BIOCHEMISTRY - Textbook - Ostapchenko L. I. - 2012

Chapter 5. STRUCTURE, PROPERTIES, AND FUNCTIONS OF PROTEINS

5.6. Protein diversity and classification

5.6.3. Functional classification of proteins

Proteins perform a wide variety of biological Functions within Cells. Based on functional similarities, Proteins can be divided into several major groups: Enzymes, regulatory proteins, receptors, transport proteins, structural proteins, protective proteins, and contractile proteins.

Enzymes are specialized proteins that accelerate Chemical Reactions. Thanks to enzymes, The rate of cellular chemical reactions increases by millions of times. Since enzymes, like any proteins, possess an Active Site, they specifically bind a certain Ligand (or a group of similar ligands) and catalyze a specific type of chemical transformation of that molecule. Currently, over 2,000 different enzymes are known that accelerate various enzymatic reactions. For example, the proteolytic enzyme Trypsin cleaves peptide bonds in proteins formed by the carboxyl group of basic Amino Acids, namely Arginine or Lysine, whereas the enzyme Ribonuclease cleaves the phosphoester bond between NUCLEOTIDES in a polynucleotide chain. The Complement of enzymes in cells ensures that metabolic transformations occur not chaotically, but in precisely directed pathways.

Regulatory proteins include a large group of protein Hormones Involved in maintaining the body's internal environment and acting on specific target cells. For instance, the hormone Insulin is released into the bloodstream when Blood glucose concentrations rise. By stimulating glucose utilization by cells, it lowers its concentration back to normal, thereby restoring Homeostasis.

Furthermore, regulatory proteins also include those whose binding to other proteins or certain cellular structures regulates their function. For example, the protein calmodulin, in a complex with four Ca²⁺ ions, can bind to certain enzymes, altering their activity.

Regulatory DNA-binding proteins, by binding to specific DNA regions at certain moments, can regulate the rate at which Genetic information is read.

Signaling molecules (hormones, Neurotransmitters) influence intracellular processes through interaction with specific receptor proteins. Thus, hormones circulating in the bloodstream locate target cells and act upon them by specifically binding to receptor proteins embedded in The Cell membrane. For hydrophobic regulatory molecules that cross the cell membrane, receptors are localized in the Cell Cytoplasm.

Blood transport proteins are involved in carrying specific ligands from one organ to another. Molecules that are poorly soluble in Water are often transported in a complex with proteins. For example, the Blood Plasma protein albumin transports Fatty acids and bilirubin (a heme breakdown product), while erythrocyte Hemoglobin participates in transporting O₂ from the Lungs to Tissues. Steroid Hormones are transported in the blood by specific transport proteins.

Transport proteins also participate in carrying hydrophilic substances across hydrophobic membranes. Because transport proteins have the property of specifically interacting with ligands, their set in the cell membrane determines which hydrophilic molecules can enter the cell. Glucose, amino acids, ions, and other molecules penetrate the cell with the aid of carrier proteins.

Structural proteins, positioned in specific ways within tissues, impart shape, form a framework, and determine the mechanical properties of a given tissue. For instance, the primary component of Cartilage and tendons, as mentioned above, is the highly durable fibrous protein Collagen. Another structural protein, Elastin, due to its unique Structure, provides certain tissues with The ability to stretch in all directions (Blood Vessels, lungs).

Protective proteins, particularly IMMUNOGLOBULINS, have the ability to recognize and bind foreign molecules, Viral Particles, and Bacteria, resulting in their neutralization. In addition, the complex formed by a foreign particle and an immunoglobulin is readily recognized and destroyed by cells of The Immune System.

Proteins of the blood clotting system, such as fibrinogen and Thrombin, also possess protective properties. They participate in forming a blood clot that plugs the damaged vessel and prevents blood loss.

Contractile proteins grant the cell the ability to either contract or move when performing their functions. These include Actin and Myosin, which are Fibrous proteins involved in skeletal Muscle contraction. Another example of such proteins is tubulin, which makes up cellular Organelles known as microtubules. The latter regulate chromatid segregation during Cell Division. Microtubules are also essential elements of Cilia and flagella, which enable cellular motility.

However, There is a vast number of proteins with unique functions that do not fit into this relatively simple Classification.

Families of related proteins. During evolution within a single biological species, Amino Acid Substitutions can lead to The Emergence of different proteins that perform related functions and possess homologous Amino acid sequences. Sequences that share many similarities are termed homologous. They contain the same amino acids—referred to as invariant residues—at many positions, while at other positions they may feature different amino acid residues that are similar in their physicochemical properties.

