Fundamentals of Biochemistry - A. A. Anisimov 1986

Proteins
Classification, Characteristics, and Representatives

All Proteins are generally divided into two groups: simple proteins, or proteins proper (consisting solely of Amino Acids), and Conjugated Proteins¹ (whose molecules contain a non-protein, prosthetic moiety In addition to the protein part). Each of these groups is further subdivided into subgroups. Simple proteins are classified according to their solubility in various substances, whereas conjugated proteins are categorized based on the Chemical Nature of their non-protein moiety. It should be noted at once that the Classification of simple proteins is rather arbitrary and imperfect, as it relies on comparing relatively minor characteristics. Many proteins previously assigned to the simple protein group (such as Blood globulins) have been shown by molecular Structure studies to be two-component systems. Free Peptides also occur naturally; as mentioned earlier, their molecules contain no more than 50 amino acid residues.

¹ For a long time, all conjugated proteins were referred to as proteids (chromoproteids, Lipoproteins, nucleoproteids, etc.). In recent years, names ending in -ins (Chromoproteins, lipoproteins, Nucleoproteins, etc.) have become increasingly standard for conjugated proteins. This is mainly because the term proteids implies protein-like substances.

2.5.1. Natural peptides. Several hundred free peptides have been discovered in living organisms. These include certain Hormones, toxins, Antibiotics, Neuropeptides, and a range of other BIOLOGICALLY ACTIVE SUBSTANCES.

Class="center">Glutathione (y-glutaminylcysteinylglycine, glu-cys-gly G-SH).

Found in animal and human Cells (especially abundant in the Brain and lens of the eye), Bacteria, Yeasts, Fungi, and green plants. It plays an active role in oxidation-reduction processes:

The primary function of glutathione within cells is to protect protein sulfhydryl groups from oxidation. It also acts as a coenzyme in A number of enzymatic reactions (see Section 3.4.3). In animals and humans, glutathione participates in The breakdown of H₂O₂ produced in erythrocytes through metabolic processes or the autoxidation of drugs. It is involved in the Detoxification of Various compounds foreign to living cells (halogenated aliphatic or aromatic Hydrocarbons), converting them into a Water-soluble form that can be excreted by the Kidneys.

Gramicidin S is an antibiotic actively synthesized by the bacterium Bac. brevis. Chemically, it is a cyclic decapeptide containing Ornithine (orn) in addition to Proteinogenic Amino Acids:

Peptide antibiotics of the gramicidin type function as ionophores, acting on Introduction/36.html">Biological Membranes by forming complexes with Metal Ions, thereby disrupting The regulation of ion permeability in bacterial membranes.

Amanitins are toxic octapeptides produced by poisonous mushrooms of the genus Amanita. A typical representative of this group is a-amanitin, which blocks Protein Synthesis in Eukaryotic cells at the METABOLISM/31.html">Transcription stage. Replacing the dioxyisoleucine residue with leucine in the a-amanitin molecule yields a non-toxic compound.

These same mushrooms contain a number of toxic heptapeptides known as phalloidins, which have a bicyclic structure similar to that of amanitins. Phalloidins cause irreversible Liver damage in mammals. The antidote to phalloidins is antamanide, a cyclic decapeptide found in the same mushrooms as the toxin. Antamanide tightens liver Cell membranes and reduces their permeability to toxins. It acts as an ionophore that binds Na⁺ or Ca²⁺ ions adsorbed on the membrane surface, thereby shielding specific membrane regions, tightening the structure, and altering its properties—particularly its permeability. Another peptide toxic to humans, muscarine, has been isolated from Amanita muscaria.

A large number of Peptide Hormones are known and will be discussed in Chapter 12.

Free peptides include kallidin and Blood Plasma bradykinin, which belong to the kinins. They increase capillary permeability, exhibit potent vasodilating activity, and act as powerful pain triggers. Both peptides are generated from a common precursor, kininogen, via proteolytic Cleavage. Bradykinin is a linear nonapeptide: arg — pro — pro — gly — phe — ser — pro — phe — arg; kallidin differs by the presence of an additional amino acid residue (lys) at the N-terminus. These peptides are inactivated by the Cleavage of the C-terminal Arginine mediated by carboxypeptidase B.

Opioid peptides (endopioids) are a group of neuropeptides that exert a modulatory effect on Nerve Impulse transmission across various Regions of the Central Nervous system (CNS). These peptides interact with the same receptors as opiate compounds (such as morphine) and exhibit similar physiological effects.

The highest density of endopioid binding receptors is found in the thalamus, Hypothalamus, neurohypophysis, and several other regions of the CNS. Opioid peptide receptors are also located in the Peripheral Nervous System. Endopioids participate in regulating processes related to pain perception, the metabolic effects of hormones, cardiovascular activity, stress responses, and various other physiological functions. For instance, during acupuncture analgesia (pain relief via needle stimulation), the concentration of endopioids in the CEREBROSPINAL FLUID increases, indicating the activation of this system. Opioid peptides also exert some influence on emotional states and behavior.

Currently, several dozen opioid peptides are known. Specific Examples include a-, ß-, and y-endorphins, a- and ß-neoendorphins, dynorphin, and the pentapeptides Methionine-enkephalin (tyr — gly — gly — phe — met) and leucine-enkephalin (tyr — gly — gly — phe — leu).

Of particular interest is The Biosynthesis of opioid peptides. The core principle involves the post-translational cleavage of high-molecular-weight protein precursors by proteases. One such endopioid precursor is proopiomelanocortin, a pre-hormonal protein with a Molecular Weight of approximately 31,000. Its molecule contains the Amino acid sequences of melanocyte-stimulating hormones (MSH), adrenocorticotropic hormone (ACTH), and ß-lipotropic hormone. In turn, ß-lipotropin—a hormone that stimulates the release of Fatty acids from adipose tissue and consists of 91 amino acid residues in humans—contains sequences analogous to ß-MSH (41–58), ACTH4-10 (47–53), as well as endopioids such as a-endorphin (61–76), ß-endorphin (61–91), y-endorphin (61–77), and methionine-enkephalin (61–65). The Amino Acid Sequence of human ß-lipotropin, which encompasses the ß-MSH, ß-endorphin, and methionine-enkephalin sequences, is presented below.

However, because ß-lipotropin and enkephalins exhibit different tissue localizations, it is believed that the primary precursor for enkephalins is not proopiomelanocortin, but rather other Polypeptides containing multiple copies of methionine-enkephalin and leucine-enkephalin.

Chalones are tissue-specific, locally acting hormones represented by proteins or peptides of varying molecular weights. Chalones suppress the mitotic activity of other cells within the same tissue. By participating in the Regulation of Cell division, these hormones presumably prevent malignant cell growth.

Folic acid, which functions biologically as a vitamin (see Section 10.3), can also be classified among peptides. Its structure contains up to 7 glu residues in the form of a y-glutaminyl peptide (meaning the glutamic acid residues are linked via y-carboxyl groups rather than a-groups).

