The bond underlying the primary structure of a protein. Secondary, tertiary, quaternary protein structures

The primary structure of proteins is a linear polypeptide chain of amino acids connected by peptide bonds. Primary structure - simplest level structural organization of a protein molecule. High stability is given to it by covalent peptide bonds between the α-amino group of one amino acid and the α-carboxyl group of another amino acid.

If the imino group of proline or hydroxyproline is involved in the formation of a peptide bond, then it has a different form

When peptide bonds form in cells, the carboxyl group of one amino acid is first activated, and then it combines with the amino group of another. Laboratory synthesis of polypeptides is carried out in approximately the same way.

A peptide bond is a repeating fragment of a polypeptide chain. It has a number of features that affect not only the shape of the primary structure, but also the higher levels of organization of the polypeptide chain:

· coplanarity - all atoms included in the peptide group are in the same plane;

· ability to exist in two resonant forms (keto or enol form);

· trans position of the substituents relative to the C-N bond;

· the ability to form hydrogen bonds, and each of the peptide groups can form two hydrogen bonds with other groups, including peptide ones.

The exception is peptide groups involving the amino group of proline or hydroxyproline. They are only able to form one hydrogen bond (see above). This affects the formation of the secondary structure of the protein. The polypeptide chain in the area where proline or hydroxyproline is located easily bends, since it is not held, as usual, by a second hydrogen bond.

tripeptide formation scheme:

Levels of spatial organization of proteins: secondary structure of proteins: concept of α-helix and β-sheet layer. Tertiary structure of proteins: the concept of native protein and protein denaturation. Quaternary structure of proteins using the example of the structure of hemoglobin.

Secondary structure of protein. The secondary structure of a protein refers to the way the polypeptide chain is arranged into an ordered structure. According to configuration, the following elements of the secondary structure are distinguished: α -spiral and β - folded layer.

Building model α-helices, taking into account all the properties of the peptide bond, was developed by L. Pauling and R. Corey (1949 - 1951).

In Figure 3, A diagram shown α -spiral, giving an idea of ​​its main parameters. The polypeptide chain folds into α -spiral in such a way that the turns of the spiral are regular, therefore the spiral configuration has helical symmetry (Fig. 3, b). For every turn α -helix has 3.6 amino acid residues. The distance between the turns or the helix pitch is 0.54 nm, the angle of the turn is 26°. Formation and maintenance α -helical configuration occurs due to hydrogen bonds formed between the peptide groups of each n-th and ( P+ 3)-th amino acid residues. Although the energy of hydrogen bonds is small, a large number of them leads to a significant energetic effect, resulting in α -spiral configuration is quite stable. Side radicals of amino acid residues are not involved in maintaining α -helical configuration, so all amino acid residues in α -spirals are equivalent.

In natural proteins, only right-handed ones exist. α -spirals.

β-fold layer- the second element of the secondary structure. Unlike α -spirals β -the folded layer has a linear rather than rod shape (Fig. 4). The linear structure is maintained due to the formation of hydrogen bonds between peptide groups located at different parts of the polypeptide chain. These areas turn out to be close to the distance of the hydrogen bond between - C = O and HN - groups (0.272 nm).

Rice. 4. Schematic illustration β -folded layer (arrows indicate

o direction of the polypeptide chain)

Rice. 3. Scheme ( A) and model ( b) α -spirals

The secondary structure of a protein is determined by the primary structure. Amino acid residues are capable of forming hydrogen bonds to varying degrees, which affects the formation α -spirals or β -layer. Helix-forming amino acids include alanine, glutamic acid, glutamine, leucine, lysine, methionine and histidine. If a protein fragment consists mainly of the amino acid residues listed above, then a α -spiral. Valine, isoleucine, threonine, tyrosine and phenylalanine contribute to the formation β -layers of the polypeptide chain. Disordered structures arise in sections of the polypeptide chain where amino acid residues such as glycine, serine, aspartic acid, asparagine, and proline are concentrated.

Many proteins simultaneously contain α -spirals, and β -layers. The proportion of helical configuration varies among proteins. Thus, the muscle protein paramyosin is almost 100% helical; the proportion of helical configuration in myoglobin and hemoglobin is high (75%). On the contrary, in trypsin and ribonuclease, a significant part of the polypeptide chain fits into layered β -structures. Supporting tissue proteins - keratin (hair protein), collagen (skin and tendon protein) - have β -configuration of polypeptide chains.

Tertiary structure of a protein. The tertiary structure of a protein is the way the polypeptide chain is arranged in space. In order for a protein to acquire its inherent functional properties, the polypeptide chain must fold in a certain way in space, forming a functionally active structure. This structure is called native. Despite the enormous number of spatial structures theoretically possible for an individual polypeptide chain, protein folding leads to the formation of a single native configuration.

The tertiary structure of the protein is stabilized by interactions that occur between the side radicals of amino acid residues of different parts of the polypeptide chain. These interactions can be divided into strong and weak.

Strong interactions include covalent bonds between the sulfur atoms of cysteine ​​residues located in different parts of the polypeptide chain. Otherwise, such bonds are called disulfide bridges; The formation of a disulfide bridge can be depicted as follows:

In addition to covalent bonds, the tertiary structure of a protein molecule is maintained by weak interactions, which, in turn, are divided into polar and nonpolar.

Polar interactions include ionic and hydrogen bonds. Ionic interactions are formed by the contact of positively charged groups of side radicals of lysine, arginine, histidine and the negatively charged COOH group of aspartic and glutamic acids. Hydrogen bonds arise between the functional groups of side radicals of amino acid residues.

Nonpolar or van der Waals interactions between hydrocarbon radicals of amino acid residues contribute to the formation hydrophobic core (fat drop) inside the protein globule, because hydrocarbon radicals strive to avoid contact with water. The more nonpolar amino acids a protein contains, the big role Van der Waals bonds play a role in the formation of its tertiary structure.

Numerous bonds between the side radicals of amino acid residues determine the spatial configuration of the protein molecule (Fig. 5).

Rice. 5. Types of bonds that support the tertiary structure of a protein:
A- disulfide bridge; b - ionic bond; c, d - hydrogen bonds;
d - van der Waals connections

The tertiary structure of an individual protein is unique, as is its primary structure. Only the correct spatial arrangement of the protein makes it active. Various violations of the tertiary structure lead to changes in protein properties and loss of biological activity.

Quaternary protein structure. Proteins with a molecular weight of more than 100 kDa 1 consist, as a rule, of several polypeptide chains with a relatively small molecular weight. A structure consisting of a certain number of polypeptide chains occupying a strictly fixed position relative to each other, as a result of which the protein has one or another activity, is called the quaternary structure of the protein. A protein with a quaternary structure is called epimolecule or multimer , and its constituent polypeptide chains - respectively subunits or protomers . A characteristic property of proteins with a quaternary structure is that an individual subunit does not have biological activity.

Stabilization of the quaternary structure of the protein occurs due to polar interactions between the side radicals of amino acid residues localized on the surface of the subunits. Such interactions firmly hold the subunits in the form of an organized complex. The areas of subunits where interactions occur are called contact areas.

A classic example of a protein with a quaternary structure is hemoglobin. The hemoglobin molecule with a molecular weight of 68,000 Da consists of four subunits of two different types - α And β / α -The subunit consists of 141 amino acid residues, a β - from 146. Tertiary structure α - And β -subunits are similar, as is their molecular weight (17,000 Da). Each subunit contains a prosthetic group - heme . Since heme is also present in other proteins (cytochromes, myoglobin), which will be studied further, we will at least briefly discuss the structure of the topic (Fig. 6). The heme group is a complex coplanar cyclic system consisting of a central atom that forms coordination bonds with four pyrrole residues connected by methane bridges (= CH -). In hemoglobin, iron is usually in an oxidized state (2+).

Four subunits - two α and two β - are connected into a single structure in such a way that α -subunits contact only with β -subunits and vice versa (Fig. 7).

Rice. 6. Structure of heme hemoglobin

Rice. 7. Schematic representation of the quaternary structure of hemoglobin:
Fe - hemoglobin heme

As can be seen from Figure 7, one hemoglobin molecule is capable of carrying 4 oxygen molecules. Both the binding and release of oxygen are accompanied by conformational changes in the structure α - And β -hemoglobin subunits and their relative arrangement in the epimolecule. This fact indicates that the quaternary structure of the protein is not absolutely rigid.

Related information.

5. Regulatory function. Proteins perform the functions of signaling substances - some hormones, histohormones and neurotransmitters, are receptors for signaling substances of any structure, and ensure further signal transmission in the biochemical signal chains of the cell. Examples include growth hormone somatotropin, the hormone insulin, H- and M-cholinergic receptors.

6. Motor function . With the help of proteins, the processes of contraction and other biological movement are carried out. Examples include tubulin, actin, and myosin.

7. Spare function. Plants contain reserve proteins, which are valuable nutrients; in animal bodies, muscle proteins serve as reserve nutrients that are mobilized when absolutely necessary.

Proteins are characterized by the presence of several levels of structural organization.

Primary structure A protein is a sequence of amino acid residues in a polypeptide chain. A peptide bond is a carboxamide bond between the α-carboxyl group of one amino acid and the α-amino group of another amino acid.

alanylphenylalanylcysteylproline

U p eptide bond there are several features:

a) it is resonantly stabilized and therefore is located practically in the same plane - planar; rotation around the C-N bond requires a lot of energy and is difficult;

b) the -CO-NH- bond has a special character, it is smaller than a regular one, but larger than a double one, that is, there is keto-enol tautomerism:

c) substituents in relation to the peptide bond are in trance-position;

d) the peptide backbone is surrounded by side chains of various nature, interacting with the surrounding solvent molecules, free carboxyl and amino groups are ionized, forming cationic and anionic centers of the protein molecule. Depending on their ratio, the protein molecule receives a total positive or negative charge, and is also characterized by one or another pH value of the environment when it reaches the isoelectric point of the protein. Radicals form salt, ether, and disulfide bridges inside the protein molecule, and also determine the range of reactions characteristic of proteins.

Currently agreed to consider polymers consisting of 100 or more amino acid residues as proteins, polypeptides - polymers consisting of 50-100 amino acid residues, low molecular weight peptides - polymers consisting of less than 50 amino acid residues.

Some low molecular weight peptides play an independent biological role. Examples of some of these peptides:

Glutathione - γ-glu-cis-gly - one one of the most widespread intracellular peptides, takes part in redox processes in cells and the transfer of amino acids across biological membranes.

Carnosine - β-ala-his - peptide, contained in the muscles of animals, eliminates the products of lipid peroxide breakdown, accelerates the process of breakdown of carbohydrates in the muscles and is involved in the energy metabolism in the muscles in the form of phosphate.

Vasopressin is a hormone of the posterior lobe of the pituitary gland, involved in the regulation of water metabolism in the body:

Phalloidin- poisonous fly agaric polypeptide, in negligible concentrations causes the death of the body due to the release of enzymes and potassium ions from cells:

Gramicidin - antibiotic, acting on many gram-positive bacteria, changes the permeability of biological membranes for low molecular weight compounds and causes cell death:

Meth-enkephalin - tyr-gly-gly-phen-met - a peptide synthesized in neurons and reducing pain.

Protein secondary structure is a spatial structure formed as a result of interactions between functional groups of the peptide backbone.

The peptide chain contains many CO and NH groups of peptide bonds, each of which is potentially capable of participating in the formation of hydrogen bonds. There are two main types of structures that allow this to happen: an α-helix, in which the chain is coiled up like a telephone cord, and a folded β-structure, in which elongated sections of one or more chains are laid side by side. Both of these structures are very stable.

The α-helix is ​​characterized by extremely dense packing of a twisted polypeptide chain; for each turn of a right-handed helix there are 3.6 amino acid residues, the radicals of which are always directed outward and slightly backward, that is, to the beginning of the polypeptide chain.

Main characteristics of the α-helix:

1) the α-helix is ​​stabilized by hydrogen bonds between the hydrogen atom at the nitrogen of the peptide group and the carbonyl oxygen of the residue located four positions along the chain;

2) all peptide groups participate in the formation of a hydrogen bond, this ensures maximum stability of the α-helix;

3) all nitrogen and oxygen atoms of peptide groups are involved in the formation of hydrogen bonds, which significantly reduces the hydrophilicity of α-helical regions and increases their hydrophobicity;

4) the α-helix is ​​formed spontaneously and is the most stable conformation of the polypeptide chain, corresponding to the minimum free energy;

5) in a polypeptide chain of L-amino acids, the right-handed helix, usually found in proteins, is much more stable than the left-handed one.

Possibility of α-helix formation determined by the primary structure of the protein. Some amino acids prevent the peptide backbone from twisting. For example, the adjacent carboxyl groups of glutamate and aspartate mutually repel each other, which prevents the formation of hydrogen bonds in the α-helix. For the same reason, chain helicalization is difficult in places where positively charged lysine and arginine residues are located close to each other. However, proline plays the largest role in α-helix disruption. Firstly, in proline the nitrogen atom is part of a rigid ring, which prevents rotation around N-C connections, secondly, proline does not form a hydrogen bond due to the absence of hydrogen at the nitrogen atom.

β-sheeting is a layered structure, formed by hydrogen bonds between linearly arranged peptide fragments. Both chains can be independent or belong to the same polypeptide molecule. If the chains are oriented in the same direction, then such a β-structure is called parallel. In the case of opposite chain directions, that is, when the N-terminus of one chain coincides with the C-terminus of another chain, the β-structure is called antiparallel. An antiparallel β-sheet with nearly linear hydrogen bridges is energetically more preferable.

parallel β-sheet antiparallel β-sheet

Unlike the α-helix saturated with hydrogen bonds, each section of the β-sheet chain is open to the formation of additional hydrogen bonds. The side radicals of amino acids are oriented almost perpendicular to the plane of the sheet, alternately up and down.