Homologous proteins share similar Conformations: the number and relative arrangement of α-helices and/or β-structures, as well as most turns and bends of The polypeptide chains, are similar or identical. Proteins with homologous Regions of the polypeptide chain, a similar conformation, and related functions are grouped into a protein family.

An example of a family of related proteins is the Myoglobin family, which includes, alongside myoglobin itself, all types of hemoglobin.

Serine proteases are classified among related Protein Families. This is a family of enzymes that utilize a specifically activated serine residue located in the active site. The targets for serine proteases are specific peptide bonds in proteins (often in other serine proteases).

All proteins of this family are characterized by the presence of Ser₁₉₅, His₅₇, and Asp₁₀₂ residues in their active site (this numbering is used regardless of their exact position in the Introduction/19.html">Primary Structure of specific serine proteases). A high degree of Spatial Structure similarity has also been revealed, despite the fact that only 40% of their positions contain identical amino acids (Fig. 5.43). The catalytic domain of serine proteases is located in a cleft between two domains.

Class="center">

Fig. 5.43. Spatial structures of Elastase (A) and Chymotrypsin (B)

Certain amino acid substitutions have led to Changes in the substrate Specificity of these proteins and to the emergence of functional diversity within the family itself. For instance, digestive serine proteases participate in Digestion (the hydrolytic Cleavage of peptide bonds) of denatured dietary proteins. These include trypsin, chymotrypsin, and elastase, yet each of these enzymes cleaves peptide bonds formed by specific amino acids.

Serine proteases also exhibit substrate specificity and are involved in strictly regulated physiological processes, such as the activation of the Blood Coagulation cascade, Fibrinolysis, The Complement System, and The formation of Peptide Hormones. During the activation of native proteins, serine proteases hydrolyze one or two specific peptide bonds out of hundreds present in the protein substrate. This occurs because the enzyme recognizes not only the amino acids directly forming the peptide bonds in the native protein, but also certain amino acid residues surrounding the bond undergoing Enzymatic Hydrolysis.

Proteins belonging to the immunoglobulin superfamily play a major role in the immune system. This superfamily comprises three large families of proteins involved in the body's immune defense: the immunoglobulin family, the T-cell antigen receptor family, and the Major Histocompatibility Complex (MHC) class I and class II proteins. The superfamily also includes the family of adhesion proteins, which are involved in cell-type recognition and intercellular interactions.

The primary criteria for including proteins in the immunoglobulin superfamily are their domain Organization, significant Amino Acid Sequence Homology, and the spatial arrangement of individual Structural domains. Furthermore, proteins of this superfamily share similar functions: immunoglobulins interact with foreign structures found in blood, lymph, interstitial fluid, or glandular secretions, whereas T-cell receptors and major histocompatibility complex proteins interact with Antigens located On the surface of the Organism's own cells.

The immunoglobulin family. Immunoglobulins, or Antibodies, are specific proteins produced by B lymphocytes in response to The entry of foreign structures—antigens—into the body. The Human Body produces approximately 10⁷ clones of B lymphocytes, each specialized in producing one of the 10⁷ types of immunoglobulins.

All immunoglobulins share a common structural plan, which can be illustrated using The structure of IgG. An IgG molecule consists of four polypeptide chains: two identical light (L) chains, containing about 220 amino acid residues, and two heavy (H) chains, consisting of 440 amino acids each. All four chains are linked together by numerous non-covalent bonds and four Disulfide Bonds. Consequently, the IgG molecule is classified as a monomer.

The light chains of IgG consist of two domains: a variable domain (V_L) located at the N-terminal region of the polypeptide chain, and a constant domain (C_L) situated at the C-terminus. Each domain consists of two anti-parallel β-sheet layers. The β-sheet is covalently linked by a disulfide bond located approximately in the middle of the domain (Fig. 5.44).

The heavy chains of IgG contain four domains: one variable domain (V_H) located at the N-terminus, and three constant domains (C_H1, C_H2, C_H3). The domains of IgG heavy chains are homologous in structure to those of the light chains. Between the two constant domains of the heavy chains, C_H1 and C_H2, lies a region rich in Proline residues that prevent the Formation of secondary structure and the interaction of adjacent H-chains in this segment. This region is called the hinge region, and it imparts flexibility to the molecule.