In addition to those listed, peptides perform a number of other fascinating and crucial functions. Hormones responsible for Sleep induction have been discovered. Certain specific peptides may act as substances involved in memory phenomena and conditioned reflex acquisition. It is quite possible that long-term memory is linked to the synthesis of these peptides in specific Neurons. For example, when rats are trained to avoid the dark, a 15-residue peptide (scotophobin) accumulates in their brains; administering this peptide to untrained animals induces the same avoidance behavior. A correlation has also been observed between deviations in human mental activity from the norm and the levels of specific peptides in the brain.

Thus, the biological activity of peptides is tied to their regulatory role, with their sites of action and physiological efficacy being highly diverse. Natural peptides are currently attracting immense research interest, and active work is underway to isolate them, determine their structures and functions, and chemically synthesize them for practical medical Applications as therapeutic agents.

2.5.2. Proteins (Simple Proteins). Below are the groups of simple proteins, classified According to the solubility of these compounds.

Albumins. These are water-soluble proteins that precipitate upon saturation of solutions with neutral salts, such as (NH₄)₂SО₄. The addition of a single salt usually does not lead to protein precipitation (with the exception of (NH₄)₂SО₄); a mixture of salts is required, predominantly mono- and divalent cations (NaCl and MgSО₄ or Na₂SО₄ and MgCl₂). Ammonium sulfate begins to precipitate albumins at 65% saturation, and complete precipitation occurs at 100% saturation.

Albumins are widely distributed in nature. They make up about 50% of all human Blood Plasma Proteins. Egg white has a high albumin content (up to 50%). A protein with similar solubility, named lactalbumin, has been isolated from milk; plants are also rich in albumins.

Globulins. They are soluble in dilute solutions of neutral salts, although high concentrations of the latter precipitate globulins. (NH₄)₂SО₄ salts out globulins at as low as 50% saturation, although it should be noted that complete Separation of albumins and globulins does not occur either at this concentration of ammonium sulfate or at others. Globulins are insoluble in water, and therefore they precipitate out when salts are removed by dialysis. Globulins constitute the bulk of the seed proteins in many plants, especially legumes and oilseeds, such as legumin in pea seeds, phaseolin in beans, and edestin in hemp.

Prolamins. They are readily soluble in 60–80% ethyl alcohol and contain high amounts of The amino acid Proline, as well as glutamic acid. These proteins contain very minor amounts of lys, arg, and gly. Prolamins are exclusively characteristic of cereal seeds, where they serve as storage proteins: gliadin is found in wheat and rye seeds, hordein in barley seeds, and zein in corn. All of these prolamins represent Protein Complexes that differ in composition and molecular weight.

Glutelins. They are readily soluble in alkaline solutions (0.2–2% NaOH). These are plant proteins found in the seeds of cereals and other crops, as well as in the green parts of plants. The complex of alkali-soluble wheat seed proteins is called glutenin, and that of rice is called oryzenin. Wheat seed gliadin, when combined with glutenin, forms gluten, The properties of which largely determine the technological qualities of flour and dough.

Histones. These are basic proteins with a molecular weight of 12,000–30,000, in which basic amino acids account for 20–30%. Histones are soluble in weak acids (0.2 N HCl) and are precipitated by ammonia and alcohol. They contain no Tryptophan and, in most cases, no Cysteine or cystine. Histones are present mainly in The Cell nuclei of animals and plants and play a crucial role in Chromatin Structure, as they quantitatively predominate among chromosomal proteins.

Histones are relatively evolutionarily conservative proteins. It has been shown that animal and plant histones are characterized by close values of the arg-to-lys ratio and contain a fairly similar set of fractions.

According to X-Ray Diffraction Analysis and Electron Microscopy data, histones are not found in the Chromosomes of organisms lacking a well-defined Cell Nucleus (bacteria, blue-green Algae). At the same time, it should be noted that protein fractions rich in lys and arg have been isolated from the cells of certain bacteria and some blue-green algae. Data on the histone content in fungi remain contradictory.

Protamines. These are strongly basic proteins with a low molecular weight (up to 12,000), due to which some of them pass through cellophane during dialysis. Protamines are soluble in weak acids and are not precipitated by boiling; basic amino acids make up about 80% of their molecules, with arg being particularly abundant. Protamines lack cys, trp, and asp, and very often lack tyr and phe, which is why they do not give many Color Reactions for proteins. Due to the high concentration of basic amino acids in their molecules, they act as polyvalent organic cations and readily react with molecules possessing an excess of negatively charged groups, particularly Nucleic Acids.

Protamines are widely distributed in nature. They are found in the Germ Cells of animals and humans and constitute the bulk of the chromatin proteins of this type. Protamines impart biochemical inertness to DNA, which is a prerequisite for preserving the hereditary traits of an Organism. It has been shown that protamine synthesis occurs during Spermatogenesis in the Cytoplasm of the germ cell; subsequently, protamines are phosphorylated, penetrate the cell nucleus, and, as the sperm matures, displace histones from the nucleoprotein to form a stable complex with DNA. As a result of this interaction, the hereditary traits of the organism are protected from adverse influences. Protamines are found in large quantities in fish sperm; salmine from salmon and clupeine from herring are the most thoroughly studied among them. Protamines have also been discovered in Representatives of the plant kingdom—they were isolated from clubmoss spores.

Proteinoids. These are poorly soluble proteins characterized by a high sulfur content. Proteinoids include Fibrous proteins: Fibroin, the protein of silk; Keratins, the proteins of Hair, horns, and hooves; collagens, the proteins of Connective Tissue; spongin, the protein of sea Sponges, etc.

2.5.3. Complex Proteins; Lipoproteins. The prosthetic group in these complex proteins consists of various fat-like substances—Lipids. The strength of the bond between lipoprotein components can vary. It is believed that weak interaction forces (hydrophobic, ionic, hydrogen) make the primary contribution to stabilizing these complexes, while The Role of covalent bonds is negligible.

Both polar and neutral lipids, as well as Cholesterol and its esters, have been found in the composition of lipoproteins. Lipoproteins are widely distributed in nature and are found in All living organisms. They are obligate components of all cell membranes, where their non-protein moiety is represented mainly by polar lipids—Phospholipids and Glycolipids. Lipoproteins are always present in the blood (see Section 2.5.4). An Inositol diphosphate-containing lipoprotein has been isolated from white brain matter, while the lipoproteins of gray brain matter contain Sphingolipids. In plants, a significant portion of phospholipids in the protoplasm is also present in the form of complexes with proteins.

Lipid-protein complexes are known in which the protein moiety contains many hydrophobic amino acids, and the lipid component often predominates over the protein one. As a result, such complex proteins are soluble in organic Solvents, such as a chloroform-methanol mixture. Such complexes are called proteolipids. They are present in large quantities in the myelin sheaths of Nerve Cells, as well as in synaptic membranes and the inner membranes of Mitochondria.