In those areas where the peptide chain bends quite sharply, often containing a β-loop. This is a short fragment in which 4 amino acid residues are bent by 180° and are stabilized by one hydrogen bridge between the first and fourth residues. Large amino acid radicals interfere with the formation of the β-loop, so it most often includes the smallest amino acid, glycine.

Protein suprasecondary structure- this is some specific order of alternation of secondary structures. A domain is understood as a separate part of a protein molecule that has a certain degree of structural and functional autonomy. Domains are now considered fundamental elements of the structure of protein molecules, and the relationship and nature of the arrangement of α-helices and β-sheets provides more for understanding the evolution of protein molecules and phylogenetic relationships than a comparison of primary structures.

The main task of evolution is designing more and more new proteins. There is an infinitesimal chance of accidentally synthesizing an amino acid sequence that would satisfy the packaging conditions and ensure the fulfillment of functional tasks. Therefore, it is common to find proteins with different functions but so similar in structure that they appear to have had a common ancestor or to have evolved from each other. It seems that evolution, when faced with the need to solve a certain problem, prefers not to design proteins for this purpose from the beginning, but to adapt already well-established structures for this purpose, adapting them for new purposes.

Some examples of frequently repeated suprasecondary structures:

1) αα’ - proteins containing only α-helices (myoglobin, hemoglobin);

2) ββ’ - proteins containing only β-structures (immunoglobulins, superoxide dismutase);

3) βαβ’ - β-barrel structure, each β-layer is located inside the barrel and is connected to an α-helix located on the surface of the molecule (triose phosphoisomerase, lactate dehydrogenase);

4) “zinc finger” - a protein fragment consisting of 20 amino acid residues, the zinc atom is linked to two cysteine ​​residues and two histidine residues, resulting in the formation of a “finger” of approximately 12 amino acid residues, which can bind to the regulatory regions of the DNA molecule;

5) “leucine zipper” - interacting proteins have an α-helical region containing at least 4 leucine residues, they are located 6 amino acids apart, that is, they are on the surface of every second turn and can form hydrophobic bonds with leucine residues another protein. With the help of leucine zippers, for example, molecules of strongly basic histone proteins can be complexed, overcoming a positive charge.

Protein tertiary structure- this is the spatial arrangement of the protein molecule, stabilized by the bonds between the side radicals of amino acids.

Types of bonds that stabilize the tertiary structure of a protein:

electrostatic hydrogen hydrophobic disulfide interaction bonds interactions bonds

Depending on folding The tertiary structure of proteins can be classified into two main types - fibrillar and globular.

Fibrillar proteins- long, thread-like molecules insoluble in water, the polypeptide chains of which are elongated along one axis. These are mainly structural and contractile proteins. Some examples of the most common fibrillar proteins:

1. α- Keratins. Synthesized by epidermal cells. They account for almost all the dry weight of hair, fur, feathers, horns, nails, claws, quills, scales, hooves and turtle shell, as well as a significant portion of the weight of the outer layer of skin. This is a whole family of proteins; they are similar in amino acid composition, contain many cysteine ​​residues and have the same spatial arrangement of polypeptide chains.

In hair cells, polypeptide chains of keratin first organized into fibers, from which structures are then formed like a rope or twisted cable, eventually filling the entire space of the cell. The hair cells become flattened and finally die, and the cell walls form a tubular sheath called the cuticle around each hair. In α-keratin, the polypeptide chains have the shape of an α-helix, twisted around one another into a three-core cable with the formation of cross-disulfide bonds.

The N-terminal residues are located on one side (parallel). Keratins are insoluble in water due to the predominance of amino acids in their composition with non-polar side radicals that face the aqueous phase. During perm, the following processes occur: first, disulfide bridges are destroyed by reduction with thiols, and then, when the hair is given the required shape, it is dried by heating, while due to oxidation with atmospheric oxygen, new disulfide bridges are formed, which retain the shape of the hairstyle.

2. β-Keratins. These include silk and spider web fibroin. They are antiparallel β-pleated layers with a predominance of glycine, alanine and serine in the composition.

3. Collagen. The most common protein in higher animals and the main fibrillar protein of connective tissues. Collagen is synthesized in fibroblasts and chondrocytes - specialized connective tissue cells, from which it is then expelled. Collagen fibers are found in skin, tendons, cartilage and bones. They do not stretch, are stronger than steel wire, and collagen fibrils are characterized by transverse striations.

When boiled in water, fibrous, insoluble and indigestible collagen is converted into gelatin by hydrolysis of some covalent bonds. Collagen contains 35% glycine, 11% alanine, 21% proline and 4-hydroxyproline (an amino acid unique to collagen and elastin). This composition determines the relatively low nutritional value of gelatin as a food protein. Collagen fibrils are composed of repeating polypeptide subunits called tropocollagen. These subunits are arranged along the fibril in the form of parallel bundles in a head-to-tail fashion. The displacement of the heads gives the characteristic transverse striations. The voids in this structure, if necessary, can serve as a site for the deposition of crystals of hydroxyapatite Ca 5 (OH)(PO 4) 3, which plays an important role in bone mineralization.

Tropocollagen subunits consist of of three polypeptide chains tightly coiled into a three-strand rope, distinct from α- and β-keratins. In some collagens, all three chains have the same amino acid sequence, while in others only two chains are identical, and the third is different. The polypeptide chain of tropocollagen forms a left-handed helix, with only three amino acid residues per turn due to the chain bends caused by proline and hydroxyproline. The three chains are connected to each other, in addition to hydrogen bonds, by a covalent-type bond formed between two lysine residues located in adjacent chains:

As we get older, in tropocollagen subunits and between them everything is formed larger number cross-links, which makes collagen fibrils more rigid and brittle, and this changes the mechanical properties of cartilage and tendons, makes bones more brittle and reduces the transparency of the cornea.

4. Elastin. Contained in the yellow elastic tissue of ligaments and the elastic layer of connective tissue in the walls of large arteries. The main subunit of elastin fibrils is tropoelastin. Elastin is rich in glycine and alanine, contains a lot of lysine and little proline. The spiral sections of elastin stretch when tension is applied, but return to their original length when the load is removed. The lysine residues of four different chains form covalent bonds with each other and allow elastin to stretch reversibly in all directions.

Globular proteins- proteins, the polypeptide chain of which is folded into a compact globule, are capable of performing a wide variety of functions.

Tertiary structure of globular proteins It is most convenient to consider using the example of myoglobin. Myoglobin is a relatively small oxygen-binding protein found in muscle cells. It stores bound oxygen and promotes its transfer to mitochondria. The myoglobin molecule contains one polypeptide chain and one hemogroup (heme) - a complex of protoporphyrin with iron.

Basic properties myoglobin:

a) the myoglobin molecule is so compact that only 4 water molecules can fit inside it;

b) all polar amino acid residues, with the exception of two, are located on the outer surface of the molecule, and all of them are in a hydrated state;

c) most of the hydrophobic amino acid residues are located inside the myoglobin molecule and, thus, are protected from contact with water;

d) each of the four proline residues in the myoglobin molecule is located at the bend site of the polypeptide chain; serine, threonine and asparagine residues are located at other bend sites, since such amino acids prevent the formation of an α-helix if they are located next to each other;

e) a flat heme group lies in a cavity (pocket) near the surface of the molecule, the iron atom has two coordination bonds directed perpendicular to the heme plane, one of them is connected to histidine residue 93, and the other serves to bind an oxygen molecule.

Starting with protein tertiary structure becomes capable of performing its inherent biological functions. The basis of the functioning of proteins is that when a tertiary structure is laid on the surface of the protein, areas are formed that can attach other molecules, called ligands. The high specificity of the interaction of the protein with the ligand is ensured by the complementarity of the structure of the active center to the structure of the ligand. Complementarity is the spatial and chemical correspondence of interacting surfaces. For most proteins, tertiary structure is the maximum level of folding.

Quaternary protein structure- characteristic of proteins consisting of two or more polypeptide chains connected to each other exclusively covalent bonds, mainly electrostatic and hydrogen. Most often, proteins contain two or four subunits; more than four subunits usually contain regulatory proteins.

Proteins with quaternary structure, are often called oligomeric. There are homomeric and heteromeric proteins. Homomeric proteins include proteins in which all subunits have the same structure, for example, the enzyme catalase consists of four absolutely identical subunits. Heteromeric proteins have different subunits; for example, the enzyme RNA polymerase consists of five structurally different subunits that perform different functions.

Single subunit interaction with a specific ligand causes conformational changes in the entire oligomeric protein and changes the affinity of other subunits for ligands; this property underlies the ability of oligomeric proteins for allosteric regulation.

The quaternary structure of a protein can be examined using the example of hemoglobin. Contains four polypeptide chains and four heme prosthetic groups, in which the iron atoms are in the ferrous form Fe 2+. The protein part of the molecule - globin - consists of two α-chains and two β-chains containing up to 70% α-helices. Each of the four chains has a characteristic tertiary structure, and one hemogroup is associated with each chain. The hemes of different chains are located relatively far from each other and have different inclination angles. Few direct contacts are formed between two α-chains and two β-chains, while numerous contacts of the α 1 β 1 and α 2 β 2 type formed by hydrophobic radicals arise between the α and β chains. Between α 1 β 1 and α 2 β 2 a channel remains.

Unlike myoglobin hemoglobin characterized significantly lower affinity for oxygen, which allows it, at the low partial pressures of oxygen existing in the tissues, to give them a significant part of the bound oxygen. Oxygen is more easily bound by hemoglobin iron at higher pH values ​​and low CO 2 concentrations characteristic of the alveoli of the lungs; the release of oxygen from hemoglobin is favored by lower pH values ​​and high concentrations of CO 2 characteristic of tissues.

In addition to oxygen, hemoglobin carries hydrogen ions, which bind to histidine residues in the chains. Hemoglobin also carries carbon dioxide, which attaches to the terminal amino group of each of the four polypeptide chains, resulting in the formation of carbaminohemoglobin:

IN red blood cells in fairly high concentrations the substance 2,3-diphosphoglycerate (DPG) is present, its content increases with rising to high altitudes and during hypoxia, facilitating the release of oxygen from hemoglobin in the tissues. DPG is located in the channel between α 1 β 1 and α 2 β 2, interacting with positively contaminated groups of β-chains. When hemoglobin binds oxygen, DPG is forced out of the cavity. The red blood cells of some birds contain not DPG, but inositol hexa-phosphate, which further reduces the affinity of hemoglobin for oxygen.

2,3-Diphosphoglycerate (DPG)

HbA - normal adult hemoglobin, HbF - fetal hemoglobin, has a greater affinity for O 2, HbS - hemoglobin in sickle cell anemia. Sickle cell anemia is a serious inherited disease caused by a genetic abnormality of hemoglobin. In the blood of sick people there is an unusually large number of thin sickle-shaped red blood cells, which, firstly, easily rupture, and secondly, clog the blood capillaries.

At the molecular level, hemoglobin S is different from hemoglobin A there is one amino acid residue at position 6 of the β-chains, where instead of a glutamic acid residue there is valine. Thus, hemoglobin S contains two less negative charges; the appearance of valine leads to the appearance of a “sticky” hydrophobic contact on the surface of the molecule; as a result, during deoxygenation, deoxyhemoglobin S molecules stick together and form insoluble, abnormally long thread-like aggregates, leading to deformation of red blood cells.

There is no reason to think that there is independent genetic control over the formation of levels of protein structural organization above the primary, since the primary structure determines the secondary, tertiary, and quaternary (if any). The native conformation of a protein is the thermodynamically most stable structure under given conditions.

LECTURE 6

There are physical, chemical and biological properties of proteins.

Physical properties of proteins are the presence of molecular weight, birefringence (a change in the optical characteristics of a protein solution in motion compared to a solution at rest), due to the nonspherical shape of proteins, mobility in an electric field, due to the charge of protein molecules. In addition, proteins are characterized by optical properties consisting in the ability to rotate the plane of polarization of light, scatter light rays due to the large size of protein particles, and absorb ultraviolet rays.

One of the characteristic physical properties proteins are the ability to adsorb on the surface, and sometimes to capture molecules, low molecular weight organic compounds and ions.

The chemical properties of proteins differ exceptional diversity, since proteins are characterized by all reactions of amino acid radicals and are characterized by the reaction of hydrolysis of peptide bonds.

Having a significant number of acidic and basic groups, proteins exhibit amphoteric properties. Unlike free amino acids, the acid-base properties of proteins are determined not by α-amino and α-carboxy groups involved in the formation of peptide bonds, but by charged radicals of amino acid residues. The main properties of proteins are determined by arginine, lysine and histidine residues. The acidic properties are due to aspartic and glutamic acid residues.

Protein titration curves are sufficient difficult to interpret, since any protein contains too many big number titratable groups, there are electrostatic interactions between the ionized groups of the protein; the pK of each titratable group is influenced by nearby hydrophobic residues and hydrogen bonds. Greatest practical use has the isoelectric point of a protein - the pH value at which the total charge of the protein is zero. At the isoelectric point, the protein is maximally inert, does not move in an electric field, and has the thinnest hydration shell.

Proteins exhibit buffering properties, but their buffer capacity is insignificant. The exception is proteins containing a large number of histidine residues. For example, the hemoglobin contained in erythrocytes, due to the very high content of histidine residues, has a significant buffer capacity at a pH of about 7, which is very important for the role that erythrocytes play in the transport of oxygen and carbon dioxide in the blood.

Proteins are characterized by solubility in water, and from a physical point of view they form true molecular solutions. However, protein solutions are characterized by some colloidal properties: the Tendahl effect (light scattering phenomenon), the inability to pass through semi-permeable membranes, high viscosity, and the formation of gels.

