecular weight 17,000, and hemoglobin - 64,500 (574 amino acid residues). The molecular weights of other proteins are higher: -globulin (forms antibodies) consists of 1250 amino acids and has a molecular weight of about 150,000, and the molecular weight of the glutamate dehydrogenase enzyme exceeds 1,000,000.

The determination of the molecular weight is carried out by various methods: osmometric, gel filtration, optical, etc. however, the most accurate is the sedimentation method proposed by T. Svedberg. It is based on the fact that during ultracentrifugation with an acceleration of up to 900,000 g, the rate of protein precipitation depends on their molecular weight.

The most important property of proteins is their ability to show both acidic and basic, that is, to act as amphoteric electrolytes. This is ensured by various dissociating groups that make up the amino acid radicals. For example, acidic properties are imparted to a protein by carboxyl groups of aspartic glutamic amino acids, while alkaline properties are imparted by arginine, lysine, and histidine radicals. The more dicarboxylic amino acids a protein contains, the stronger its acidic properties are and vice versa.

These groups also have electric charges that form the overall charge of the protein molecule. In proteins where aspartic and glutamine amino acids predominate, the charge of the protein will be negative; an excess of basic amino acids gives a positive charge to the protein molecule. As a result, in an electric field, proteins will move towards the cathode or anode, depending on the magnitude of their total charge. So, in an alkaline environment (pH 7 - 14), the protein donates a proton and becomes negatively charged, while in an acidic environment (pH 1 - 7), the dissociation of acid groups is suppressed and the protein becomes a cation.

Thus, the factor that determines the behavior of a protein as a cation or anion is the reaction of the medium, which is determined by the concentration of hydrogen ions and is expressed by the pH value. However, at certain pH values, the number of positive and negative charges equalizes and the molecule becomes electrically neutral, i.e. it will not move in an electric field. This pH value of the medium is defined as the isoelectric point of proteins. In this case, the protein is in the least stable state and, with slight changes in pH to the acidic or alkaline side, it easily precipitates. For most natural proteins, the isoelectric point is in a slightly acidic environment (pH 4.8 - 5.4), which indicates the predominance of dicarboxylic amino acids in their composition.

The amphoteric property underlies the buffering properties of proteins and their participation in the regulation of blood pH. The pH value of human blood is constant and is in the range of 7.36 - 7.4, despite various substances of an acidic or basic nature, regularly supplied with food or formed in metabolic processes - therefore, there are special mechanisms for regulating the acid-base balance of the internal environment of the body. Such systems include the one considered in Chap. “Classification” hemoglobin buffer system (page 28). A change in blood pH by more than 0.07 indicates the development of a pathological process. A shift in pH to the acid side is called acidosis, and to the alkaline side is called alkalosis.

Of great importance for the body is the ability of proteins to adsorb on their surface certain substances and ions (hormones, vitamins, iron, copper), which are either poorly soluble in water or are toxic (bilirubin, free fatty acids). Proteins transport them through the blood to places of further transformations or neutralization.

Aqueous solutions of proteins have their own characteristics. First, proteins have a high affinity for water, i.e. They hydrophilic. This means that protein molecules, like charged particles, attract water dipoles, which are located around the protein molecule and form a water or hydrate shell. This shell protects the protein molecules from sticking together and precipitating. The size of the hydration shell depends on the structure of the protein. For example, albumins more easily bind to water molecules and have a relatively large water shell, while globulins, fibrinogen attach water worse, and the hydration shell is smaller. Thus, the stability of an aqueous solution of a protein is determined by two factors: the presence of a charge on the protein molecule and the water shell around it. When these factors are removed, the protein precipitates. This process can be reversible and irreversible.

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Squirrels- high-molecular organic compounds, consisting of residues of α-amino acids.

IN protein composition includes carbon, hydrogen, nitrogen, oxygen, sulfur. Some proteins form complexes with other molecules containing phosphorus, iron, zinc and copper.

Proteins have a large molecular weight: egg albumin - 36,000, hemoglobin - 152,000, myosin - 500,000. For comparison: the molecular weight of alcohol is 46, acetic acid - 60, benzene - 78.