Phosphoproteins. A characteristic feature of phosphoproteins is the presence of significant amounts of orthophosphoric acid, which is usually bound via an ester linkage to the hydroxyl group of ser, and less frequently thr. Other hydroxyamino acids (Tyrosine, hydroxyproline) do not form phosphate esters.

Phosphoproteins include many proteins that play an essential role in the Nutrition of young organisms. These are casein, the main milk protein that precipitates upon curdling; vitellin and phosvitin from egg yolk; and ichthulin from fish roe. They contain 1–10% phosphorus. Phosphoproteins have also been detected in the brain.

In addition to phosphoric acid, casein contains a carbohydrate component in its molecule, making it a phosphoglycoprotein. Relatively short-term phosphorylation of A wide variety of proteins (enzymatic, membrane, ribosomal, etc.) occurs with the participation of a special group of Enzymes known as protein Kinases. Such phosphorylation is of a regulatory, transient nature, and these proteins should presumably be distinguished from structurally permanent phosphoproteins.

Metalloproteins. Complexes of metal ions with proteins, in which the metal ions are directly attached to the protein and constitute an integral part of the protein molecule's structure, are called metalloproteins. Some authors also classify proteins that possess metal-binding prosthetic groups in addition to the metal—such as the porphyrin group in Hemoglobin or chlorophyll—as metalloproteins. However, this view is not universally accepted.

Metals such as Cu, Fe, Zn, Mo, and others are frequently found in the composition of metalloproteins. Typical metalloproteins include certain enzymes containing the aforementioned metals, as well as Mn, Ni, Se, Ca, etc. (see Section 3.4.2).

Copper-containing proteins include, for example, cytochrome c oxidase, plastocyanin (electron carriers), and the blood protein ceruloplasmin; iron-containing proteins include lactoferrin (a milk protein), transferrin (a blood protein), ferritin, etc. A specific nickel-containing protein belonging to the macroglobulin class, named nickeloplasmin, has been found in blood serum.

Proteins known as selenoproteins have been discovered in which selenium is most likely covalently attached to an aromatic or heterocyclic (heme) group. One of the selenoproteins is found in animal Muscles.

A vanadium-containing protein, vanadochrome—which is most likely an oxygen carrier—has been discovered in certain marine animals.

Glycoproteins. These are complex proteins that contain a carbohydrate component. In these compounds, the protein serves as a kind of backbone to which carbohydrate groups are attached. There is no universally recognized and established classification for these proteins. Based on their chemical structure, glycoproteins can be divided into true Glycoproteins and Proteoglycans (glycosaminoproteoglycans).

The main difference between them is that the carbohydrate groups of true glycoproteins typically contain up to 15–20 monosaccharide units that do not form repeating oligosaccharide fragments, whereas in proteoglycans they are constructed from a very large number of repeating units, mostly having a characteristic disaccharide nature.

True glycoproteins. The molecular weight of true glycoproteins varies widely, sometimes reaching 1 million or more. The molecular weight is especially large in salivary glycoproteins. The carbohydrate component accounts for anywhere from 1–3% (Ovalbumin) to 80–90% (blood group substances) of the total molecular mass. The number of carbohydrate chains per molecule also varies significantly. For instance, transferrin and Ribonuclease contain no more than 1–4 chains, whereas blood group substances and salivary mucins contain up to 300–800. The carbohydrate chains, whether branched or linear, are always covalently linked to the peptide portion of the molecule.

More than 10 different Monosaccharides have been identified within the carbohydrate components: D-galactose, D-manose, D-glucose, N-acetylglucosamine and N-acetylgalactosamine, deoxy sugars (L-fucose, L-rhamnose), D-xylose, and L-arabinose. Neuraminic acid (see Section 6.2.3) is also a typical component of glycoproteins; due to its carboxyl group (at physiological pH values), it imparts a negative charge to the carbohydrate-containing molecule. Neuraminic acid most commonly occurs in the form of sialic acids, which, along with fucose, typically occupy a terminal position in the carbohydrate chains of glycoproteins.

Various chemical groupings can form the covalent bond between the carbohydrate and protein parts of glycoproteins. The glycosylamide type of linkage between N-acetylglucosamine and the ß-amide nitrogen of asparagine is very common.

Another type of carbohydrate-peptide bond in glycoproteins is the O-glycosidic bond. This linkage most frequently involves Serine or Threonine and N-acetylgalactosamine or galactose.

Collagen contains a galactosyl-hydroxylysine O-glycosidic bond, while Carbohydrate-Containing Proteins of higher plants contain an arabinosyl-hydroxyproline bond. Furthermore, an S-glycosidic bond between galactose and cysteine, as well as an O-glycosidic bond between fucose and threonine, has been discovered in human urinary glycoproteins. More than one type of carbohydrate-peptide linkage may occur within the molecules of the same glycoprotein.

Proteoglycans. The molecular weight of proteoglycans is large, sometimes reaching several millions due to a high number of repeating disaccharide units. Solutions of these carbohydrate-protein complexes exhibit high viscosity. Proteoglycans consist of a small protein core to which a significant number (several dozens) of heteropolysaccharide chains, containing amino sugar and uronic acid residues, are covalently attached.

The carbohydrate components of proteoglycans, known as glycosaminoglycans, are primarily represented by the following Polysaccharides: hyaluronic acid, chondroitin sulfates, heparin, heparan sulfate, and keratan sulfates. All of these polysaccharides contain repeating paired units composed of amino sugar residues, hexuronic acids, and, less frequently, monosaccharides (see Section 6.4). In proteoglycans containing hyaluronic acid, the protein moiety accounts for 0.4–2% of the total molecular mass, whereas for chondroitin sulfates, it makes up 17–22%.

The carbohydrate-peptide linkage can be established via the O-glycosidic bond of D-xylose to a serine residue of the peptide chain (in chondroitin sulfates and heparin) or N-acetylglucosamine to threonine, as well as the glycosylamide bond of N-acetylglucosamine to asparagine (in keratan sulfates).

Xylose is not a constituent of glycosaminoglycans; instead, it acts as an intercalated linking component between the polysaccharide and the protein. Proteoglycans exhibit polyanionic properties due to the presence of uronic acid carboxyl groups and amino sugar sulfate groups.

Biological Role of glycoproteins. Carbohydrate-containing proteins are widely distributed in living organisms, occurring in animals, plants, and microorganisms, where they perform a wide variety of functions.

1. Function of selective interaction and highly specific recognition. Along with other components, surface membranes contain glycoproteins that participate in the subtle processes of biological recognition and intercellular interaction, serving as receptor systems for specific compounds and cells. For instance, the epithelial Cells of the intestinal mucosa are equipped with receptors that specifically bind cells of bacteria and Viruses infecting the organism.

Certain glycoproteins of Erythrocyte membranes are responsible for the selective attachment of the Influenza virus. The specific binding of hormones to The surface of target cells is mediated in many cases by the carbohydrate-containing compounds of these cells. A receptor of this type for Insulin exists On the surface of liver cells, adipose tissue, and lymphocytes. Hormones with pre-cleaved terminal sialic acids fail to reach target cells when introduced into the bloodstream. The crucial role of the carbohydrate moiety of glycoproteins in determining the Specificity of many Antigens is also unquestionable. This applies primarily to blood group substances and soluble group substances of biological fluids (saliva, milk, seminal fluid).