Protein solubility is highly dependent on the concentration of salts, that is, on the ionic strength of the solution. In distilled water, proteins are most often poorly soluble, but their solubility increases as the ionic strength increases. At the same time, an increasing number of hydrated inorganic ions bind to the surface of the protein and thereby the degree of its aggregation decreases. At high ionic strength, salt ions take away the hydration shell from protein molecules, which leads to aggregation and precipitation of proteins (salting out phenomenon). Using differences in solubility, it is possible to separate a mixture of proteins using common salts.

Among the biological properties of proteins primarily include their catalytic activity. Another important biological property of proteins is their hormonal activity, that is, the ability to influence entire groups of reactions in the body. Some proteins have toxic properties, pathogenic activity, protective and receptor functions, and are responsible for cell adhesion phenomena.

Another unique biological property of proteins- denaturation. Proteins in their natural state are called native. Denaturation is the destruction of the spatial structure of proteins under the action of denaturing agents. The primary structure of proteins is not damaged during denaturation, but their biological activity is lost, as well as solubility, electrophoretic mobility and some other reactions. When denatured, the amino acid radicals that form the active center of the protein are spatially distant from each other, that is, the specific binding center of the protein with the ligand is destroyed. Hydrophobic radicals, usually located in the hydrophobic core of globular proteins, upon denaturation end up on the surface of the molecule, thereby creating conditions for aggregation of proteins that precipitate.

Reagents and conditions that cause protein denaturation:

Temperature above 60 o C - destruction of weak bonds in the protein,

Acids and alkalis - change in ionization of ionogenic groups, breaking of ionic and hydrogen bonds,

Urea - destruction of intramolecular hydrogen bonds as a result of the formation of hydrogen bonds with urea,

Alcohol, phenol, chloramine - destruction of hydrophobic and hydrogen bonds,

Salts of heavy metals - the formation of insoluble salts of proteins with heavy metal ions.

When denaturing agents are removed, renativation is possible, since the peptide chain tends to adopt the conformation with the lowest free energy in solution.

Under cellular conditions, proteins can spontaneously denature, although at a lower rate than at high temperature. Spontaneous renativation of proteins in the cell is difficult, since due to the high concentration there is a high probability of aggregation of partially denatured molecules.

Cells contain proteins- molecular chaperones that have the ability to bind to partially denatured proteins that are in an unstable state, prone to aggregation, and restore their native conformation. Initially, these proteins were discovered as heat shock proteins, since their synthesis increased when the cell was exposed to stress, for example, when the temperature increased. Chaperones are classified according to the mass of their subunits: hsp-60, hsp-70 and hsp-90. Each class includes a family of related proteins.

Molecular chaperones ( hsp-70) a highly conserved class of proteins found in all parts of the cell: cytoplasm, nucleus, endoplasmic reticulum, mitochondria. At the C-terminus of the single polypeptide chain, hsp-70 has a region that is a groove capable of interacting with peptides 7-9 amino acid residues long, enriched in hydrophobic radicals. Such regions in globular proteins occur approximately every 16 amino acids. Hsp-70 is able to protect proteins from temperature inactivation and restore the conformation and activity of partially denatured proteins.

Chaperones-60 (hsp-60) participate in the formation of the tertiary structure of proteins. Hsp-60 functions as oligomeric proteins consisting of 14 subunits. Hsp-60 forms two rings, each ring consists of 7 subunits connected to each other.

Each subunit consists of three domains:

The apical domain has a number of hydrophobic amino acid residues facing the inside of the cavity formed by the subunits;

The equatorial domain has ATPase activity and is necessary for the release of the protein from the chaperonin complex;

The intermediate domain connects the apical and equatorial domains.

A protein that has fragments on its surface, enriched with hydrophobic amino acids, enters the cavity of the chaperonin complex. In the specific environment of this cavity, in conditions of isolation from other molecules of the cell cytosol, the selection of possible protein conformations occurs until an energetically more favorable conformation is found. The chaperone-dependent formation of the native conformation is associated with the expenditure of a significant amount of energy, the source of which is ATP.

1. Primary structure of the protein. Dependence of the properties and conformation of proteins on the primary structure. Examples of protein polymorphisms, hemoglobin A and F, structural and functional differences. The role of fetal hemoglobin in period of intrauterine development of the fetus . Hereditary changes in the primary structure - molecular diseases (sickle cell anemia).

The primary structure of a protein is the sequence of alternating amino acids in a polypeptide chain. This structure is formed by peptide bonds between the α-amino and α-carboxyl groups of amino acids (see 1.4.2). Keep in mind that even small changes in the primary structure of a protein can significantly change its properties. An example of diseases that develop as a result of changes in the primary structure of a protein are hemoglobinopathies (hemoglobinoses).

Hemoglobin A (Hb A) is present in the red blood cells of healthy adults. Some people's blood contains abnormal (altered) hemoglobin - hemoglobin (Hb S). The only difference between the primary structure of Hb S and Hb A is the replacement of the hydrophilic glutamic acid residue with a hydrophobic valine residue in the terminal region of their β-chains.

As you know, the main function of hemoglobin is to transport oxygen to tissues. Under conditions of reduced partial pressure of O2, the solubility of hemoglobin S in water and its ability to bind and transport oxygen decreases. The red blood cells take on a sickle shape and are quickly destroyed, resulting in anemia (sickle cell anemia).

It has been established that the sequence of amino acid residues of the polypeptide chain of a protein carries the information necessary for the formation of the spatial structure of the protein. It has been established that each polypeptide sequence corresponds to only one stable variant of the spatial structure. The process of folding a polypeptide chain into a regular three-dimensional structure is called folding

Until recently, it was believed that the formation of the spatial structure of a protein occurs spontaneously, without the participation of any components. However, relatively recently it was discovered that this is true only for relatively small proteins (about 100 amino acid residues). In the process of folding larger proteins, special proteins take part - chaperones, which create the possibility of rapid formation of the correct spatial structure of the protein.

An example of protein polymorphism is hemoglobin, which has many forms. Hemoglobin A is the normal hemoglobin of an adult. This protein is a tetramer consisting of two pairs of polypeptide chains - monomers: two α-chain monomers and two β-chain monomers, or two α monomers and two δ monomers. Hemoglobin F is the fetal type of human hemoglobin. Hemoglobin F is a heterotetramer protein of two α-chains and two γ-chains of globin. Hemoglobin F has an increased affinity for oxygen (it contains serine instead of lysine) and allows the relatively small volume of fetal blood to perform oxygen supply functions more efficiently. However, hemoglobin F is less resistant to degradation and less stable. During the last trimester of pregnancy and after birth, hemoglobin F is gradually replaced by “adult” hemoglobin A (HbA), a less active oxygen transporter, but more resistant to destruction and more stable. Molecular diseases are hereditary disorders in the primary structure of the bun. For example, replacing the sixth glutamine amino acid with valine in the β-subunit of hemoglobin leads to the formation of hemoglobin S and the fact that the hemoglobin molecule as a whole cannot perform its main function - oxygen transport; In such cases, the person develops a disease called sickle cell anemia.

2. Conformation of the protein molecule (secondary and tertiary structures). Types of intramolecular bonds in proteins. Fibrillar and globular proteins (examples). Quaternary structure of protein. Examples of the structure and functioning of oligomeric proteins.

Protein secondary structure represents a method of folding a polypeptide chain into a helical or other conformation. In this case, hydrogen bonds are formed between the CO and NH groups of the peptide backbone of one chain or adjacent polypeptide chains. Several types of secondary structure of peptide chains are known, among which the main ones are the α-helix and β-sheet layer.

α-Helix- rigid structure, looks like a rod. The inner part of this rod is created by a tightly twisted peptide backbone, the amino acid radicals are directed outward. In this case, the CO group of each amino acid residue interacts with the NH group of the fourth residue from it. There are 3.6 amino acid residues per turn of the helix, and the helix pitch is 0.54 nm (Figure 2.1).

Figure 2.1.α-Helix.

Some amino acids prevent the chain from folding into an α-helix, and at their location the continuity of the helix is ​​disrupted. These amino acids include proline (in which the nitrogen atom is part of a rigid ring structure and rotation around the N - C α bond becomes impossible), as well as amino acids with charged radicals that electrostatically or mechanically prevent the formation of an α-helix. If there are two such radicals (or more) within one turn (about 4 amino acid residues), they interact and deform the helix.

β-fold layer differs from an α-helix in that it is flat rather than rod-shaped. Formed by hydrogen bonds within one or more polypeptide chains. The peptide chains can be arranged in the same direction (parallel) or in opposite directions (antiparallel), resembling the bellows of an accordion. Lateral radicals are located above and below the plane of the layer.

Figure 2.2.β-folded layer.

Note that the type of secondary structure of a protein is determined by its primary structure. For example, at the location of the proline residue (the atoms of the pyrrolidine ring in proline lie in the same plane), the peptide chain bends, and hydrogen bonds between amino acids are not formed. Therefore, proteins with a high proline content (for example, collagen) are not able to form an α-helix. Amino acid radicals carrying electric charge, also prevent spiralization.

2.1.3. The tertiary structure of a protein is the distribution in space of all the atoms of the protein molecule, or in other words, spatial packing of a helical polypeptide chain. The main role in the formation of the tertiary structure of a protein is played by hydrogen, ionic, hydrophobic and disulfide bonds, which are formed as a result of the interaction between amino acid radicals.

• Hydrogen bonds are formed between two polar uncharged radicals or between an uncharged and a charged radical, for example, serine and glutamine radicals:

• Ionic bonds can occur between oppositely charged radicals, for example, glutamate and arginine radicals:

• Hydrophobic interactions are typical for non-polar radicals, for example, valine and leucine:

• Disulfide bonds are formed between the SH groups of two cysteine ​​radicals located in different parts of the polypeptide chain:
  .

Based on the shape of the molecule and the characteristics of the formation of the tertiary structure, proteins are divided into globular and fibrillar.

Globular proteins- have a spherical or ellipsoidal molecule shape (globule). During the formation of a globule, hydrophobic amino acid radicals are immersed in the internal regions, while hydrophilic radicals are located on the surface of the molecule. When interacting with the aqueous phase, polar radicals form numerous hydrogen bonds. Proteins are held in a dissolved state due to their charge and hydration shell. In the body, globular proteins perform dynamic functions(transport, enzymatic, regulatory, protective). Globular proteins include:

• Albumen - blood plasma protein; contains many glutamate and aspartate residues; precipitates at 100% saturation of the solution with ammonium sulfate.
• Globulins - blood plasma proteins; Compared to albumin, they have a higher molecular weight and contain fewer glutamate and aspartate residues; they precipitate at 50% saturation of the solution with ammonium sulfate.
• Histones - are part of cell nuclei, where they form a complex with DNA. Contains many arginine and lysine residues.

Fibrillar proteins- have a thread-like shape (fibrils), form fibers and bundles of fibers. There are many covalent cross-links between adjacent polypeptide chains. Insoluble in water. The transition into solution is prevented by non-polar amino acid radicals and cross-links between peptide chains. In the body they perform mainly a structural function, providing mechanical strength to tissues. Fibrillar proteins include:

• Collagen - connective tissue protein. Its composition is dominated by the amino acids glycine, proline, and hydroxyproline.
• Elastin - more elastic than collagen, it is part of the walls of arteries and lung tissue; its composition is dominated by the amino acids glycine, alanine, and valine.
• Keratin - protein of the epidermis and skin derivatives; the amino acid cysteine ​​predominates in its structure.

3. Hemoglobin is an allosteric protein. Conformational changes in the hemoglobin molecule. Cooperative effect. Regulators of hemoglobin affinity for oxygen. Structural and functional differences between myoglobin and hemoglobin.

Hemoproteins include: hemoglobin, myoglobin, cytochromes, peroxidase, catalase. These proteins contain as a prosthetic group heme.

According to its chemical structure, heme is protoporphyrin IX, associated with ferrous iron. Protoporphyrin IX is an organic compound belonging to the class of porphyrins. Protoporphyrin IX contains four substituted pyrrole rings connected by methine bridges =CH—. The substituents on the pyrrole rings are: four methyl groups CH3—, two vinyl groups CH2=CH— and two propionic acid residues — CH2—CH2—COOH. Heme is connected to the protein part as follows. Non-polar groups. Protoporphyrin IX interacts with hydrophobic regions of amino acids using hydrophobic bonds. In addition, there is a coordination bond between the iron atom and the imidazole histidine radical in the protein chain. Another coordination bond of the iron atom can be used to bind oxygen and other ligands.

The presence of heme-containing proteins in biological material is detected using a benzidine test (when benzidine and hydrogen peroxide are added, the test solution turns blue-green).

compare the structure and function of myoglobin and hemoglobin, remember the characteristic features of each of these proteins.

Myoglobin- a chromoprotein present in muscle tissue and having a high affinity for oxygen. The molecular weight of this protein is about 16,000 Da. The myoglobin molecule has a tertiary structure and represents one polypeptide chain connected to heme. Myoglobin does not have allosteric properties (see 2.4.), its oxygen saturation curve has the shape of a hyperbola (Figure 4). The function of myoglobin is to create an oxygen reserve in the muscles, which is consumed as needed, replenishing the temporary lack of oxygen.

Hemoglobin (Hb)- a chromoprotein present in red blood cells and involved in the transport of oxygen to tissues. Hemoglobin in adults is called hemoglobin A (Hb A). Its molecular weight is about 65,000 Da. The Hb A molecule has a quaternary structure and includes four subunits - polypeptide chains (designated α1, α2, β1 and β2, each of which is associated with heme.

Remember that hemoglobin is an allosteric protein; its molecules can reversibly change from one conformation to another. This changes the affinity of the protein for ligands. The conformation with the least affinity for the ligand is called the tense, or T-conformation. The conformation with the greatest affinity for the ligand is called the relaxed, or R-conformation.