Amino acid composition of proteins

Squirrels- non-periodic polymers, the monomers of which are α-amino acids. Usually, 20 types of α-amino acids are called protein monomers, although more than 170 of them have been found in cells and tissues.

Depending on whether amino acids can be synthesized in the body of humans and other animals, there are: non-essential amino acids- can be synthesized essential amino acids- cannot be synthesized. Essential amino acids must be ingested with food. Plants synthesize all kinds of amino acids.

Depending on the amino acid composition, proteins are: complete- contain the entire set of amino acids; defective- some amino acids are absent in their composition. If proteins are made up of only amino acids, they are called simple. If proteins contain, in addition to amino acids, also a non-amino acid component (a prosthetic group), they are called complex. The prosthetic group can be represented by metals (metalloproteins), carbohydrates (glycoproteins), lipids (lipoproteins), nucleic acids (nucleoproteins).

All amino acids contain: 1) a carboxyl group (-COOH), 2) an amino group (-NH 2), 3) a radical or R-group (the rest of the molecule). The structure of the radical in different types of amino acids is different. Depending on the number of amino groups and carboxyl groups that make up amino acids, there are: neutral amino acids having one carboxyl group and one amino group; basic amino acids having more than one amino group; acidic amino acids having more than one carboxyl group.

Amino acids are amphoteric compounds, since in solution they can act as both acids and bases. In aqueous solutions, amino acids exist in different ionic forms.

Peptide bond

Peptides- organic substances consisting of amino acid residues connected by a peptide bond.

The formation of peptides occurs as a result of the condensation reaction of amino acids. When the amino group of one amino acid interacts with the carboxyl group of another, a covalent nitrogen-carbon bond arises between them, which is called peptide. Depending on the number of amino acid residues that make up the peptide, there are dipeptides, tripeptides, tetrapeptides etc. The formation of a peptide bond can be repeated many times. This leads to the formation polypeptides. At one end of the peptide there is a free amino group (it is called the N-terminus), and at the other end there is a free carboxyl group (it is called the C-terminus).

Spatial organization of protein molecules

The performance of certain specific functions by proteins depends on the spatial configuration of their molecules, in addition, it is energetically unfavorable for the cell to keep proteins in an expanded form, in the form of a chain, therefore, polypeptide chains are folded, acquiring a certain three-dimensional structure, or conformation. Allocate 4 levels spatial organization of proteins.

Primary structure of a protein- the sequence of amino acid residues in the polypeptide chain that makes up the protein molecule. The bond between amino acids is peptide.

If a protein molecule consists of only 10 amino acid residues, then the number of theoretically possible variants of protein molecules that differ in the order of alternation of amino acids is 10 20 . With 20 amino acids, you can make even more diverse combinations of them. About ten thousand different proteins have been found in the human body, which differ both from each other and from the proteins of other organisms.

It is the primary structure of the protein molecule that determines the properties of the protein molecules and its spatial configuration. The replacement of just one amino acid for another in the polypeptide chain leads to a change in the properties and functions of the protein. For example, the replacement of the sixth glutamine amino acid in the β-subunit of hemoglobin with valine leads to the fact that the hemoglobin molecule as a whole cannot perform its main function - oxygen transport; in such cases, a person develops a disease - sickle cell anemia.

secondary structure- ordered folding of the polypeptide chain into a spiral (looks like a stretched spring). The coils of the helix are strengthened by hydrogen bonds between carboxyl groups and amino groups. Almost all CO and NH groups take part in the formation of hydrogen bonds. They are weaker than peptide ones, but, repeating many times, they impart stability and rigidity to this configuration. At the level of the secondary structure, there are proteins: fibroin (silk, web), keratin (hair, nails), collagen (tendons).

Tertiary structure- packing of polypeptide chains into globules, resulting from the occurrence of chemical bonds (hydrogen, ionic, disulfide) and the establishment of hydrophobic interactions between radicals of amino acid residues. The main role in the formation of the tertiary structure is played by hydrophilic-hydrophobic interactions. In aqueous solutions, hydrophobic radicals tend to hide from water, grouping inside the globule, while hydrophilic radicals tend to appear on the surface of the molecule as a result of hydration (interaction with water dipoles). In some proteins, the tertiary structure is stabilized by disulfide covalent bonds that form between the sulfur atoms of the two cysteine ​​residues. At the level of the tertiary structure, there are enzymes, antibodies, some hormones.