Bacterial antigenic complexes possess a highly complex chemical structure, consisting of protein, polysaccharide, and lipid components. Complex antigens of many bacteria that are toxic to humans and animals are termed endotoxins. The determining regions of the polysaccharide components of endotoxin macromolecules dictate their antigen specificity. Peptides also act as immunodeterminants and enhance the immunogenicity of macromolecules. All or most of the active toxic sites are localized within the lipid component, which also determines its pyrogenic effect.

The uptake of Bacterial toxins by host cells also occurs with the involvement of glycoproteins.

The carbohydrate component of glycoproteins serves as a sort of postal code in which The sequence of sugars in the carbohydrate chain dictates where the molecule should go—whether to exit the cell, incorporate into cellular membranes of various Organs, or integrate into subcellular membranes.

Glycoproteins are common components of plant seeds. For example, bean seeds contain vicilin, a protein composed of mannose and N-acetylglucosamine, while castor bean seeds contain ricin, a protein capable of selectively precipitating blood group-specific substances and agglutinating erythrocytes. Plant-derived compounds exhibiting this property are called phytohemagglutinins and belong to the lectin group. Lectins are defined as non-immune proteins, including glycoproteins, capable of specifically binding polysaccharides and carbohydrate-containing Biopolymers without causing their chemical transformation. Specifically, the interaction of lectins with carbohydrate-containing compounds manifests as the agglutination of particles and cells, such as erythrocytes, or the precipitation of polysaccharides and glycoproteins.

To date, lectins have been discovered in organisms at various Stages of the evolutionary ladder: bacteria, algae, fungi, Mollusks, fish, mammals, and other animals. Consequently, it has been proposed to use terms such as "phytolectins," "zoolectins," "mycolectins," etc.

In mammals, certain lectins likely participate in Organogenesis during Embryogenesis, as they are found in large quantities in Embryonic and Fetal Tissues. Furthermore, in animals, lectins presumably take part in the intracellular compartmentalization of specific glycoproteins. The diverse functions of lectins are intimately linked to their ability to recognize cells and cellular components and to orchestrate their selective interactions.

Lectins are widespread in plants, and it is hypothesized that they may be involved in regulating Cell Division and seed germination. Lectins play a vital role in recognition processes during The formation of symbiotic associations, particularly between legumes and nitrogen-fixing ROOT nodule bacteria. Fungal lectins fulfill a similar role in the formation of Lichens. Lectins also participate in the establishment of parasitic interspecies associations, such as higher plant–bacteria and higher plant–fungi. Furthermore, lectins are important in Cell Recognition during the formation of colonial microorganisms. Cell Adhesion driven by lectin-carbohydrate recognition is well demonstrated in the aggregation of cellular slime Molds from single-celled myxamoebae. It has also been suggested that lectins take part in the Transport of Assimilates through the plant phloem, which is likewise explained by their role in selective uptake.

Because the binding sites of Lectins are specific to certain carbohydrate residues, they are currently used as tools to probe the architectonics of cell surfaces.

Thus, in many instances where living organisms exhibit fine biochemical specificity and highly selective recognition, CARBOHYDRATES are involved. Carbohydrate components of glycoproteins are especially important in this regard, containing a wide variety of monosaccharide residues in various combinations, which, in complex with proteins, provides the potential for high biochemical specificity.

2. Transport function. A number of glycoproteins circulating in the bloodstream of humans and animals act as transport proteins; for example, transferrin functions as an iron carrier, ceruloplasmin transports copper, and transcortin carries Steroid Hormones. The major integral protein of The erythrocyte membrane is involved in The transport of anions and, potentially, glucose.

3. Catalytic function. The carbohydrate component has been identified within several enzymes, notably enterokinase, taka-amylase, peroxidase, glucose oxidase, serum cholinesterase (which hydrolyzes acetylcholine and participates in nerve impulse transmission), ribonuclease B, and others.

4. Structural and mechanical function. In various living organisms, this function is performed primarily by proteoglycans. In vertebrates, they are constituents of the intercellular matrix of connective tissue. Proteoglycans are found in the Skin, bones, Cartilage, synovial fluid of joint capsules, tendons, Heart Valves, vitreous humor, Cytology/practical/76.html">Cornea of the eye, and other tissues, imparting them with elasticity and resistance to compression. Proteoglycans containing hyaluronic acid form highly viscous solutions, which increases the tissue's resistance to infection. In many bacteria, the protective tissue capsule is largely built from complex proteins that include hyaluronic acid. Furthermore, this proteoglycan, along with chondroitin sulfate A, acts as a lubricant in joints. The function of protective lubrication is also performed by glycoproteins, which are the main constituents of salivary mucins, as well as gastric and intestinal mucins. Glycoproteins are widely distributed Structural components of various cell membranes.

In addition to those listed, glycoproteins perform several other functions in the body. Glycoproteins include fibrinogen, prothrombin, and certain other blood clotting factors. The liver synthesizes glycosaminoglycan, or heparin, a crucial anticoagulant substance. IMMUNOGLOBULINS (see Section 5.5), certain hormones such as gonadotropic and follicle-stimulating hormones, thyroglobulin, and interferon (an inhibitor of animal virus Replication) are also glycoproteins. The hemagglutinating activity of certain viruses is associated with the presence of glycoproteins in their envelope.

The carbohydrate component enhances the stability of glycoprotein molecules, protecting them from Proteolytic Enzymes. The non-protein moiety of several serum glycoproteins determines their half-life. Antifreeze glycoproteins, which protect cells from freezing, have been found in the blood serum and muscles of Antarctic fish. Some connective tissue proteoglycans (chondroitin sulfates) are capable of Swelling by absorbing large amounts of water, and therefore sometimes act as water reserves.

Avidin, an egg white protein, is a glycoprotein. It interacts with vitamin H (biotin) and prevents its absorption from the intestine, leading to acute biotin deficiency. Avidin is used in in vitro experiments as an inhibitor of biotin-containing enzymes.

Chromoproteins. Chromoproteins are conjugated proteins whose non-protein moiety consists of colored compounds belonging to various classes of organic substances: porphyrin structures, flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), etc. The porphyrin ring with a coordinately bound iron ion serves as the prosthetic group for a number of oxidation-reduction enzymes (catalase, peroxidase) and a group of electron carriers known as Cytochromes. Flavine dehydrogenases, or "yellow respiratory enzymes"—flavoproteins (FP)—are also chromoproteins. The protein moiety of their molecules is bound to FAD or FMN. Flavoproteins are discussed in more detail in Chapter 3. Typical chromoproteins include rhodopsin (see Section 10.2) and blood hemoglobin.

The most important group of conjugated proteins, nucleoproteins, is discussed in Chapter 4.