Various environmental factors can shift this balance in one direction or another. Allosteric regulators affecting the affinity of Hb for O2 are: 1) oxygen; 2) H+ concentration (medium pH); 3) carbon dioxide (CO2); 4) 2,3-diphosphoglycerate (DPG). The attachment of an oxygen molecule to one of the hemoglobin subunits promotes the transition of a tense conformation to a relaxed one and increases the affinity for oxygen of other subunits of the same hemoglobin molecule. This phenomenon is called the cooperative effect. The complex nature of the binding of hemoglobin to oxygen is reflected by the hemoglobin O2 saturation curve, which has an S-shape (Figure 3.1).

Figure 3.1. Curves of myoglobin (1) and hemoglobin (2) oxygen saturation.

4. Biological functions of proteins. The role of the spatial organization of the polypeptide chain in the formation of active centers. Interaction of proteins with ligands. Denaturation of proteins.

Proteins play a vital role in the body, performing a variety of biological functions. Remember the most important ones and examples of corresponding proteins by studying Table 2.2.

Table 2.2 Functional classification of proteins

Protein function Essence Examples Catalytic (enzymatic) Acceleration of chemical reactions in the body Pepsin, trypsin, catalase, cytochrome oxidase Transport Transport (transfer) of chemical compounds in the body Hemoglobin, albumin, transferrin Structural plastic Ensuring the strength and elasticity of fabrics Collagen, elastin, keratin Contractive Shortening of muscle sarcomeres (contraction) Actin, myosin Hormonal (regulatory) Regulation of metabolism in cells and tissues insulin, somatotropin, glucagon, corticotropin Protective Protecting the body from damaging factors Interferons, immunoglobulins Energy Release of energy due to the breakdown of amino acids Food and tissue proteins 2.2.2. Please note that the basis for the functioning of any protein is its ability to selectively interact with strictly defined molecules or ions (ligands). For example, for enzymes that catalyze chemical reactions, the ligands will be the substances participating in these reactions (substrates), for transport proteins - the substances being transported, etc. 2.2.3. The ligand is able to interact not with the entire surface of the protein molecule, but only with a certain part of it, which is the binding center or active center. This center is formed by spatially close amino acid radicals at the level of the secondary or tertiary structure of the protein. The ability of a ligand to interact with the binding site is determined by its complementarity, that is, the mutual correspondence of their spatial structure (similar to the “key-lock” interaction). Non-covalent (hydrogen, ionic, hydrophobic) as well as covalent bonds are formed between the functional groups of the ligand and the binding center. The complementarity of the ligand and the binding site can explain the high specificity (selectivity) of the protein-ligand interaction. It is important to note that a change in the spatial structure of the protein during denaturation (see 2.4) leads to the destruction of binding centers and loss biological function squirrel.

Denaturation of proteins This is called a change in the native (natural) physicochemical and, most importantly, biological properties of a protein due to a violation of its quaternary, tertiary and even secondary structure. Protein denaturation can be caused by:

• temperature above 60° C;
• ionizing radiation;
• concentrated acids and alkalis;
• salts of heavy metals (mercury, lead, cadmium);
• organic compounds (alcohols, phenols, ketones).

Denatured proteins are characterized by:

• change in the conformation of the molecule;
• decreased solubility in water;
• change in molecule charge;
• less resistance to the action of proteolytic enzymes;
• loss of biological activity.

Please note that under certain conditions it is possible to restore the original (native) conformation of the protein after removing the factor that caused the denaturation. This process is called regeneration.

Remember some examples of the use of protein denaturation in medicine:

• for sedimentation of blood plasma proteins when determining the content of non-protein substances in the blood;
• during disinfection and sanitization;
• in the treatment and prevention of poisoning with salts of heavy metals (milk or egg white is used as an antidote);
• to obtain medicinal substances of protein nature (denaturation under mild conditions followed by renativation is used).

5. Structure and biological role of nucleotides.

Nucleic acids or polynucleotides are high-molecular substances consisting of nucleotides connected in a chain by 3", 5" phosphodiester bonds. Each nucleotide consists of a nitrogenous base, a carbohydrate (pentose) and a phosphoric acid residue. The nitrogenous bases that make up nucleotides have the following structure: Carbohydrates are represented by ribose and deoxyribose: 4.1.2. The nitrogenous base and pentose, connected by an N-glycosidic bond, form nucleoside. If ribose is present as a pentose in a nucleoside, then it is a ribonucleoside, and if deoxyribose is present, then it is a deoxyribonucleoside.

4.1.3. Nucleotides are phosphorylated nucleosides. The phosphoric acid residue is usually attached to the hydroxyl group of the pentose at the 5" position via an ester bond. Examples:

Nucleoside diphosphates and nucleoside triphosphates, containing two and three phosphoric acid residues, respectively, are also found in cells. Biological role These compounds will be discussed further.

6. Primary and secondary structures of DNA. Chargaff's rules. Principle of complementarity . Types of bonds in a DNA molecule. Biological role of DNA. Molecular diseases are a consequence of gene mutations.

Primary structure of nucleic acids called sequence of arrangement of mononucleotides in a DNA or RNA chain . The primary structure of nucleic acids is stabilized by 3",5" phosphodiester bonds. These bonds are formed by the interaction of the hydroxyl group in the 3" position of the pentose residue of each nucleotide with the phosphate group of the neighboring nucleotide (Figure 3.2),

Thus, at one end of the polynucleotide chain there is a free 5"-phosphate group (5"-end), and at the other there is a free hydroxyl group in the 3" position (3"-end). Nucleotide sequences are usually written in the direction from the 5" end to the 3" end.

DNA (deoxy ribonucleic acid) found in the cell nucleus and has molecular weight about 1011 Yes. Its nucleotides contain nitrogenous bases adenine, guanine, cytosine, thymine , carbohydrate deoxyribose and phosphoric acid residues. The content of nitrogenous bases in a DNA molecule is determined by Chargaff’s rules:

1) the number of purine bases is equal to the number of pyrimidine bases (A + G = C + T);

2) the amount of adenine and cytosine is equal to the amount of thymine and guanine, respectively (A = T; C = G);

3) DNA isolated from cells of different biological species differ from each other in the specificity coefficient:

(G + C) / (A + T)

These patterns in the structure of DNA are explained by the following features of its secondary structure:

1) a DNA molecule is built from two polynucleotide chains connected to each other by hydrogen bonds and oriented antiparallel (that is, the 3" end of one chain is located opposite the 5" end of the other chain and vice versa);

2) hydrogen bonds are formed between complementary pairs of nitrogenous bases. Thymine is complementary to adenine; this pair is stabilized by two hydrogen bonds. Cytosine is complementary to guanine; this pair is stabilized by three hydrogen bonds (see figure b). The more DNA there is in a molecule G-C steam, the greater its resistance to high temperatures and ionizing radiation;

3) both DNA strands are twisted into a helix that has a common axis. The nitrogenous bases face the inside of the helix; In addition to hydrogen interactions, hydrophobic interactions also arise between them. The ribose phosphate moieties are located along the periphery, forming the core of the helix (see Figure 3.4).

Figure 3.4. DNA structure diagram.

7. Primary and secondary structures of RNA. Types of RNA: structural features. The main components of the protein synthesizing system. Function of ribosomes. Adapter function of tRNA and the role of mRNA in protein synthesis.

RNA (ribonucleic acid) is found predominantly in the cytoplasm of the cell and has a molecular weight in the range of 104 - 106 Da. Its nucleotides contain nitrogenous bases adenine, guanine, cytosine, uracil , carbohydrate ribose and phosphoric acid residues. Unlike DNA, RNA molecules are built from a single polynucleotide chain, which can contain sections that are complementary to each other (Figure 3.5). These regions can interact with each other, forming double helices alternating with non-helical regions.

Figure 3.5. Scheme of the structure of transfer RNA.

Based on their structure and function, there are three main types of RNA:

1) messenger RNA (mRNA) transmit information about the structure of the protein from the cell nucleus to ribosomes;

2) transfer RNAs (tRNAs) transport amino acids to the site of protein synthesis;

3) ribosomal RNA (rRNA) are part of ribosomes and participate in protein synthesis.

8. Biosynthesis of DNA (replication) and mRNA (transcription). Processes of “maturation” of the primary transcript during the formation of mRNA.

Matrix biosynthesis- the process of assembling new macromolecules from monomers, the sequence of which is programmed using nucleic acids. Molecules used as programs in template biosynthesis are called matrices.

The three main template biosyntheses inherent in all living organisms without exception are DNA replication, transcription and translation.

• DNA replication occurs in the nucleus, precedes cell division, resulting in daughter cells receiving a full set of genes;
• transcription also occurs in the nucleus, during which matrix, transport and ribosomal RNAs are formed, which are involved in protein synthesis in the cell;
• broadcast occurs on ribosomes and leads to the formation of specific cellular proteins.

The connection between these processes is reflected in basic postulate of molecular biology: direction of information flow from genotype to phenotype: DNA → RNA → protein(arrows indicate the direction of information transfer).

4.3.2. In addition, some types of viruses are characterized by two more types of matrix syntheses:

• RNA replication - RNA synthesis on an RNA template;
• reverse transcription - DNA synthesis using RNA molecules as a template.

4.3.3. Let us try to formulate general patterns that are characteristic of all matrix biosyntheses.

1. Monomers (nucleotides, amino acids) cannot directly participate in the synthesis of polymers; they must be in active form - nucleotides - in the form of nucleoside triphosphates, amino acids - in the form of compounds with tRNA.
2. The synthesis of all polynucleotide and polypeptide chains consists of three main stages- initiation, elongation and termination.
3. The matrix has special signal or a group of signals allowing identification of a coding element, where does it start information about the synthesized biopolymer chain. This signal, as a rule, does not coincide with the physical start point of the polymer chain of the matrix. Initiation is the process in which the first monomer unit is attached to the matrix molecule.
4. For each act of initiation of biosynthesis, there is a large number of elongation events, i.e. connection of the next monomer with the growing chain. Three components are involved in elongation: a) the terminal group of the synthesized polymer, b) the coding element of the matrix, c) the next molecule of the active monomer. All of them must be fixed in a certain way in the active center of the enzyme or ribosome.
5. Each act of elongation begins with substrate selection by searching through all the substrates present in the system. The entry of the desired substrate into the active center is a signal for the enzymatic reaction to occur. connection of the monomer fragment with the end of the synthesized polymer chain. The addition of a monomer to a growing chain serves as a signal for moving the active center to one coding element of the matrix.
6. The end of the product most often does not correspond to the end of the matrix; there should be a special signal that ensures that the chain stops growing, i.e. termination.
7. The synthesis of a biologically active molecule is usually does not end with termination. The resulting polymer undergoes a number of transformations, such as partial hydrolysis and combination of several chains into one, modification of monomers in the polymer, attachment of a prosthetic part (to a polypeptide) or an apoprotein (to a polynucleotide).

Replication- the process of DNA self-duplication, or the biosynthesis of a daughter DNA molecule that is completely identical to the original molecule (matrix). Localization of the process is the cell nucleus. Basic principles of DNA replication:

• complementarity
• antiparallelism
• unipolarity
• need for priming- enzymes that synthesize DNA are only capable of extending the existing polynucleotide chain, so first a short RNA chain (seed or primer) is synthesized, to which deoxyribonucleotides are attached; the RNA primer that has fulfilled its role is removed;
• intermittency- one of the daughter strands (leading) grows continuously during the replication process, and the other (lagging) grows in the form of fragments several hundred nucleotides long (Okazaki fragments);
• semi-conservative- as a result of replication, two double daughter DNAs are formed, each of which retains (preserves) unchanged one of the halves of the maternal DNA.

4.4.2. Conditions required for DNA replication:

1) Matrix - DNA molecule (Figure 26.1, a);

2) Unraveling squirrels - break hydrogen bonds between the complementary bases of the DNA double helix, resulting in the formation of a replication fork (Figure 26.1, b);

3) DNA binding proteins- attach to separated DNA strands and prevent their reunification;

4) Primase (RNA polymerase)- an enzyme that synthesizes seed RNA.

5) - deoxyribonucleoside triphosphates (dATP, dGTP, dTTP, dCTP). They attach to the nitrogenous bases of polynucleotide chains using hydrogen bonds according to the principle of complementarity;

6) DNA polymerase - an enzyme that forms new polynucleotide chains from nucleoside triphosphates due to the formation of 3’,5’-phosphodiester bonds. The source of energy is the high-energy bonds of nucleoside triphosphates. On one branch of the replication fork, a continuous chain is synthesized, on the other - Okazaki fragments (Figure 26.1, c);

7) DNA ligase - an enzyme that connects Okazaki fragments into a single chain (Figure 26.1, d).

As a result, two identical DNA molecules are formed (Figure 26.1, e).

Transcription- RNA biosynthesis on a DNA matrix. The process of transcription also occurs in the cell nucleus. Basic principles of transcription:

• complementarity- the synthesized chains are complementary to the matrix;
• antiparallelism- the 5" end of the synthesized polynucleotide chain is opposite the 3" end of the template and vice versa;
• unipolarity- the synthesis of polynucleotide chains always occurs in the direction 5" → 3";
• bareness- RNA biosynthesis does not require a primer;
• asymmetry- the synthesis of the daughter chain occurs only on one strand of the DNA template, while the second is blocked.

4.5.2. Conditions required for transcription:

• Matrix- a section of one of the DNA chains (Figure 8.2, a);
• DNA-dependent RNA polymerase- the main enzyme involved in transcription. The place where the enzyme attaches to DNA is the promoter;
• Substrates and energy sources- ribonucleoside triphosphates (ATP, GTP, UTP, CTP). They bind to the nitrogenous bases of the transcribed DNA chain by hydrogen bonds according to the principle of complementarity.