Quaternary structure characteristic of complex proteins, the molecules of which are formed by two or more globules. Subunits are held in the molecule by ionic, hydrophobic, and electrostatic interactions. Sometimes, during the formation of a quaternary structure, disulfide bonds occur between subunits. The most studied protein with a quaternary structure is hemoglobin. It is formed by two α-subunits (141 amino acid residues) and two β-subunits (146 amino acid residues). Each subunit is associated with a heme molecule containing iron.

If for some reason the spatial conformation of proteins deviates from normal, the protein cannot perform its functions. For example, the cause of "mad cow disease" (spongiform encephalopathy) is an abnormal conformation of prions, the surface proteins of nerve cells.

Protein properties

The amino acid composition, the structure of the protein molecule determine its properties. Proteins combine basic and acidic properties determined by amino acid radicals: the more acidic amino acids in a protein, the more pronounced its acidic properties. The ability to give and attach H + determine buffer properties of proteins; one of the most powerful buffers is hemoglobin in erythrocytes, which maintains the pH of the blood at a constant level. There are soluble proteins (fibrinogen), there are insoluble proteins that perform mechanical functions (fibroin, keratin, collagen). There are chemically active proteins (enzymes), there are chemically inactive, resistant to various environmental conditions and extremely unstable.

External factors (heat, ultraviolet radiation, heavy metals and their salts, pH changes, radiation, dehydration)

can cause a violation of the structural organization of the protein molecule. The process of losing the three-dimensional conformation inherent in a given protein molecule is called denaturation. The cause of denaturation is the breaking of bonds that stabilize a particular protein structure. Initially, the weakest ties are torn, and when conditions become tougher, even stronger ones. Therefore, first the quaternary, then the tertiary and secondary structures are lost. A change in the spatial configuration leads to a change in the properties of the protein and, as a result, makes it impossible for the protein to perform its biological functions. If denaturation is not accompanied by the destruction of the primary structure, then it can be reversible, in this case, self-healing of the conformation characteristic of the protein occurs. Such denaturation is subjected, for example, to membrane receptor proteins. The process of restoring the structure of a protein after denaturation is called renaturation. If the restoration of the spatial configuration of the protein is impossible, then denaturation is called irreversible.

Functions of proteins

Function	Examples and explanations
Construction	Proteins are involved in the formation of cellular and extracellular structures: they are part of cell membranes (lipoproteins, glycoproteins), hair (keratin), tendons (collagen), etc.
Transport	The blood protein hemoglobin attaches oxygen and transports it from the lungs to all tissues and organs, and from them carbon dioxide transfers to the lungs; The composition of cell membranes includes special proteins that provide an active and strictly selective transfer of certain substances and ions from the cell to the external environment and vice versa.
Regulatory	Protein hormones are involved in the regulation of metabolic processes. For example, the hormone insulin regulates blood glucose levels, promotes glycogen synthesis, and increases the formation of fats from carbohydrates.
Protective	In response to the penetration of foreign proteins or microorganisms (antigens) into the body, special proteins are formed - antibodies that can bind and neutralize them. Fibrin, formed from fibrinogen, helps to stop bleeding.
Motor	The contractile proteins actin and myosin provide muscle contraction in multicellular animals.
Signal	Molecules of proteins are embedded in the surface membrane of the cell, capable of changing their tertiary structure in response to the action of environmental factors, thus receiving signals from the external environment and transmitting commands to the cell.
Reserve	In the body of animals, proteins, as a rule, are not stored, with the exception of egg albumin, milk casein. But thanks to proteins in the body, some substances can be stored in reserve, for example, during the breakdown of hemoglobin, iron is not excreted from the body, but is stored, forming a complex with the ferritin protein.
Energy	With the breakdown of 1 g of protein to the final products, 17.6 kJ is released. First, proteins break down into amino acids, and then to the end products - water, carbon dioxide and ammonia. However, proteins are used as an energy source only when other sources (carbohydrates and fats) are used up.
catalytic	One of the most important functions of proteins. Provided with proteins - enzymes that accelerate the biochemical reactions that occur in cells. For example, ribulose bisphosphate carboxylase catalyses CO2 fixation during photosynthesis.