Hemoglobin and other respiratory pigments. Hemoglobin (abbreviated as Hb) forms the Molecular Basis of the RESPIRATORY FUNCTION OF Blood, i.e., The ability to transport O₂ and CO₂.

The first detailed studies on The structure of hemoglobin were conducted between 1896 and 1901 by Professor M. V. Nencki at the St. Petersburg Institute of Experimental Medicine. He was also the first to propose a structural formula for the non-protein component of hemoglobin, based on four pyrrole rings arranged around an iron atom, which correctly reflects the main Structural Features of the non-protein part of the molecule. It has now been established that Hemoglobins from various species consist of the protein globin and heme (ferroprotoporphyrin) linked non-covalently. Differences in the properties of hemoglobins among various animal species are primarily due to the globin component.

Heme. Heme is a relatively flat, nearly square molecule in which the Fe²⁺ ion is located at the center of the protoporphyrin IX core, with its four coordination sites occupied by the nitrogen atoms of the pyrrole rings. Protoporphyrins can exist in fifteen isomeric forms because they contain three Different types of substituents: four methyl groups, two vinyl groups, and two propionic acid residues. Protoporphyrin IX is the most common of the fifteen possible isomers. In addition to hemoglobin, it is found in Myoglobin and most cytochromes. The chelate complex of protoporphyrin with Fe²⁺ is called protoheme or heme.

In the presence of Hydrochloric acid or alkalis, heme is oxidized by molecular oxygen and other oxidizing agents to hemin, in which trivalent iron is bound to a chlorine anion, or to hematin, in which the third valence is occupied by a hydroxyl group. The porphyrin ring consisting of four pyrrole nuclei serves as the basis not only for heme, but also for plant chlorophyll, a fact already pointed out in 1896 by M. V. Nencki in his paper "On the Biological Relationship between Blood Pigment and Leaf Pigment." In it, the author made a broad generalization regarding The Unity of the PLANT AND ANIMAL worlds, linking it to Darwin's theory.

Globin. The protein component of hemoglobin, globin, in adults consists of two α-chains and two β-chains. The subunits are packed within the tetramer to form a spherical macromolecule measuring 6.4 nm in length and 5.5 nm in width. Each subunit contains a flat heme ring within one of its "folds" (the so-called "heme pocket"). The "heme pocket" is lined with a large number of non-polar amino acid groups that engage in hydrophobic interactions with the pyrrole rings of heme. The latter also forms a coordination bond with globin, established between the iron atom of heme and a nitrogen atom in the Histidine residue of globin. The four Hemes of the four subunits are oriented relative to the molecule as a whole such that one edge of each heme faces the interior of the molecule, while the other faces outward toward the aqueous-salt medium.

Inside the globin tetramer, There is a free cavity running through the entire height of the molecule, spanning 5 nm. This cavity is lined predominantly with non-polar amino acid residue groups and only a few polar ones. In an aqueous environment, hydrophobic interactions occur between these non-polar groups, which appear to partially shield the interior of the molecule from water contact and stabilize its overall structure. Using radioisotope labeling, experiments have shown that once formed, the hemoglobin in each erythrocyte does not undergo renewal and remains practically unchanged until the erythrocyte is destroyed. The average "lifespan" of erythrocytes is about 4 months. Following their destruction, the bulk of Fe (90%) is not excreted from the body, but is instead utilized to form a specialized protein called ferritin, which contains up to 30% iron. Thus, this protein serves as an iron storage depot. Iron from ferritin is subsequently supplied to the Bone Marrow for The production of new erythrocytes.

Hemoglobin Heterogeneity. Two or more hemoglobin fractions may simultaneously be present in an organism, differing primarily in the Primary Structure of their globin protomers (i.e., the sequence of their amino acid residues) and occasionally in their quaternary structure. This phenomenon is known as hemoglobin heterogeneity. Three types of heterogeneity are distinguished in human populations: 1. Heterogeneity caused by the presence of minor components: while major forms (HbA) consist of two a-chains and two ß-chains, minor Hbs contain two δ-chains instead of ß-chains, which differ from ß-chains by ten amino acid residues. 2. Genetic heterogeneity: up to 300 variants of genetically determined hemoglobins have been identified in adults. Many of these are functionally normal, while some cause diseases. 3. Embryonic heterogeneity: represented by two fractions of the so-called "primitive" hemoglobin, the principal one being HbF (fetal hemoglobin). Normally at birth, its content accounts for 60—70% of the total Hb. With age, The amount of HbF drops sharply, and it is absent in adults.

Properties of Hemoglobin. Cooperative Subunit Interaction. In the Hb macromolecule, subunits exhibit mobility during function, Ligand binding, and release. As they move, the subunits interact to produce a cooperative effect, The Essence of which is that the affinity of each subunit for a ligand is not constant, but changes under The Influence of neighboring subunits. Due to the cooperative effect, the relationship between Hb and O₂ changes under the influence of O₂ itself during oxygenation: initial O₂ binding induces a conformational change in Hb, facilitating the binding of subsequent oxygen molecules. As a result, the dependence of the Hb oxygenation degree on the partial pressure of O₂ is represented by a sigmoid curve.

The cooperative nature of O₂ binding by hemoglobin is of great physiological significance. In the pulmonary capillaries, at an O₂ partial pressure of about 100 mm Hg, cooperativity leads to nearly complete oxygen saturation of hemoglobin. When erythrocytes pass through the capillaries of oxygen-consuming tissues, the partial pressure of O₂ drops to approximately 5 mm Hg, and in this case, cooperativity facilitates a more complete unloading of oxygen from hemoglobin than would occur if the four heme groups acted independently.

Effect of pH. The concentration of H+ significantly alters the properties of Hb, most notably its ability to bind O₂. This phenomenon is known as the Bohr effect. In general terms, the Bohr effect is viewed as the Influence of Environmental pH on the interaction of the Fe atom in respiratory pigment molecules with various ligands—O₂, CO, NO. This also encompasses the pH dependence of The oxidation of Fe²⁺ to Fe³⁺.

In capillaries where the partial pressure of O₂ is low and CO₂ and lactic acid may accumulate, a decrease in pH causes oxyhemoglobin to release its oxygen more readily. In the Bohr effect, protons play the role of Allosteric regulators that bind to the amino and imidazole groups of globin. Another allosteric regulator of conformational equilibria in hemoglobin is 2,3-diphosphoglycerate (bisphosphoglycerate), the content of which in human erythrocytes is approximately equimolar relative to hemoglobin. Diphosphoglycerate binds between the two ß-chains of deoxyhemoglobin, thereby reducing the latter's affinity for oxygen, enabling erythrocytes to release a larger fraction of the transported O₂ to tissues. The concentration of diphosphoglycerate in erythrocytes varies depending on physiological conditions: individuals living in high-altitude regions have higher concentrations. The presence of diphosphoglycerate in erythrocytes is not characteristic of all animal species; in birds and turtles, it is apparently replaced by inositol pentaphosphate.