9. Biosynthesis of proteins. Genetic code. The sequence of reactions during the synthesis of a polypeptide chain (initiation, elongation, termination) during translation on ribosomes. Post-translational modification of protein molecules. Protein synthesis disorders in childhood (kwashiorkor) .

Broadcast(from English translation- translation) - translation genetic information enclosed in mRNA, a linear sequence of amino acids in a polypeptide chain. This translation is carried out through the genetic (biological) code. 5.1.2.Genetic code - a sequence of nucleotides corresponding to certain amino acids. The genetic code is characterized by the following properties: triplet code - each amino acid corresponds to a triple ( triplet) nucleotides - codon . There are 43 = 64 codons in total. Of these, 61 are semantic (that is, encode a specific amino acid) and 3 are meaningless (terminating); non-overlapping code - the same DNA or RNA nucleotide cannot belong to two adjacent codons at the same time; continuous code - there are no “punctuation marks” or insertions between codons in the polynucleotide chain; degenerate code (multiple) - some amino acids can be encoded by more than one triplet of nucleotides (since there are 61 codons and 20 amino acids); universal code - the meaning of codons is the same for organisms of all species. 5.1.3. Amino acids and the nucleotide triplets that encode them are not complementary to each other. Therefore, there must be adapter molecules, each of which can interact with both a specific codon and the corresponding amino acid. Such molecules are transfer RNAs(Figure 8.3). Each tRNA contains a triplet of nucleotides - anticodon , which is complementary to a strictly defined mRNA codon. The 3' end of tRNA (acceptor region) is the site of attachment of the amino acid corresponding to the codon of the mRNA.

Amino acid activation - preparatory stage protein biosynthesis - involves binding them to specific tRNAs with the participation of an enzyme aminoacyl-tRNA synthetases. The reaction occurs in the cytoplasm of cells.

The translation process itself includes 3 stages - initiation, elongation, termination and occurs on ribosomes.

Each ribosome consists of a large and small subunit (40S and 60S) and contains an aminoacyl (A) and peptidyl (P) region. The peptidyl site binds the initiating aminoacyl-tRNA, all other aminoacyl-tRNAs are attached to the aminoacyl site.

1) Initiation stage - start of the broadcast. Conditions required for initiation:

• mRNA start codon (AUG);
• protein initiation factors;
• small and large ribosomal subunits;
• GTP (energy source for the closure of ribosomal subparticles);
• magnesium ions;
• initiating aminoacyl-tRNA (methionyl-tRNA) - binds with its anticodon to the initiating codon of mRNA in the peptidyl region of the ribosome.

As a result, initiation complex : mRNA - ribosome - methionyl-tRNA (Figure 5.3, a).

2) Elongation stage - lengthening of the polypeptide chain by 1 amino acid residue occurs in three steps:

• attachment to the initiating complex of aminoacyl-tRNA corresponding to the codon located in the aminoacyl region of the ribosome (Figure 5.3, b);
• transpeptidation - the formation of a peptide bond between amino acid residues (Figure 5.3, c). Energy source - GTP;
• translocation - movement of the ribosome relative to the mRNA by 1 triplet (Figure 5.3, d). The energy source is GTP. Protein factors take part in elongation.

The described process is repeated many times (according to the number of amino acids in the chain).

3) Termination stage - end of broadcast. It is ensured by the presence in the mRNA chain of one of the termination (nonsense) codons - UAA, UGA or UAG. Protein termination factors are involved in the release of the polypeptide (Figure 5.3e). When one of the nonsense codons appears in the aminoacyl region, termination factors stimulate the hydrolase activity of peptidyl transferase. Due to this, the bond between tRNA and peptide is hydrolyzed. GTP is not required for this reaction. After this, the peptide chain, tRNA and mRNA leave the ribosome, and its subparticles dissociate.

Thus, translation of mRNA leads to the formation of a peptide chain with a strictly defined sequence of amino acid residues. The next stage of protein formation is folding, i.e. folding of the peptide chain into a regular three-dimensional structure. If a protein consists of several subunits, then folding also includes combining them into a single macromolecule.

It is believed that small protein molecules containing about 100 aminoacyl residues can independently adopt a three-dimensional structure; folding of larger polypeptide chains requires the participation of special proteins - chaperones.

Chaperones are called differently heat shock proteins since they not only ensure the correct folding of newly formed proteins, but also the renaturation of previously synthesized proteins that have undergone partial denaturation in the cell under the influence of various factors(overheating, irradiation, action of free radicals, etc.).

5.2.2. Post-translational modifications protein molecules may include:

• partial proteolysis (for example, the conversion of a proenzyme to an enzyme);
• addition of a prosthetic group (phosphoric acid residues, carbohydrate residues, heme groups, etc.);
• modifications of side chains of amino acid residues:
  • hydroxylation of proline to hydroxyproline in collagen,
  • arginine methylation in histone,
  • iodination of tyrosine in thyroglobulin).

5.2.3. The effect of toxic and medicinal substances on protein biosynthesis. Protein biosynthesis is one of the most complex processes occurring in cells. Its interruption or perversion is possible as a result of disruption of any of the three matrix syntheses.
Thus, mutagens (benz(a)pyrene, lindane) disrupt DNA replication and thus interrupt protein synthesizing processes.
Some toxic substances (gossypol) can change the rate of transcription.
Drugs that affect protein biosynthesis include antibiotics and interferons.
Antibiotics that block matrix biosynthesis are used in the treatment of infectious diseases and malignant tumors. (see table 5.1).

Table 5.1

Antibiotics that inhibit matrix biosynthesis

10. Regulation of protein synthesis. Introduction to the operon. Induction and repression of synthesis in the human body. The role of hormones in the regulation of gene action. Inhibitors of matrix synthesis - antibiotics, interferons.

5.3.1. Operon (transcripton)- a set of genes that can be turned on and off depending on the metabolic needs of the cell. The composition of the operon, along with structural genes (SG) , encoding the structure of certain proteins, includes DNA sections that perform regulatory functions (Figure 5.4). A group of structural genes responsible for the synthesis of enzymes of one metabolic pathway is under the control gene operator (GO) located nearby. The function of the operator gene is controlled by something spatially distant from it gene regulator (GR) , which produces repressor protein , in active or inactive form. An active repressor protein is able to bind to the operator gene and inhibit the transcription of structural genes, therefore, suppress protein synthesis. Substances that cause inactivation of the repressor protein are inductors protein synthesis, which have the opposite effect - corepressors. The initial substrates of metabolic pathways can act as inducers, and the end products of these pathways can act as corepressors. 5.3.2. There are two mechanisms for regulating protein synthesis - induction and repression . An example of an operon that is regulated by an inductive mechanism is lactose operon , which, along with the operator gene, includes 3 structural genes encoding enzymes for lactose catabolism (see Figure 5.4). Lactose is the inducer of this operon. At a high concentration of lactose in the medium, enzymes are synthesized, but at a low concentration - not.

5.3.3. According to the mechanism of repression it is regulated histidine operon , containing an operator gene and 10 structural genes encoding enzymes necessary for the biosynthesis of histidine (see Figure 5.5). Histidine is a corepressor of this operon. At a high concentration of histidine in the medium, the synthesis of enzymes stops; in the absence of histidine, they are synthesized.

11. The role of enzymes in metabolism. Hereditary enzymopathies V early childhood . Variety of enzymes. Specificity of enzyme action. Classification of enzymes. Isoenzymes, multienzymes.

The course of metabolic processes in the body is determined by the action of numerous enzymes - biological catalysts of protein nature. They speed up chemical reactions without being consumed. Term "enzyme" comes from the Latin word fermentum - sourdough. Along with this concept, the equivalent term is used in the literature "enzyme" (en zyme - in yeast) of Greek origin. Hence the branch of biochemistry that studies enzymes is called “enzymology”.

Enzymology forms the basis for knowledge at the molecular level of the most important problems of human physiology and pathology. Digestion of nutrients and their use for energy production, formation of structural and functional components of tissues, muscle contraction, transmission of electrical signals along nerve fibers, perception of light by the eye, blood clotting - each of these physiological mechanisms is based on the catalytic action of certain enzymes. Numerous diseases have been shown to directly impair enzymatic catalysis; determination of enzyme activity in blood and other tissues provides valuable information for medical diagnostics; enzymes or their inhibitors can be used as medicinal substances. Thus, knowledge the most important features enzymes and the reactions they catalyze are necessary for a rational approach to the study of human diseases, their diagnosis and treatment.

The classification is based on the most important sign The way one enzyme differs from another is the reaction it catalyzes. The number of types of chemical reactions is relatively small, which made it possible to divide all currently known enzymes into 6 most important classes, depending on the type of reaction being catalyzed. These classes are:

• oxidoreductases (redox reactions);
• transferases (transfer of functional groups);
• hydrolases (cleavage reactions involving water);
• lyases (breaking bonds without the participation of water);
• isomerases (isomeric transformations);
• ligases (synthesis with the consumption of ATP molecules).

7.4.3. Enzymes of each class are divided into subclasses, guided by the structure of the substrates. Subclasses combine enzymes that act on similarly constructed substrates. Subclasses are divided into subclasses, V which further refine the structure of chemical groups that distinguish substrates from each other. Within the subclasses they list individual enzymes. All classification divisions have their own numbers. Thus, any enzyme receives its own unique code number, consisting of four numbers separated by dots. The first number indicates the class, the second the subclass, the third the subsubclass, and the fourth the number of the enzyme within the subclass. For example, the enzyme α-amylase, which breaks down starch, is designated as 3.2.1.1, where:
3 — type of reaction (hydrolysis);
2 - type of bond in the substrate (glycosidic);
1 - type of bond (O-glycosidic);
1 - enzyme number in the subclass

The above-described decimal numbering method has one important advantage: it allows one to bypass the main inconvenience of continuous numbering of enzymes, namely: the need to change the numbers of all subsequent ones when including a newly discovered enzyme in the list. A new enzyme may be placed at the end of the corresponding subclass without disturbing the rest of the numbering. Likewise, when new classes, subclasses and sub-subclasses are identified, they can be added without disturbing the numbering order of previously established divisions. If after receiving new information It becomes necessary to change the numbers of some enzymes; the previous numbers are not assigned to new enzymes in order to avoid misunderstandings.

Speaking about the classification of enzymes, it should also be noted that enzymes are classified not as individual substances, but as catalysts for certain chemical transformations. Enzymes isolated from different biological sources and catalyzing identical reactions can differ significantly in their primary structure. However, in the classification list they all appear under the same code number.

So, knowing the enzyme code number allows you to:

• eliminate ambiguities if different researchers use the same name for different enzymes;
• make searching for information in literary databases more efficient;
• obtain additional information about the amino acid sequence, spatial structure of the enzyme, and genes encoding enzyme proteins from other databases.

Isoenzymes - these are multiple forms of one enzyme that catalyze the same reaction, but differ in physical and chemical properties (affinity for the substrate, maximum speed of the catalyzed reaction, electrophoretic mobility, different sensitivity to inhibitors and activators, pH optimum and thermal stability). Isoenzymes have a quaternary structure, which is formed by an even number of subunits (2, 4, 6, etc.). Enzyme isoforms are formed by different combinations of subunits.

As an example, consider lactate dehydrogenase (LDH), an enzyme that catalyzes a reversible reaction:

NADH 2 NAD +

pyruvate ← LDH → lactate

LDH exists in the form of 5 isoforms, each of which consists of 4 protomers (subunits) of 2 types M (muscle) and H (heart). The synthesis of M and H type protomers is encoded by two different genetic loci. LDH isoenzymes differ at the level of quaternary structure: LDH 1 (NNNN), LDH 2 (NNMM), LDH 3 (NNMM), LDH 4 (NMMM), LDH 5 (MMMM).

Polypeptide chains of the H and M types have the same molecular weight, but the former are dominated by carboxylic amino acids, the latter by diamino acids, so they carry different charges and can be separated by electrophoresis.

Oxygen metabolism in tissues affects the isoenzyme composition of LDH. Where aerobic metabolism dominates, LDH 1, LDH 2 predominate (myocardium, adrenal glands), where anaerobic metabolism - LDH 4, LDH 5 (skeletal muscles, liver). In progress individual development The oxygen content and LDH isoforms change in the body's tissues. In the embryo, LDH 4 and LDH 5 predominate. After birth, the content of LDH 1 and LDH 2 increases in some tissues.

The existence of isoforms increases the adaptive capacity of tissues, organs, and the body as a whole to changing conditions. The metabolic state of organs and tissues is assessed by changes in isoenzyme composition.

12. Properties of enzymes. Dependence of the rate of enzymatic reaction on the concentration of enzyme and substrate, temperature and pH of the environment.

The protein nature of enzymes determines the appearance of a number of properties in them that are generally uncharacteristic of inorganic catalysts: oligodynamicity, specificity, dependence of the reaction rate on temperature, pH of the medium, concentration of the enzyme and substrate, the presence of activators and inhibitors.

Under oligodynamism Enzymes are highly effective in very small quantities. This high efficiency is explained by the fact that enzyme molecules continuously regenerate during their catalytic activity. A typical enzyme molecule can regenerate millions of times per minute. It must be said that inorganic catalysts are also capable of accelerating the transformation of a quantity of substances that is many times greater than their own mass. But no inorganic catalyst can compare with enzymes in terms of efficiency.

An example is the enzyme rennin, produced by the gastric mucosa of ruminants. One molecule of it in 10 minutes at 37°C is capable of causing coagulation (curdling) of about a million molecules of milk caseinogen.