Enzymes

Enzymes, or enzymes, is a special class of proteins that are biological catalysts. Thanks to enzymes, biochemical reactions proceed at a tremendous speed. The rate of enzymatic reactions is tens of thousands of times (and sometimes millions) higher than the rate of reactions involving inorganic catalysts. The substance on which an enzyme acts is called substrate.

Enzymes are globular proteins structural features Enzymes can be divided into two groups: simple and complex. simple enzymes are simple proteins, i.e. consist only of amino acids. Complex enzymes are complex proteins, i.e. in addition to the protein part, they include a group of non-protein nature - cofactor. For some enzymes, vitamins act as cofactors. In the enzyme molecule, a special part is isolated, called the active center. active center- a small section of the enzyme (from three to twelve amino acid residues), where the binding of the substrate or substrates occurs with the formation of an enzyme-substrate complex. Upon completion of the reaction, the enzyme-substrate complex decomposes into the enzyme and reaction product(s). Some enzymes have (other than active) allosteric centers- sites to which regulators of the rate of enzyme work are attached ( allosteric enzymes).

Enzymatic catalysis reactions are characterized by: 1) high efficiency, 2) strict selectivity and direction of action, 3) substrate specificity, 4) fine and precise regulation. The substrate and reaction specificity of enzymatic catalysis reactions is explained by the hypotheses of E. Fischer (1890) and D. Koshland (1959).

E. Fisher (key-lock hypothesis) suggested that the spatial configurations of the active site of the enzyme and the substrate should correspond exactly to each other. The substrate is compared to the "key", the enzyme - to the "lock".

D. Koshland (hypothesis "hand-glove") suggested that the spatial correspondence between the structure of the substrate and the active center of the enzyme is created only at the moment of their interaction with each other. This hypothesis is also called induced fit hypothesis.

The rate of enzymatic reactions depends on: 1) temperature, 2) enzyme concentration, 3) substrate concentration, 4) pH. It should be emphasized that since enzymes are proteins, their activity is highest under physiologically normal conditions.

Most enzymes can only work at temperatures between 0 and 40°C. Within these limits, the reaction rate increases by about 2 times for every 10 °C rise in temperature. At temperatures above 40 °C, the protein undergoes denaturation and the activity of the enzyme decreases. At temperatures close to freezing, the enzymes are inactivated.

With an increase in the amount of substrate, the rate of the enzymatic reaction increases until the number of substrate molecules becomes equal to the number of enzyme molecules. With a further increase in the amount of substrate, the rate will not increase, since the active sites of the enzyme are saturated. An increase in the enzyme concentration leads to an increase in catalytic activity, since a larger number of substrate molecules undergo transformations per unit time.

For each enzyme, there is an optimal pH value at which it exhibits maximum activity (pepsin - 2.0, salivary amylase - 6.8, pancreatic lipase - 9.0). At higher or lower pH values, the activity of the enzyme decreases. With sharp shifts in pH, the enzyme denatures.

The speed of allosteric enzymes is regulated by substances that attach to allosteric centers. If these substances speed up the reaction, they are called activators if they slow down - inhibitors.

Enzyme classification

According to the type of catalyzed chemical transformations, enzymes are divided into 6 classes:

1. oxidoreductase(transfer of hydrogen, oxygen or electron atoms from one substance to another - dehydrogenase),
2. transferase(transfer of a methyl, acyl, phosphate or amino group from one substance to another - transaminase),
3. hydrolases(hydrolysis reactions in which two products are formed from the substrate - amylase, lipase),
4. lyases(non-hydrolytic addition to the substrate or the elimination of a group of atoms from it, while C-C, C-N, C-O, C-S bonds can be broken - decarboxylase),
5. isomerase(intramolecular rearrangement - isomerase),
6. ligases(the connection of two molecules as a result of the formation of C-C, C-N, C-O, C-S bonds - synthetase).

Classes are in turn subdivided into subclasses and subsubclasses. In the current international classification, each enzyme has a specific code, consisting of four numbers separated by dots. The first number is the class, the second is the subclass, the third is the subclass, the fourth is the serial number of the enzyme in this subclass, for example, the arginase code is 3.5.3.1.

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