Forms of Hemoglobin. Oxyhemoglobin (HbO₂). The Fe atoms of each heme group in the Hb molecule can reversibly bind an O₂ molecule. Fully oxygenated Hb is called oxyhemoglobin and contains four O₂ molecules per hemoglobin molecule.

The valence of heme iron does not change during the formation of oxyhemoglobin. Hemoglobin possesses The unique ability to reversibly bind O₂, forming a stable complex without oxidizing Fe²⁺ to Fe³⁺. In heme, the four porphyrin ligand groups form a planar complex with iron. The remaining fifth and sixth coordination bonds of iron are positioned perpendicularly to the plane of the porphyrin ring. The fifth bond is occupied by the histidine imidazole residue, while the sixth either remains unoccupied (deoxyhemoglobin) or is replaced by oxygen (oxyhemoglobin). The oxidation of Fe²⁺ to Fe³⁺ in the reaction of O₂ with heme is possible only in solutions with a high dielectric constant, whereas the hydrophobic heme pocket, from which water is excluded, provides an environment with a low dielectric constant.

Carbaminohemoglobin. CO₂ can bind to Hb to form carbaminohemoglobin:

CO₂ binds exclusively to the N-terminal a-amino groups. The reaction is readily reversible. The formation of carbaminohemoglobin is determined by the partial pressure of CO₂ and is directly relevant to the transport of CO₂ by the blood.

Carboxyhemoglobin (HbCO). Hb can combine with four molecules of CO (carbon monoxide) to form CO-hemoglobin or carboxyhemoglobin, which is photosensitive and dissociates in the light with the release of CO. The heme Fe remains divalent in the process. The affinity of human Hb for CO is more than 200 times greater than its affinity for O₂; that is, the formation of HbCO requires a CO partial pressure 200 times lower than that required for HbO₂—O₂, respectively. As a result, when breathing air containing carbon monoxide, the bulk of blood hemoglobin converts to carboxyhemoglobin, oxyhemoglobin is not formed, and the transport of O₂ from the Lungs to tissues is impaired, which constitutes The Mechanism of Carbon monoxide poisoning. Death occurs due to inadequate oxygen supply to tissues (primarily the brain) once 70% of Hb is bound to carbon monoxide. Timely elevation of O₂ partial pressure (breathing pure oxygen) can induce sufficient conversion of HbCO back to HbO₂.

Methemoglobin (Met-Hb). Peroxides, ferricyanide, nitrogen oxides, and Quinones can oxidize Fe²⁺ in hemoglobin to Fe³⁺, forming methemoglobin, which binds neither O₂ nor CO. It has a brown color, forms normally in vivo in small amounts, and is enzymatically reduced back to Hb. Since Met-Hb cannot serve as an O₂ carrier, the accumulation of significant amounts of it in the body (e.g., in poisonings with aniline, nitrobenzene, or nitrogen oxides) leads to oxygen starvation and, in severe cases, death. However, The conversion of a small fraction of hemoglobin into methemoglobin is less dangerous than the formation of HbCO, because Met-Hb is gradually reduced back to hemoglobin in the body.

Functions of hemoglobins. The Main Functions of hemoglobins are: 1) transport of O₂, 2) transport of CO₂, and 3) Maintenance of the constant buffer capacity of the blood.

In the lungs, due to the existing O₂ gradient, oxygen diffuses through the capillary walls and plasma into the erythrocytes. Arterial blood Hb is 96% saturated with O₂. In tissues, O₂ diffuses from erythrocytes through plasma into the interstitial fluid and then into tissue cells, while CO₂ diffuses in the opposite direction.

Transport of CO₂ from tissues to alveolar air is carried out in small part as carbaminohemoglobin. The major part of CO₂ is hydrated by Carbonic anhydrase into H₂CO₃, which then dissociates. About 60% of CO₂ is transported as HCO^-₃ by venous plasma, and about 32% as carbamino-CO₂ and HCO3- by erythrocytes. However, the transport of 60% HCO^-₃ by venous plasma is also indirectly supported by Hb, as it acts as a hydrogen acceptor during the dissociation of H₂CO₃.

Hemoglobin is a functionally weaker acid than HbO₂. These two proteins form a buffer system that helps maintain blood pH at a constant level.

Hemoglobin metabolism. The lifespan of erythrocytes is 120 days. Every day, 8–9 g of Hb is released from destroyed erythrocytes. Erythrocyte destruction and hemoglobin breakdown occur mainly in the liver, Spleen, and bone marrow. Hb breakdown begins with the cleavage of a single α-methine bridge (between the 1st and 2nd pyrrole rings) by the action of an NADP-containing oxidase. This yields a green pigment called verdoglobin, which still contains iron and globin. Further degradation presumably proceeds spontaneously. Iron and globin are split off, forming biliverdin, one of the Bile pigments. Biliverdin is reduced by an NADPH-dependent dehydrogenase to bilirubin, which passes from the liver along with bile into the Gallbladder.

In the blood of an adult human, the bilirubin level is relatively constant, ranging from 2.5 to 12 mg/L. An increase in blood bilirubin concentration up to 20 mg/L leads to jaundice, and a further rise causes severe toxic symptoms. Upon entering the liver with the blood flow, bilirubin is detoxified by conjugation with glucuronic acid. The final breakdown of bilirubin takes place in the intestine under the action of bacteria: glucuronic acid is cleaved off, and bilirubin is reduced to stercobilinogen, which is excreted in the feces.

Myoglobin. Myoglobin is a small globular protein (M 17,000); its molecule consists of a single polypeptide chain (153 amino acid residues) and a single heme group. Thus, myoglobin represents approximately ¹/₄ of an Hb molecule. The complete amino acid sequence of the myoglobin polypeptide chain is known. It varies slightly among humans and different animal species, but the folding pattern of the chain into a secondary helix and tertiary configuration is largely identical. The molecule forms eight segments of right-handed α-helices. The heme is located in the cleft between the segments. Similar to hemoglobin, it forms oxymyoglobin, carboxymyoglobin, and metmyoglobin.

Types of Oxygen transport to tissues in animals of various evolutionary positions. According to P. A. Korzhuev (1964), in approximately 86% of animal species, O₂ transport occurs without the participation of Hb-type respiratory pigments. Among these, in 78% of species (insects, myriapods, most arachnids), O₂ is delivered to all tissues and organs by a tracheal system, while in the remaining 8% of "pigment-free" species (Protozoans, sponges, Coelenterates, mollusks, Echinoderms), it occurs via diffusion through the body surface. Respiratory pigments are present in the blood or body cavity fluid of only 14% of species, but these are the most highly organized animals, including all vertebrates.

In the most primitive animals, the O₂ carrier is enclosed within cells suspended in the coelomic fluid. With The Development of Circulation, O₂ carriers soluble in the circulating plasma emerged. A major milestone in evolution was the appearance of specialized cells—erythrocytes—which concentrate high levels of carriers, thereby preventing a sharp increase in the viscosity and colloid Osmotic Pressure of the circulating blood.