Another example of the high efficiency of enzymes is provided by catalase. One molecule of this enzyme at 0°C breaks down about 50,000 molecules of hydrogen peroxide per second:

2 H2O2 2 H2O + O2

The effect of catalase on hydrogen peroxide is to change the activation energy of this reaction from approximately 75 kJ/mol without a catalyst to 21 kJ/mol in the presence of the enzyme. If colloidal platinum is used as a catalyst for this reaction, then the activation energy is only 50 kJ/mol.

7.2.2. When studying the influence of any factor on the rate of an enzymatic reaction, all other factors should remain unchanged and, if possible, have an optimal value.

The rate of enzymatic reactions is measured by the amount of substrate converted per unit of time, or the amount of product formed. The speed change is carried out at the initial stage of the reaction, when the product is still practically absent and the reverse reaction does not occur. In addition, at the initial stage of the reaction, the concentration of the substrate corresponds to its original amount.

7.2.3. Dependence of the rate of enzymatic reaction (V) on enzyme concentration [E](Figure 7.3). At a high substrate concentration (multiple times the enzyme concentration) and other factors remaining constant, the rate of the enzymatic reaction is proportional to the enzyme concentration. Therefore, knowing the rate of the reaction catalyzed by the enzyme, we can draw a conclusion about its amount in the material under study.

Figure 7.3.Dependence of the rate of enzymatic reaction on enzyme concentration

7.2.4. Dependence of reaction rate on substrate concentration [S]. The dependence graph looks like a hyperbola (Figure 7.4). At a constant enzyme concentration, the rate of the catalyzed reaction increases with increasing substrate concentration up to the maximum value Vmax, after which it remains constant. This should be explained by the fact that at high substrate concentrations, all active centers of enzyme molecules are associated with substrate molecules. Any excess substrate can combine with the enzyme only after the reaction product is formed and the active site is freed.

Figure 7.4.Dependence of the rate of enzymatic reaction on the concentration of the substrate.

The dependence of the reaction rate on the substrate concentration can be expressed by the Michaelis-Menten equation:

,

where V is the reaction rate at the substrate concentration [S], Vmax is the maximum speed and KM is the Michaelis constant.

The Michaelis constant is equal to the substrate concentration at which the reaction rate is half the maximum. The definition of KM and Vmax is important practical significance, since it allows you to quantitatively describe most enzymatic reactions, including reactions involving two or more substrates. Different chemicals that alter enzyme activity have different effects on Vmax and KM values.

7.2.5. Dependence of the reaction rate on t - the temperature at which the reaction occurs (Figure 7.5) is complex. The temperature value at which the reaction rate is maximum represents the temperature optimum of the enzyme. The temperature optimum for most enzymes in the human body is approximately 40°C. For most enzymes, the optimal temperature is equal to or higher than the temperature at which the cells are located.

Figure 7.5. Dependence of the rate of enzymatic reaction on temperature.

At lower temperatures (0° - 40°C), the reaction rate increases with increasing temperature. When the temperature rises by 10°C, the rate of the enzymatic reaction doubles (temperature coefficient Q10 is 2). The increase in reaction rate is due to an increase kinetic energy molecules. With a further increase in temperature, the bonds that support the secondary and tertiary structure of the enzyme are broken, that is, thermal denaturation. This is accompanied by a gradual loss of catalytic activity.

7.2.6. Dependence of the reaction rate on the pH of the medium (Figure 7.6). At a constant temperature, the enzyme works most efficiently within a narrow pH range. The pH value at which the reaction rate is maximum represents the optimum pH of the enzyme. Most enzymes in the human body have an optimum pH within the range of pH 6 - 8, but there are enzymes that are active at pH values ​​outside this range (for example, pepsin, which is most active at pH 1.5 - 2.5).

A change in pH, either in the acidic or alkaline direction from the optimum, leads to a change in the degree of ionization of the acidic and basic groups of amino acids that make up the enzyme (for example, COOH groups of aspartate and glutamate, NH2 groups of lysine, etc.). This causes a change in the conformation of the enzyme, resulting in a change in the spatial structure of the active center and a decrease in its affinity for the substrate. In addition, at extreme pH values, the enzyme is denatured and inactivated.

Figure 7.6. Dependence of the rate of enzymatic reaction on the pH of the medium.

It should be noted that the pH optimum characteristic of an enzyme does not always coincide with the pH of its immediate intracellular environment. This suggests that the environment in which the enzyme is located regulates its activity to some extent.

7.2.7. Dependence of the reaction rate on the presence of activators and inhibitors . Activators increase the rate of the enzymatic reaction. Inhibitors reduce the rate of enzymatic reactions.

Inorganic ions can act as enzyme activators. It is believed that these ions cause the enzyme or substrate molecules to adopt a conformation that promotes the formation of an enzyme-substrate complex. This increases the probability of interaction between the enzyme and the substrate, and consequently the rate of the reaction catalyzed by the enzyme. For example, salivary amylase activity increases in the presence of chloride ions.

13. Mechanism of action of enzymes. Catalytic (active) center. Coenzymes and cofactors. Competitive and non-competitive inhibition. Use of competitive inhibitors as drugs.

Active center (Ac) is a part of the enzyme molecule that specifically interacts with the substrate and is directly involved in catalysis. Ats, as a rule, is located in a niche (pocket). Two regions can be distinguished in Ac: the substrate binding site - substrate area (contact pad) and actually catalytic center .

Most substrates form at least three bonds with the enzyme, due to which the substrate molecule attaches to the active site in the only possible way, which ensures the substrate specificity of the enzyme. The catalytic center provides the choice of the chemical transformation path and the catalytic specificity of the enzyme.

A group of regulatory enzymes has allosteric centers , which are located outside the active center. “+” or “-” modulators that regulate enzyme activity can be attached to the allosteric center.

There are simple enzymes, consisting only of amino acids, and complex enzymes, which also include low molecular weight organic compounds of a non-protein nature (coenzymes) and (or) metal ions (cofactors).

Coenzymes - This organic matter non-protein nature, taking part in catalysis as part of the catalytic site of the active center. In this case, the protein component is called apoenzyme , and the catalytically active form of a complex protein is holoenzyme . Thus: holoenzyme = apoenzyme + coenzyme.

The following function as coenzymes:

• hemes,
• nucleotides,
• coenzyme Q,
• FAFS,
• Glutathione
• derivatives of water-soluble vitamins:

A coenzyme that is attached to the protein part by covalent bonds is called prosthetic group . These are, for example, FAD, FMN, biotin, lipoic acid. The prosthetic group is not separated from the protein part. A coenzyme that is attached to the protein part by non-covalent bonds is called cosubstrate . These are, for example, NAD +, NADP +. The cosubstrate attaches to the enzyme at the time of reaction.

Enzyme cofactors are metal ions necessary for the catalytic activity of many enzymes. Potassium, magnesium, calcium, zinc, copper, iron, etc. ions act as cofactors. Their role is diverse; they stabilize substrate molecules, the active center of the enzyme, its tertiary and quaternary structure, and ensure substrate binding and catalysis. For example, ATP binds to kinases only in conjunction with Mg 2+ .

14. Basic mechanisms for regulating the action of enzymes and their role in the regulation of metabolism. Proenzymes.

8.4.1. As already noted, enzymes are catalysts whose activity can be regulated. Therefore, through enzymes it is possible to control the speed of chemical reactions in the body. Regulation of enzyme activity can be carried out by interaction with them of various biological components or foreign compounds (for example, drugs and poisons), which are commonly called modifiers or enzyme regulators. Under the influence of modifiers on the enzyme, the reaction can be accelerated (in this case they are called activators) or slow down (in this case they are called inhibitors).

8.4.2. Enzyme activation is determined by the acceleration of biochemical reactions that occurs after the action of the modifier. One group of activators consists of substances that affect the region of the active center of the enzyme. These include enzyme cofactors and substrates. Cofactors (metal ions and coenzymes) are not only obligatory structural elements of complex enzymes, but also essentially their activators.

Of the metal ions, the activity of many enzymes is affected by: NH4+, Na+, Mg2+, K+, Ca2+, Mn2+, Zn2+, Fe2+, Fe3+, Co2+. Heavy metal ions, as a rule, have an inhibitory effect. The actions of cations are generally quite specific, but in most cases the enzyme is activated by more than one cation. The phenomenon of antagonism between ions is also observed. The best known is the antagonism between Na+ and K+ and between Mg2+ and Ca2+.

Magnesium is a natural activator of enzymes acting on phosphorylated substrates (phosphatases, kinases, synthetases), but under in vitro conditions it can be replaced by manganese.

Anions generally have little effect on enzyme activity, and their effects lack specificity. The exception is amylase, which is activated by chlorides and, to a lesser extent, by other halogens. The effect of the activating ion also varies depending on pH. The degree of enzyme purification also affects the activating ion concentration and the specificity of activation. Highly purified enzymes are characterized by greater selectivity towards activating ions.

8.4.3. The activating effect of metal ions is realized in various ways. The most typical mechanism is the inclusion of an ion in the structure of the catalytic center of the enzyme, which is inactive without it. This is a typical function of a metal as a coenzyme. Another fairly common function of an activating metal is to form a bond between an enzyme and a substrate, or between an enzyme, a coenzyme and a substrate. For example, Zn2+ ions in the alcohol dehydrogenase enzyme form 2 coordination bonds with the NAD+ coenzyme molecule, 3 coordination bonds with the apoenzyme molecule, and the sixth coordination bond attaches the substrate.

Metal ions, as well as substrates, coenzymes, their precursors and structural analogs, can be used in practice as drugs that regulate enzyme activity.

While the catalytically active protein is called an enzyme (or enzyme), the inactive enzyme precursor is called a proenzyme (or zymogen).

Activation of proteins by partial proteolysis is a process widespread in biological systems. Here are some examples.

• digestive enzymes that hydrolyze proteins are synthesized in the stomach and pancreas in the form of proenzymes: pepsin - in the form of pepsinogen, trypsin - in the form of trypsinogen, etc.
• Blood coagulation is a cascade of reactions of proteolytic activation of proenzymes. This provides a quick response to blood vessel damage.
• some protein hormones are synthesized as inactive precursors. For example, insulin is formed from proinsulin.
• fibrillar connective tissue protein collagen is also formed from a precursor - procollagen.

15. Principles of quantitative determination of enzymes. Units of enzyme activity. Main directions of use of enzymes in medicine. Enzymodiagnostics, enzyme therapy, use of enzymes as reagents.

The unique property of enzymes to accelerate chemical reactions can be used to quantify the content of these biocatalysts in biological material (tissue extract, blood serum, etc.). Under correctly selected experimental conditions, there is almost always a proportionality between the amount of enzyme and the rate of the catalyzed reaction, therefore, by the activity of the enzyme, one can judge its quantitative content in the test sample.

Enzyme activity measurement is based on speed comparison chemical reaction in the presence of an active biocatalyst with the reaction rate in a control solution in which the enzyme is absent or inactivated.

The material under study is placed in an incubation environment where optimal temperature, pH, concentrations of activators and substrates are created. At the same time, a control sample is carried out, to which the enzyme is not added. After some time, the reaction is stopped by adding various reagents (changing the pH of the medium, causing denaturation of proteins, etc.) and analyzing the samples.

In order to determine the rate of an enzymatic reaction, you need to know:

• the difference in concentrations of the substrate or reaction product before and after incubation;
• incubation time;
• amount of material taken for analysis.

Most often, enzyme activity is assessed by the amount of reaction product formed. This is done, for example, when determining the activity of alanine aminotransferase, which catalyzes the following reaction:

Enzyme activity can also be calculated based on the amount of substrate consumed. An example is a method for determining the activity of α-amylase, an enzyme that breaks down starch. By measuring the starch content in the sample before and after incubation and calculating the difference, the amount of substrate broken down during incubation is found.

There are a large number of methods for measuring enzyme activity, differing in technique, specificity, and sensitivity.

Most often used to determine photoelectrocolorimetric methods . These methods are based on color reactions with one of the products of enzyme action. In this case, the color intensity of the resulting solutions (measured on a photoelectrocolorimeter) is proportional to the amount of the product formed. For example, during reactions catalyzed by aminotransferases, α-keto acids accumulate, which give red-brown compounds with 2,4-dinitrophenylhydrazine:

If the biocatalyst under study has low specificity of action, then it is possible to select a substrate whose reaction results in the formation of a colored product. An example is the determination of alkaline phosphatase, an enzyme widely distributed in human tissues; its activity in blood plasma changes significantly in diseases of the liver and skeletal system. This enzyme, in an alkaline environment, hydrolyzes a large group of phosphate esters, both natural and synthetic. One of the synthetic substrates is paranitrophenyl phosphate (colorless), which in an alkaline environment breaks down into orthophosphate and paranitrophenol (yellow).

The progress of the reaction can be monitored by measuring the gradually increasing color intensity of the solution:

For enzymes with high specificity of action, such a selection of substrates is usually impossible.

Spectrophotometric methods are based on changes in the ultraviolet spectrum of chemicals taking part in the reaction. Most compounds absorb ultraviolet rays, and the absorbed wavelengths are characteristic of certain groups of atoms present in the molecules of these substances. Enzymatic reactions cause intramolecular rearrangements, as a result of which the ultraviolet spectrum changes. These changes can be recorded on a spectrophotometer.

Spectrophotometric methods, for example, determine the activity of redox enzymes containing NAD or NADP as coenzymes. These coenzymes act as acceptors or donors of hydrogen atoms and are thus either reduced or oxidized during metabolic processes. Reduced forms of these coenzymes have an ultraviolet spectrum with an absorption maximum at 340 nm; oxidized forms do not have this maximum. Thus, when lactate dehydrogenase acts on lactic acid, hydrogen is transferred to NAD, which leads to an increase in the absorption of NADH at 340 nm. The magnitude of this absorption in optical units is proportional to the amount of reduced form of the coenzyme formed.