Fig. 5.44. Structure of immunoglobulin G

Between the variable domains of the heavy and light chains lie two identical regions that bind two identical specific antigens; therefore, such antibodies are often referred to as bivalent. Antigen-antibody binding does not involve the entire amino acid sequence of the variable domains of both chains, but rather only 20–30 amino acids located in the hypervariable regions of each chain. It is these regions that determine The unique ability of each antibody clone to interact with its corresponding (complementary) antigen.

The primary functions of antibodies are the detection and binding of foreign antigens located outside the body's cells (in the blood, lymph, interstitial fluid, and mucosal secretions). This process is mediated by specific antigen-binding sites of various immunoglobulin clones. Moreover, antigen-antibody binding facilitates the subsequent destruction of foreign substances. The specific pathway of antigen-antibody complex destruction depends on the antibody class.

There are five classes of immunoglobulin heavy chains, which differ in the structure of their constant domains: α, δ, ε, γ, and μ. Correspondingly, there are five classes of immunoglobulins: A, D, E, G, and M. The Structural Features of heavy chains are determined by their hinge and C-terminal regions, which adopt a conformation characteristic of each class. The binding of an antigen to an antibody alters the conformation of the HEAVY CHAIN CONSTANT domains, thereby directing The pathway of complex degradation in the body (either via binding to complement proteins or uptake by phagocytic cells).

Immunoglobulin M represents the first class of antibodies synthesized in developing B lymphocytes. There are two forms of immunoglobulin M: a monomeric, membrane-bound form, and a pentameric form secreted by B lymphocytes into the bloodstream.

Maturing B lymphocytes synthesize monomeric bivalent IgM molecules that are structurally similar to the IgG molecules discussed above. These are embedded in The Plasma Membrane and function as primary antigen-recognition receptors. Attachment of IgM to the membrane is mediated by a hydrophobic region located in the C-terminal ("tail") portion of the heavy chains, which contains 25 hydrophobic amino acid residues.

The interaction of an antigen with a receptor on The surface of a B lymphocyte triggers its proliferation and the formation of a clone of lymphocytes derived from a single antigen-stimulated cell. This B-lymphocyte clone will produce immunoglobulins with identical antigen-binding sites. However, B lymphocytes are capable of class switching to produce other antibody classes.

When B lymphocytes first encounter a novel antigen in Body Fluids, they synthesize and secrete into the blood IgM molecules, which consist of five monomeric subunits linked together by disulfide bonds and an additional polypeptide J-chain (Fig. 5.45).

Fig. 5.45. STRUCTURE OF THE pentameric secretory immunoglobulin M molecule

The heavy chains of their monomers lack the hydrophobic "tail" region. The pentameric molecule contains ten antigen-binding sites, which increases the probability of capturing a previously unencountered antigen (Fig. 5.46).

Fig. 5.46. Binding of IgM to bacterial cell antigens and their destruction by activated complement proteins

The interaction of an antigen with IgM induces a conformational change and triggers the binding of its "tail" region to the first component of the complement system. If the antigen is located on the surface of a microorganism, complement activation leads to the disruption of the cell membrane integrity and the death of the bacterial cell.

Quantitatively, immunoglobulins G predominate in the blood, accounting for about 75% of total Serum proteins. The structure of IgG has been described in detail above. In the bloodstream, IgGs exist exclusively in monomeric form; they are secreted in large quantities by activated B lymphocytes during the secondary Immune Response upon re-exposure to an antigen.

Four subclasses of IgG have been identified in humans: IgG₁, IgG₂, IgG₃, and IgG₄. The numerical subscript reflects the relative concentration of each subclass in serum (IgG₁ is the most abundant, whereas IgG₄ is the least abundant). The degree of Sequence homology among these subclasses is very high (approximately 90–95%).

IgGs not only effectively bind and neutralize foreign molecules and cells that have entered the body, but also facilitate their subsequent elimination. Conformational Changes in the "tail" region of IgG following antigen binding lead to the recruitment and activation of complement proteins. Furthermore, the C-terminal region of IgG can interact with specific receptors on macrophages and neutrophils, driving the phagocytosis of antigen-antibody complexes and their subsequent destruction within phagolysosomes (Fig. 5.47).

IgG is the only class of antibodies capable of crossing the placental barrier and providing intrauterine protection for the fetus against infections.