The most widespread respiratory pigment is hemoglobin, which is found in almost all animal groups, ranging from protozoans to vertebrates. Chlorocruorin is structurally similar to hemoglobin. It is found in the blood of certain Annelids. It contains iron, but the porphyrin of chlorocruorin differs from the protoporphyrin of heme in that the vinyl group at the 2nd pyrrole ring is replaced by a formyl group.

Hemerythrin is found in the blood of marine worms. It occurs in a small number of polychaetes and in a single brachiopod species. The subunits of this protein typically form octamers, and each monomer possesses an active center containing two atoms of non-heme Fe²⁺. Hemocyanins—blue copper-containing pigments lacking a heme group—are found in the blood plasma of many mollusks and Arthropods.

2.5.4. Blood plasma proteins. Representatives and functions. If blood is prevented from clotting and the formed elements are removed by centrifugation, the transparent light-yellow supernatant liquid represents blood plasma. During blood clotting, fibrinogen, a plasma protein, forms fibrin threads that, together with the formed elements of the blood, produce a clot. The liquid portion of clotted blood is called serum. Consequently, plasma differs from serum by the presence of the protein fibrinogen. Proteins dissolved in plasma account for about 7% of its mass. Using Electrophoresis, five major protein fractions have been detected in serum: albumin, accounting for 54–58%; α₁-globulins, 6–7%; α₂-globulins, 8–9%; β₁-globulins, 13–14%; and γ-globulins, 11–12%.

Starch gel electrophoresis additionally reveals prealbumin, β-lipoprotein, and transferrin fractions. Immunoelectrophoresis also isolates β₁-lipoprotein, α₁- and α₂-lipoproteins, haptoglobin, ceruloplasmin, and IgG globulin.

Prealbumin. Its plasma concentration is relatively low—10–40 mg/100 mL. Its function is to bind and transport thyroxine and retinol-binding protein.

Serum albumin is present in plasma in significant amounts—3500–4500 mg/100 mL. It is one of the few plasma proteins that are not glycoproteins.

The main functions of albumins are osmotic regulation and transport. About 75–80% of the osmotic effect of plasma is associated with albumin because, among the major plasma proteins (where it constitutes more than half by mass), albumin has the lowest molecular weight. The transport function of albumins is manifested in the carrying of free fatty acids from the liver, bilirubin to the liver (where it is excreted in the bile), and steroid hormones. A number of drugs entering the bloodstream form stable complexes with albumin. Albumin can participate in the removal of toxic substances, such as heavy metals. Albumins play a major role in tissue Nitrogen metabolism, and their levels can serve as an indicator of the replenishment of the body's protein reserves.

α-Globulins. They are designated as α₁ and α₂-globulins based on their electrophoretic mobility. α₁-Globulins include: α₁-acid glycoprotein (retinol-binding protein), α₁-antitrypsin, transcortin (which binds and transports cortisol and corticosterone), and thyroxine-binding protein. α₂-Globulins include: ceruloplasmin, haptoglobins, α₂-macroglobulin, and inter-α-Trypsin inhibitor.

Ceruloplasmin is the main copper-containing blood protein, classified as a glycoprotein. It has a blue color and accounts for 3% of the copper contained in the body. Ceruloplasmin participates in Cu transport and in maintaining its level in tissues, especially the liver. Ceruloplasmin exhibits enzymatic properties, specifically polyamine oxidase and ferroxidase activity. The latter is manifested in the catalysis of the oxidation of Fe²⁺ to Fe³⁺. The Importance of this reaction lies in the fact that only Fe³⁺ can bind to transferrin, the iron-transporting protein.

Haptoglobins (Hp). These are α-globulins of glycoprotein structure capable of binding to hemoglobin, which increases the latter's stability and extends its lifespan—this is the primary function of haptoglobins. Hps also perform a nonspecific protective function by complexing with various protein and non-protein substances released during cell breakdown, protecting tissues from proteolysis and participating in detoxification processes.

β-Globulins. These comprise a wide range of proteins, including lipoproteins. Transferrin (Tf), or siderophilin, is the main component of this fraction. It readily forms a complex compound with Fe, which under certain conditions dissociates just as easily. Due to this property, transferrin performs an important physiological function by converting plasma iron into a stored form and delivering it to the bone marrow (where iron is used for hematopoiesis) and other tissues, particularly those of the reticuloendothelial system.

Hemopexin. It binds heme, preventing its excretion in the urine.

Lipoproteins. They are present in blood plasma in amounts of 700–1100 mg/100 mL. Human blood contains several lipoprotein fractions that differ in density due to the varying ratios of lipid and protein components in the molecule. The lowest density (less than 0.95 g⋅cm^-3) is characteristic of chylomicrons, in which lipids account for about 98%. Chylomicrons are lipid droplets stabilized by a thin surface layer of proteins. The lipids here are represented mainly by triacylglycerols (80%). β-Lipoproteins have a higher density, with an average molecular weight of 10 million. These plasma lipoproteins are further subdivided into pre-beta-lipoproteins, or very low-density lipoproteins (VLDL), and beta-lipoproteins, or low-density lipoproteins (LDL).

Plasma lipoproteins with a density of 1.06–1.20 g⋅cm^-3 are called α₁-lipoproteins or high-density lipoproteins (HDL), and their lipid content is approximately 65%. The lowest lipid content is characteristic of α₂-lipoproteins, or very high-density lipoproteins (VHDL, density greater than 1.2 g∙cm^-3). Lipids account for 43% of their mass. α-Lipoproteins have a molecular weight of about 300,000, with phospholipids predominating. The classification of plasma lipoproteins is conventional, as their composition and density change during the transport of lipids to tissues.

Blood lipoproteins are spherical particles whose diameter decreases with increasing density. The core of these particles contains nonpolar lipids (triacylglycerols, esterified cholesterol). The core is surrounded by a shell composed of phospholipids, protein, and free cholesterol. Currently, the role of certain lipoprotein fractions in the Pathogenesis of atherosclerosis has been proven, and they are referred to as atherogenic lipoproteins.

Atherosclerosis develops with a significant increase in the blood LDL fraction, and in many cases also VLDL. Atherosclerosis is a lipid degeneration of arterial walls, plaque formation, and luminal narrowing. Chylomicrons cannot penetrate the vessel wall due to their large size. HDL particles have the smallest size, easily penetrate the wall, but are also easily cleared from it into the Lymph. Furthermore, HDL has the highest percentage of proteins and phospholipids, which is why they are rapidly metabolized in the wall without depositing in it. Therefore, LDL and VLDL are atherogenic; they penetrate the wall quite effectively and are very rich in cholesterol and triacylglycerols (fats), which trigger atherosclerosis. There is evidence that LDL receptors—proteins located on the surface of body cells that extract LDL particles from the blood—play a crucial role in regulating blood LDL and VLDL levels. The LDL particles then enter the cells, where cholesterol is released and metabolized. The number of receptors can be genetically determined or dependent on a number of other factors.