By changing the content of the reduced form of the coenzyme, the activity of the enzyme can be determined.

Fluorimetric methods. These methods are based on the phenomenon of fluorescence, which consists in the fact that the object under study, under the influence of radiation, emits light with a shorter wavelength. Fluorimetric methods for determining enzyme activity are more sensitive than spectrophotometric methods. Relatively new and even more sensitive are chemiluminescent methods using the luciferin-luciferase system. Such methods make it possible to determine the rate of reactions that occur with the formation of ATP. When luciferin (a carboxylic acid of complex structure) interacts with ATP, luciferyl adenylate is formed. This compound is oxidized with the participation of the enzyme luciferase, which is accompanied by a flash of light. By measuring the intensity of light flashes, it is possible to determine amounts of ATP on the order of several picomoles (10-12 mol).

Titrometric methods . A number of enzymatic reactions are accompanied by a change in the pH of the incubation mixture. An example of such an enzyme is pancreatic lipase. Lipase catalyzes the reaction:

The resulting fatty acids can be titrated, and the amount of alkali used for titration will be proportional to the amount of fatty acids released and, therefore, to the lipase activity. Determination of the activity of this enzyme is of clinical importance.

Manometric methods are based on measuring in a closed reaction vessel the volume of gas released (or absorbed) during an enzymatic reaction. Using such methods, the oxidative decarboxylation reactions of pyruvic and α-ketoglutaric acids, which proceed with the release of CO2, were discovered and studied. Currently, these methods are rarely used.

The International Enzyme Commission has proposed unit of activity of any enzyme, take such an amount of enzyme that when given conditions catalyzes the conversion of one micromole (10-6 mol) of substrate per unit of time (1 min, 1 hour) or one microequivalent of the affected group in cases where more than one group in each substrate molecule is attacked (proteins, polysaccharides and others). The temperature at which the reaction is carried out must be indicated. Enzyme activity measurements can be expressed in units general, specific and molecular activity.

For a unit total enzyme activity based on the amount of material taken for research. Thus, the activity of alanine aminotransferase in the liver of rats is 1670 μmol of pyruvate per hour per 1 g of tissue; Cholinesterase activity in human serum is 250 µmol acetic acid per hour per 1 ml of serum at 37°C.

High values ​​of enzyme activity both in normal and pathological conditions require special attention of the researcher. It is recommended to work with low levels of enzyme activity. To do this, the enzyme source is taken in smaller quantities (the serum is diluted several times with physiological solution, and a smaller percentage homogenate is prepared for the tissue). In this case, conditions for saturation with the substrate are created in relation to the enzyme, which contributes to the manifestation of its true activity.

Total enzyme activity is calculated using the formula:

Where A- enzyme activity (total), ΔС- difference in substrate concentrations before and after incubation; IN- amount of material taken for analysis, t- incubation time; n- breeding.

It should be borne in mind that indicators of the activity of enzymes in blood serum and urine, studied for diagnostic purposes, are expressed in units of total activity.

Since enzymes are proteins, it is important to know not only the overall enzyme activity in the material being tested, but also the enzymatic activity of the protein present in the sample. For a unit specific activity take an amount of enzyme that catalyzes the conversion of 1 µmol of substrate per unit time per 1 mg of sample protein. To calculate the specific activity of the enzyme, it is necessary to divide the total activity by the protein content in the sample:

The worse the enzyme is purified, the more extraneous ballast proteins are in the sample, the lower the specific activity. During purification, the amount of such proteins decreases, and accordingly, the specific activity of the enzyme increases. Suppose that in the original biological material that is the source of the enzyme (chopped liver, pulp from plant tissue), the specific activity was equal to 0.5 µmol/(mg protein × min). After fractional precipitation with ammonium sulfate and gel filtration through Sephadex, it increased to 25 µmol/(mg protein x min), i.e. increased 50 times. Evaluating the efficiency of purification of enzyme preparations is used in the production of medicines of an enzymatic nature.

Specific activity is determined when it is necessary to compare the activity of different preparations of the same enzyme. If it is necessary to compare the activity of different enzymes, molecular activity is calculated.

Molecular activity (or enzyme turnover number) is the number of moles of substrate that are converted by 1 mole of enzyme per unit of time (usually 1 minute). Different enzymes have different molecular activities. A decrease in the number of enzyme turnover occurs under the influence of non-competitive inhibitors. By changing the conformation of the catalytic center of the enzyme, these substances reduce the affinity of the enzyme for the substrate, which leads to a decrease in the number of substrate molecules reacting with one enzyme molecule per unit time.

16. Nutrition is an integral part of metabolism. The main components of the diet and their role. Replaceable and irreplaceable components of the diet. Balanced diet. Consequences of unbalanced nutrition in children .

Full-fledged is a diet that meets a person’s energy needs and contains the required amount of essential nutrients to ensure normal growth and development of the body.

Factors influencing the body's need for energy and nutrients: gender, age and body weight of a person, his physical activity, climatic conditions, biochemical, immunological and morphological characteristics of the body.

All nutrients can be divided into five classes:

1. proteins; 2. fats; 3. carbohydrates; 4. vitamins; 5. minerals.

In addition, any diet must contain water, as a universal solvent.

The essential components of the diet are:

1. essential amino acids - valine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan;
2. essential fatty acids - linoleic, linolenic, arachidonic;
3. water- and fat-soluble vitamins;
4. inorganic (mineral) elements - calcium, potassium, sodium, chlorine, copper, iron, chromium, fluorine, iodine and others.

11.1.2. Balanced diet. A diet containing nutrients in a ratio optimal for maximum satisfaction of the plastic and energy needs of the human body is called balanced diet. It is believed that the most favorable ratio of proteins, fats and carbohydrates is close to 1:1:4, provided that the total calorie content of the diet corresponds to the energy expenditure of a given person. So, for a male student weighing 60 kg, energy consumption is on average 2900 kcal per day and the diet should contain: 80-100 g of proteins, 90 g of fats, 300 - 400 g of carbohydrates.

17. Biological value of food proteins. Quantity and quality of proteins in human nutrition. Replaceable and essential amino acids. Combination food products, mutually complementary in amino acid composition. Characteristics of the protein diet of children. Consequences of insufficient protein nutrition in children.

Biological role of food proteins is that they serve as a source of irreplaceable and replaceable amino acids. Amino acids are used by the body to synthesize its own proteins; as precursors of non-protein nitrogenous substances (hormones, purines, porphyrins, etc.); as a source of energy (oxidation of 1 g of proteins provides approximately 4 kcal of energy).

Food proteins are divided into complete and incomplete.

Complete food proteins - of animal origin, contain all amino acids in the required proportions and are well absorbed by the body.

Incomplete proteins - of plant origin, do not contain, or contain insufficient amounts of one or more essential amino acids. Thus, grain crops are deficient in lysine, methionine, and threonine; Potato protein contains little methionine and cysteine. To obtain protein-rich diets, you should combine plant proteins that complement each other in amino acid composition, for example, corn and beans.

Daily requirement: at least 50 g per day, on average 80-100 g.

11.2.2. Protein deficiency in childhood causes: 1. decreased body resistance to infections; 2. growth arrest due to impaired synthesis of growth factors; 3. energy deficiency of the body (depletion of carbohydrate and fat depots, catabolism of tissue proteins); 4. loss of body weight - malnutrition. During protein starvation, edema is observed, which occurs due to a decrease in protein content in the blood ( hypoalbuminemia) and disturbances in the distribution of water between blood and tissues.

18. Digestion of proteins. Proteinases. Mechanism of activation of gastrointestinal proteinases . Specificity (selectivity) of hydrolysis of peptide bonds. Features of protein digestion in infants, protein digestion disorders in children . Rotting of proteins (amino acids) in the large intestine.

The digestion of proteins, that is, their breakdown into individual amino acids, begins in the stomach and ends in the small intestine. Digestion occurs under the action of gastric, pancreatic and intestinal juices, which contain proteolytic enzymes (proteases or peptidases). Proteolytic enzymes belong to the class of hydrolases. They catalyze the hydrolysis of peptide bonds CO—NH protein molecule.

All proteolytic enzymes can be divided into two groups:

1. exopeptidases- catalyze the rupture of the terminal peptide bond with the release of the N- or C-terminal amino acid;
2. endopeptidases- hydrolyze peptide bonds within the polypeptide chain, the reaction products are peptides with a lower molecular weight.

10.1.3. Most proteolytic enzymes involved in the digestion of proteins and peptides are synthesized and secreted into the cavity of the digestive tract in the form of inactive precursors - proenzymes (zymogens). Therefore, the proteins of the cells producing proenzymes are not digested. Activation of proenzymes occurs in the lumen of the gastrointestinal tract through partial proteolysis - the cleavage of part of the zymogen peptide chain.

The bulk of amino acids formed in the digestive tract as a result of protein digestion are absorbed into the blood and replenish the body’s amino acid pool. A certain amount of unabsorbed amino acids undergo decay in the large intestine.

Rotting - transformations of amino acids caused by the activity of microorganisms in the large intestine. Strengthening the processes of decay of amino acids can be facilitated by:

• excess intake of proteins from food;
• congenital and acquired disorders of the absorption of amino acids in the intestine;
• decreased intestinal motor function.

As a result of the decay of amino acids, various substances are formed, many of which are toxic to the body. Some examples of decay products are given in Table 10.2.

Table 10.2
Products of decay of amino acids in the intestines.

10.2.2. The products of amino acid decay are xenobiotics- substances that are foreign to the human body and must be neutralized (inactivated).

19. The role of lipids in the body. Dietary lipids, daily requirements for children of different ages. Features of the use of lipids in various tissues. Brown adipose tissue. Deposition and mobilization of fats in adipose tissue. Obesity.

The composition of dietary fats consists mainly of triacylglycerols (98%), phospholipids and cholesterol. Triacylglycerols of animal origin contain many saturated fatty acids and have a solid consistency. Vegetable fats contain more unsaturated fatty acids and have a liquid consistency (oils).

Biological role: 1. are one of the main sources of energy; 2. serve as a source of essential polyunsaturated fatty acids; 3. promote the absorption of fat-soluble vitamins from the intestines. Polyunsaturated fatty acids necessary for the body to build phospholipids, which form the basis of all cell membrane structures and blood lipoproteins. In addition, linoleic acid is used for the synthesis of arachidonic acid, which serves as a precursor to prostaglandins, prostacyclins, thromboxanes and leukotrienes.

Daily requirement: 90-100 g, of which 30% should be vegetable oils. The nutritional value of vegetable fats is higher than that of animal fats, since with the same energy effect - 9 kcal per 1 g, they contain more essential fatty acids.

11.3.2. Violation of the ratio of the proportion of plant and animal fats in the diet leads to a change in the ratio of various classes of lipoproteins in the blood and, as a consequence, to coronary heart disease and atherosclerosis.

20. Digestion of fats. Lipases and phospholipases. Bile acids and paired bile acids: structure, formation, biological role. Features of lipid digestion in children . Lipid digestion disorders.

The main site of lipid digestion is the upper small intestine. The following conditions are necessary for the digestion of lipids:

• presence of lipolytic enzymes;
• conditions for lipid emulsification;
• optimal pH values ​​of the environment (within 5.5 - 7.5).

10.3.2. Various enzymes are involved in the breakdown of lipids. Dietary fats in an adult are broken down mainly by pancreatic lipase; Lipase is also found in intestinal juice and saliva; in infants, lipase is active in the stomach. Lipases belong to the class of hydrolases; they hydrolyze ester bonds -O-SO- with the formation of free fatty acids, diacylglycerols, monoacylglycerols, glycerol

Glycerophospholipids supplied with food are exposed to specific hydrolases - phospholipases, which cleave ester bonds between the components of phospholipids. The specificity of the action of phospholipases is shown in Figure 10.4.

Figure 10.4. Specificity of the action of enzymes that break down phospholipids.

The products of phospholipid hydrolysis are fatty acids, glycerol, inorganic phosphate, nitrogenous bases (choline, ethanolamine, serine).

Dietary cholesterol esters are hydrolyzed by pancreatic cholesterol esterase to form cholesterol and fatty acids.

10.3.3. Understand the structure of bile acids and their role in the digestion of fats. Bile acids are the end product of cholesterol metabolism and are formed in the liver. These include: cholic (3,7,12-trioxycholanic), chenodeoxycholic (3,7-dioxycholanic) and deoxycholic (3, 12-dioxycholanic) acids (Figure 10.5, a). The first two are primary bile acids (formed directly in hepatocytes), deoxycholic acid is secondary (as it is formed from primary bile acids under the influence of intestinal microflora).

In bile, these acids are present in conjugated form, i.e. in the form of compounds with glycine H2N-CH2-COOH or taurine H2N-CH2-CH2-SO3H(Figure 10.5, b).

Figure 10.5. The structure of unconjugated (a) and conjugated (b) bile acids.

15.1.4. Bile acids have amphiphilic properties: hydroxyl groups and side chain are hydrophilic, cyclic structure is hydrophobic. These properties determine the participation of bile acids in the digestion of lipids:

1) bile acids are capable emulsify fats, their molecules with their non-polar part are adsorbed on the surface of fat droplets, at the same time hydrophilic groups interact with the surrounding aqueous environment. As a result, the surface tension at the interface between the lipid and aqueous phases decreases, as a result of which large fat droplets are broken into smaller ones;

2) bile acids, along with bile colipase, are involved in activation of pancreatic lipase, shifting its pH optimum to the acidic side;

3) bile acids form water-soluble complexes with hydrophobic products of fat digestion, which contributes to their absorption into the wall of the small intestine.

Bile acids, which penetrate into the enterocytes during absorption along with hydrolysis products, enter the liver through the portal system. These acids can be re-secreted with bile into the intestines and participate in the processes of digestion and absorption. Such enterohepatic circulation bile acids can be carried out up to 10 or more times a day.

The name “squirrels” comes from the ability of many of them to turn white when heated. The name "proteins" comes from the Greek word for "first", indicating their importance in the body. The higher the level of organization of living beings, the more diverse the composition of proteins.

Proteins are formed from amino acids, which are linked together by covalent bonds. peptide bond: between the carboxyl group of one amino acid and the amino group of another. When two amino acids interact, a dipeptide is formed (from the residues of two amino acids, from the Greek. peptos– cooked). Replacement, exclusion or rearrangement of amino acids in a polypeptide chain causes the emergence of new proteins. For example, when replacing only one amino acid (glutamine with valine), a serious disease occurs - sickle cell anemia, when red blood cells have a different shape and cannot perform their main functions (oxygen transport). When a peptide bond is formed, a water molecule is split off. Depending on the number of amino acid residues, they are distinguished:

– oligopeptides (di-, tri-, tetrapeptides, etc.) – contain up to 20 amino acid residues;

– polypeptides – from 20 to 50 amino acid residues;

– squirrels – over 50, sometimes thousands of amino acid residues

By physical and chemical properties distinguish between hydrophilic and hydrophobic proteins.

There are four levels of organization of the protein molecule - equivalent spatial structures (configurations, conformation) proteins: primary, secondary, tertiary and quaternary.

Primary the structure of proteins is the simplest. It has the form of a polypeptide chain, where amino acids are linked to each other by a strong peptide bond. Determined by quality and quantitative composition amino acids and their sequence.

Secondary structure of proteins

Secondary the structure is formed predominantly by hydrogen bonds that were formed between the hydrogen atoms of the NH group of one helix curl and the oxygen atoms of the CO group of the other and are directed along the spiral or between parallel folds of the protein molecule. The protein molecule is partially or entirely twisted into an α-helix or forms a β-sheet structure. For example, keratin proteins form an α-helix. They are part of hooves, horns, hair, feathers, nails, and claws. The proteins that make up silk have a β-sheet. Amino acid radicals (R-groups) remain outside the helix. Hydrogen bonds are much weaker than covalent bonds, but with a significant number of them they form a fairly strong structure.

Functioning in the form of a twisted spiral is characteristic of some fibrillar proteins - myosin, actin, fibrinogen, collagen, etc.

Protein tertiary structure

Tertiary protein structure. This structure is constant and unique for each protein. It is determined by the size, polarity of R-groups, shape and sequence of amino acid residues. The polypeptide helix is ​​twisted and folded in a certain way. The formation of the tertiary structure of a protein leads to the formation of a special configuration of the protein - globules (from Latin globulus - ball). Its formation is determined by different types of non-covalent interactions: hydrophobic, hydrogen, ionic. Disulfide bridges appear between cysteine ​​amino acid residues.

Hydrophobic bonds are weak bonds between non-polar side chains that result from the mutual repulsion of solvent molecules. In this case, the protein twists so that the hydrophobic side chains are immersed deep inside the molecule and protect it from interaction with water, while the hydrophilic side chains are located outside.

Most proteins have a tertiary structure - globulins, albumins, etc.

Quaternary protein structure

Quaternary protein structure. Formed as a result of the combination of individual polypeptide chains. Together they form a functional unit. There are different types of bonds: hydrophobic, hydrogen, electrostatic, ionic.

Electrostatic bonds occur between electronegative and electropositive radicals of amino acid residues.

Some proteins are characterized by a globular arrangement of subunits - this is globular proteins. Globular proteins easily dissolve in water or salt solutions. Over 1000 known enzymes belong to globular proteins. Globular proteins include some hormones, antibodies, and transport proteins. For example, the complex molecule of hemoglobin (red blood cell protein) is a globular protein and consists of four globin macromolecules: two α-chains and two β-chains, each of which is connected to heme, which contains iron.

Other proteins are characterized by association into helical structures - this is fibrillar (from Latin fibrilla - fiber) proteins. Several (3 to 7) α-helices are twisted together, like fibers in a cable. Fibrillar proteins are insoluble in water.

Proteins are divided into simple and complex.

Simple proteins (proteins)

Simple proteins (proteins) consist only of amino acid residues. Simple proteins include globulins, albumins, glutelins, prolamins, protamines, pistons. Albumins (for example, serum albumin) are soluble in water, globulins (for example, antibodies) are insoluble in water, but soluble in aqueous solutions some salts (sodium chloride, etc.).

Complex proteins (proteids)

Complex proteins (proteids) include, in addition to amino acid residues, compounds of a different nature, which are called prosthetic group. For example, metalloproteins are proteins containing non-heme iron or linked by metal atoms (most enzymes), nucleoproteins are proteins connected to nucleic acids (chromosomes, etc.), phosphoproteins are proteins that contain phosphoric acid residues (egg proteins yolk, etc.), glycoproteins - proteins combined with carbohydrates (some hormones, antibodies, etc.), chromoproteins - proteins containing pigments (myoglobin, etc.), lipoproteins - proteins containing lipids (included in the composition of membranes).

Proteins are one of the important organic elements of any living cell in the body. They perform many functions: support, signaling, enzymatic, transport, structural, receptor, etc. The primary, secondary, tertiary and quaternary structures of proteins have become an important evolutionary adaptation. What are these molecules made of? Why is the correct conformation of proteins in the cells of the body so important?

Structural components of proteins

The monomers of any polypeptide chain are amino acids (AA). These low-molecular organic compounds are quite common in nature and can exist as independent molecules that perform their inherent functions. Among them are transport of substances, reception, inhibition or activation of enzymes.

There are about 200 biogenic amino acids, but only 20 of them can be found. They are easily soluble in water, have a crystalline structure, and many of them taste sweet.

From a chemical point of view, AA are molecules that necessarily contain two functional groups: -COOH and -NH2. With the help of these groups, amino acids form chains, connecting to each other with peptide bonds.

Each of the 20 proteinogenic amino acids has its own radical, depending on which the Chemical properties. Based on the composition of such radicals, all AAs are classified into several groups.

1. Non-polar: isoleucine, glycine, leucine, valine, proline, alanine.
2. Polar and uncharged: threonine, methionine, cysteine, serine, glutamine, asparagine.
3. Aromatic: tyrosine, phenylalanine, tryptophan.
4. Polar and negatively charged: glutamate, aspartate.
5. Polar and positively charged: arginine, histidine, lysine.

Any level of protein structure organization (primary, secondary, tertiary, quaternary) is based on a polypeptide chain consisting of AK. The only difference is how this sequence folds in space and with the help of what chemical bonds this conformation is maintained.

Primary protein structure

Any protein is formed on ribosomes - non-membrane cell organelles that participate in the synthesis of the polypeptide chain. Here, amino acids are connected to each other using a strong peptide bond, forming the primary structure. However, this primary structure of the protein is extremely different from the quaternary one, so further maturation of the molecule is necessary.

Proteins such as elastin, histones, glutathione, even with such a simple structure, are able to perform their functions in the body. For the vast majority of proteins, the next stage is the formation of a more complex secondary conformation.

Protein secondary structure

The formation of peptide bonds is the first step in the maturation of most proteins. In order for them to perform their functions, their local conformation must undergo some changes. This is achieved with the help of hydrogen bonds - fragile, but at the same time numerous connections between the basic and acidic centers of amino acid molecules.

This is how the secondary structure of the protein is formed, which differs from the quaternary structure in its simplicity of assembly and local conformation. The latter means that not the entire chain undergoes transformation. Hydrogen bonds can form at several sites at different distances from each other, and their form also depends on the type of amino acids and the method of assembly.

Lysozyme and pepsin are representatives of proteins that have a secondary structure. Pepsin is involved in digestion processes, and lysozyme performs a protective function in the body, destroying the cell walls of bacteria.

Features of the secondary structure

Local conformations of the peptide chain may differ from each other. Several dozen of them have already been studied, and three of them are the most common. These include the alpha helix, beta sheets, and beta turn.

• The alpha helix is ​​one of the common secondary structure conformations of most proteins. It is a rigid rod frame with a stroke of 0.54 nm. Amino acid radicals are directed outward.

Right-handed helices are most common, and left-handed counterparts can sometimes be found. The shape-forming function is performed by hydrogen bonds, which stabilize the curls. The chain that forms the alpha helix contains very little proline and polar charged amino acids.

• The beta turn is separated into a separate conformation, although it can be called part of the beta sheet. The essence is the bending of the peptide chain, which is supported by hydrogen bonds. Typically, the bend itself consists of 4-5 amino acids, among which the presence of proline is obligatory. This AK is the only one with a rigid and short skeleton, which allows it to form a turn.
• The beta layer is a chain of amino acids that forms several bends and stabilizes them with hydrogen bonds. This conformation is very reminiscent of a sheet of paper folded into an accordion. Most often, aggressive proteins have this form, but there are many exceptions.

There are parallel and antiparallel beta layers. In the first case, the C- and N-termini at the bend points and at the ends of the chain coincide, but in the second case they do not.

Tertiary structure

Further packaging of the protein leads to the formation of a tertiary structure. This conformation is stabilized with the help of hydrogen, disulfide, hydrophobic and ionic bonds. Their large number makes it possible to twist the secondary structure into a more complex shape and stabilize it.

They are divided into globular and The globular peptide molecule has a spherical structure. Examples: albumin, globulin, histones in the tertiary structure.

They form strong strands whose length exceeds their width. Such proteins most often perform structural and shape-forming functions. Examples are fibroin, keratin, collagen, elastin.

Structure of proteins in the quaternary structure of a molecule

If several globules combine into one complex, a so-called quaternary structure is formed. This conformation is not typical for all peptides, and it is formed when it is necessary to perform important and specific functions.

Each globule in the composition represents a separate domain or protomer. Collectively, the molecule is called an oligomer.

Typically, such a protein has several stable conformations, which constantly replace each other, either depending on the influence of any external factors, or when necessary to perform different functions.

An important difference between the tertiary structure of a protein and the quaternary one is the intermolecular bonds, which are responsible for connecting several globules. In the center of the entire molecule there is often a metal ion, which directly affects the formation of intermolecular bonds.

Additional protein structures

A chain of amino acids is not always sufficient to perform the functions of a protein. In most cases, other substances of organic and inorganic nature are attached to such molecules. Since this feature is characteristic of the vast majority of enzymes, the composition of complex proteins is usually divided into three parts:

• An apoenzyme is the protein part of a molecule, which is an amino acid sequence.
• A coenzyme is not a protein, but an organic part. It may contain various types of lipids, carbohydrates or even nucleic acids. This also includes representatives of biologically active compounds, among which are vitamins.
• Cofactor is an inorganic part, represented in the vast majority of cases by metal ions.

The structure of proteins in the quaternary structure of a molecule requires the participation of several molecules of different origins, so many enzymes have three components at once. An example is phosphokinase, an enzyme that ensures the transfer of a phosphate group from an ATP molecule.

Where is the quaternary structure of a protein molecule formed?

The polypeptide chain begins to be synthesized on the cell's ribosomes, but further protein maturation occurs in other organelles. The newly formed molecule must enter the transport system, which consists of the nuclear membrane, ER, Golgi apparatus and lysosomes.

The complication of the spatial structure of the protein occurs in the endoplasmic reticulum, where not only different kinds bonds (hydrogen, disulfide, hydrophobic, intermolecular, ionic), but also a coenzyme and a cofactor are added. This is how the quaternary structure of the protein is formed.

When the molecule is completely ready for work, it enters either the cytoplasm of the cell or the Golgi apparatus. In the latter case, these peptides are packaged into lysosomes and transported to other cell compartments.

Examples of oligomeric proteins

Quaternary structure is the structure of proteins that is designed to facilitate the performance of vital functions in a living organism. Complex conformation organic molecules allows, first of all, to influence the work of many metabolic processes (enzymes).

Biologically important proteins are hemoglobin, chlorophyll and hemocyanin. The porphyrin ring is the basis of these molecules, in the center of which is a metal ion.

Hemoglobin

The quaternary structure of the hemoglobin protein molecule consists of 4 globules connected by intermolecular bonds. In the center is porphine with ferrous iron ion. The protein is transported in the cytoplasm of red blood cells, where they occupy about 80% of the total volume of the cytoplasm.

The basis of the molecule is heme, which is more inorganic in nature and is colored red. It is also the breakdown of hemoglobin in the liver.

We all know that hemoglobin performs an important transport function - the transfer of oxygen and carbon dioxide throughout the human body. The complex conformation of the protein molecule forms special active centers, which are capable of binding the corresponding gases to hemoglobin.

When the protein-gas complex is formed, so-called oxyhemoglobin and carbohemoglobin are formed. However, there is another type of such associations that is quite stable: carboxyhemoglobin. It is a complex of protein and carbon monoxide, the stability of which explains attacks of suffocation due to excessive toxicity.

Chlorophyll

Another representative of proteins with a quaternary structure, the domain connections of which are supported by a magnesium ion. Main function the entire molecule - participation in the processes of photosynthesis in plants.

There are different types of chlorophylls, which differ from each other by the radicals of the porphyrin ring. Each of these varieties is marked with a separate letter of the Latin alphabet. For example, land plants are characterized by the presence of chlorophyll a or chlorophyll b, and other types of this protein are found in algae.

Hemocyanin

This molecule is an analogue of hemoglobin in many lower animals (arthropods, mollusks, etc.). The main difference between the protein structure and the quaternary structure of the molecule is the presence of a zinc ion instead of an iron ion. Hemocyanin has a bluish color.

Sometimes people wonder what would happen if we replaced human hemoglobin with hemocyanin. In this case, the usual content of substances in the blood, and in particular amino acids, is disrupted. Hemocyanin also complexes unstable with carbon dioxide, so blue blood would have a tendency to form blood clots.