The blood clotting mechanism. The biological and biochemical processes that prevent and stop bleeding in the body are referred to as hemostasis. It is maintained by three key components: 1) platelets, 2) the vascular wall, and 3) the plasma enzymatic coagulation system. The latter includes a number of substances (clotting factors) which, in accordance with the recommendations of the International Nomenclature Committee, are designated by Roman numerals (the letter 'a' next to a factor number indicates its active form). Below is a description of several clotting factors.

Factor I (fibrinogen) is present in the blood plasma of warm-blooded animals at a concentration ranging from 0.17 to 0.4 g/100 ml. It is a glycoprotein featuring three pairs of non-identical polypeptide chains linked by Disulfide Bonds. It is synthesized in the liver.

Factor II (prothrombin) is a glycoprotein that plays a central role in Blood Coagulation. It acts as the proenzyme of Thrombin (Factor IIa).

Factor IIa (thrombin) belongs to the serine proteinase family. Through the cleavage of peptide bonds, it converts fibrinogen into fibrin and transforms Factor XIII into Factor XIIIa.

Factor XII is a key enzyme in the intrinsic activating system of prothrombin and possesses a glycoprotein structure. The activation of Factor XII, achieved through its adsorption onto surfaces other than the normal endothelial lining of Blood Vessels, serves as the trigger for a chain reaction of sequential activations of specific proteinases within the BLOOD COAGULATION SYSTEM.

Factor XIII is the proenzyme of transglutaminase. Upon conversion into its active form (XIIIa), it catalyzes the synthesis of covalent bonds that link monomeric units within fibrin together, thereby converting soluble fibrin S into insoluble fibrin I.

The kallikrein-kinin system participates in activating the Initial Stages of blood coagulation and in forming the "kallikrein bridge" between Factors XIIa and VII (Fig. 2.22).

The most important plasma kinins are the peptides bradykinin, kallidin, and methionyl-lysyl-bradykinin (see Section 2.5.1). The substrates from which kinins are released are called kininogens. These are proteins bound in the blood plasma to the alpha-2-globulin fraction. The formation of kinins from kininogens occurs through the action of specific enzymes—kallikreins, which are trypsin-like proteinases.

Fig. 2.22. Cascade-complex scheme of blood coagulation:

fibrin S and fibrin I — soluble and insoluble fibrin, a — activated factors, pf — platelet factor

The involvement of platelets in hemostasis is determined by their core functions: a) angiotrophic, i.e., the ability to maintain the normal Structure and function of microvessels (up to 100 µm in diameter); b) the ability to form a primary platelet plug in damaged vessels (adhesive-aggregatory function); c) the ability to sustain vasospasm in injured vessels; d) participation in blood coagulation and an inhibitory effect on Fibrinolysis. If vascular damage is minor, the platelet plug can halt bleeding.

Platelets periodically make contact with endothelial cells and "discharge" their contents into them, serving as natural "feeders" for the endothelium, which is otherwise unable to extract several essential substances directly from the plasma.

The formation of a platelet plug begins with the adhesion of platelets to the subendothelial structures of the vessel wall. Vascular wall collagen serves as the primary stimulus for this process. Platelets actively adhere to one another to form aggregates. One of the aggregating agents is thrombin; it induces platelet aggregation at doses several times lower than those required for blood coagulation. Consequently, platelet plug formation outpaces blood clotting.

Today, blood coagulation is viewed as a multi-step cascade enzymatic process in which proenzymes are sequentially activated, operating via autacoidal mechanisms that function both top-down and through feedback loops. In recent years, it has become clear that protein-lipid complexes form at various stages of the clotting process, within which enzymatic factors are activated. The non-enzymatic components of these complexes accelerate the process and ensure the specificity of coagulating enzymes by creating additional binding sites within the enzyme-substrate complexes. The cascade-complex scheme of blood coagulation includes four types of complexes.

Complex 1: Factors XIIa + XI + phospholipid = Factor II activator.

Complex 1a: Factor III + Factor VII + Ca²⁺ = Factor X activator (extrinsic pathway).

Complex 2: Factors IXa + VIII + Ca²⁺ + phospholipid = Factor X activator (intrinsic pathway).

Complex 3: Factors Xa + V + Ca²⁺ + phospholipid = prothrombin activator (prothrombinase).

Coagulation is carried out through two closely interconnected mechanisms: the extrinsic and intrinsic pathways (Fig. 2.22).

The extrinsic coagulation pathway is triggered by tissue thromboplastic factor (Factor III), which interacts with Factor VII and, in the presence of Ca²⁺, forms Complex 1a (Factor X activator). The latter, acting as a constituent of Complex 3, transforms Factor II into Factor IIa.

The intrinsic coagulation pathway is triggered without the addition of exogenous tissue thromboplastin—that is, relying entirely on the internal resources of the blood or plasma. Initiation of the intrinsic pathway begins with the activation of Factor XII. This activation can be brought about by collagen from the damaged vessel wall, altered cell membranes, certain proteases, or adrenaline. Outside the body, Factor XII activation occurs upon contact with foreign surfaces such as Glass, needles, etc. Following Factor XII, Factors XI (as part of Complex 7), IX, and VIII are sequentially activated. The latter are components of Complex 2, which activates Factor X—once again generating the prothrombinase activity necessary to convert prothrombin into thrombin. Thrombin (IIa) and Factor XIII are then required to convert fibrinogen into fibrin and form a blood clot.

This process consists of three stages. The fibrinogen molecule contains three pairs of non-identical polypeptide chains linked by disulfide bonds (2aA, 2ßB, 2γ). In the first, proteolytic stage, thrombin cleaves fibrinopeptides A from the α (A)-chains and fibrinopeptides B from the β (B)-chains, leaving behind the α- and β-chains that comprise the newly formed fibrin monomer. Thus, the polypeptide formula of the fibrin monomer is (α, β, γ)2. In the subsequent polymerization stage, monomeric molecules join together side-by-side and end-to-end to form a fibrin network with the polypeptide formula (α, β, γ)_n. In the final stabilization stage, the fibrin network is stabilized by covalent bonds under the action of an enzyme generated from Factor XIII.

The enzymatic system responsible for the lysis of fibrin within the bloodstream is known as the fibrinolytic or plasmin system. This lysis is carried out by fibrinolyticin, or plasmin, which exists in blood plasma as the proenzyme plasminogen at a concentration of about 20 mg%. Plasminogen activators include proteinases, such as snake venom and streptokinase from hemolytic streptococci, while urokinase acts as the physiological activator. Plasmin hydrolyzes peptide bonds in fibrin formed by arginine and Lysine residues.

Anticoagulants are substances that prevent blood coagulation. They can be divided into two main groups: physiological and non-physiological. The former include heparin, antithrombins, antiphospholipids (or antithromboplastins), etc. Non-physiological anticoagulants include substances that precipitate Ca²⁺ from plasma (oxalates, citrates), vitamin K antagonists (coumarin derivatives), biological poisons (from snakes or leeches), and physical-Chemical factors (such as Temperature or ultrasound).