Fig. 5.47. Phagocytosis of an antigen-antibody complex by a neutrophil:

A – interaction of an IgG-coated bacterium with neutrophil receptors; B – engulfment of the bacterium by the neutrophil;

C – digestion of the bacterium within the neutrophil phagosome

Immunoglobulins A represent the primary class of antibodies found in body secretions (saliva, milk, digestive juices, and respiratory secretions). In blood serum, its concentration does not exceed 10-15% of the total immunoglobulin pool. The monomeric form structurally resembles IgG. However, in secretions, IgA predominantly exists as a dimer, where the monomers are linked by an additional J-peptide chain (Fig. 5.48).

Fig. 5.48. Structure of a dimeric immunoglobulin molecule

At the basal surface of epithelial cells, the IgA dimer specifically interacts with cell-surface proteins known as the secretory component. The resulting complex is translocated intracellularly via endocytosis and transported to the apical region. Here, the complex undergoes proteolytic cleavage, releasing the free dimer into the extracellular space (Fig. 5.49).

Fig. 5.49. Transport of immunoglobulins A across epithelial cells into glandular ducts

The complex formed during the interaction of IgA with an antigen does not activate complement proteins or phagocytic cells; however, it prevents antigens from attaching to the surface of epithelial cells and penetrating the body.

The concentration of immunoglobulin E in the blood is quite low. IgE molecules are monomers, but unlike IgG, their heavy chains contain four constant domains rather than three. Following Synthesis and Secretion into the bloodstream by B lymphocytes, IgE antibodies bind via their C-terminal regions to specific receptors on the surface of mast cells and basophils (Fig. 5.50).

Fig. 5.50. Release of BIOLOGICALLY ACTIVE SUBSTANCES by a mast cell upon binding of an antigen to surface-fixed IgE: 1 - granules filled with histamine, serotonin, and other mediators; 2 - release of histamine and other mediators into the Extracellular matrix

When an antigen binds to at least two antigen-binding sites on two adjacent IgE molecules, the cell receives a signal to secrete biologically active substances (serotonin, histamine) stored in secretory vesicles. The release of these substances is largely responsible for The Development of inflammatory responses as well as allergic conditions such as asthma, urticaria (hives), and hay fever. Elevated levels of IgE can precede the onset of allergic reactions.

Immunoglobulin D is found in the blood in very small amounts. These monomeric proteins function as B-lymphocyte receptors; no other functions for IgD have been identified as of yet.

The family of antigen-recognizing T-cell receptors. While antibodies produced by B lymphocytes bind antigens in body fluids (mediating humoral Immunity), T lymphocytes interact with antigens presented on the surface of virus-infected and tumor-transformed host cells (cell-mediated immunity). T lymphocytes recognize antigens only when presented in a complex with MHC class I or class II molecules, which are also present on the cell surface.

T-cell receptors are heterodimers, meaning they consist of α- and β-chains. Each chain features two immunoglobulin-like domains: a variable (V) and a constant (C) domain (Fig. 5.51). The C-terminal regions of each chain are embedded in the plasma membrane. A single antigen-binding site is located between the two variable domains, V_α and V_β. The Diversity of T-cell receptors with different antigen-binding sites is comparable to that of immunoglobulins.

Fig. 5.51. Structure of the T-cell receptor

The family of major histocompatibility complex proteins. Major histocompatibility complex (MHC) proteins were discovered during studies on intraspecific tissue transplantation, which gave rise to their name. They are also referred to as MHC proteins (see above) or HLA proteins (human leukocyte antigens), as they were first identified on human lymphocytes.

There are two main classes of MHC molecules: class I and class II. MHC class I molecules are expressed on the surface of virtually all cells in the human body, whereas MHC class II proteins are found exclusively on certain Cells of the immune system known as antigen-presenting cells. These primarily include macrophages and B lymphocytes that interact with antigens.

MHC class I molecules are heterodimers. They consist of a single polypeptide α-chain non-covalently linked to a small extracellular protein, β₂-microglobulin. The polypeptide α-chain possesses three extracellular globular domains (α₁, α₂, α₃), a transmembrane region, and a carboxy-terminal tail located in the cytoplasm (Fig. 5.52, A). The α₃ domain and β₂-microglobulin adopt an immunoglobulin-like conformation. The α₁ and α₂ domains contain variable regions capable of binding an "unfolded" antigen (most commonly a peptide fragment of a foreign protein) displayed on the cell surface.

MHC class II molecules are likewise heterodimers. They are composed of two polypeptide chains, α and β, each containing one conserved immunoglobulin-like domain and one variable domain at their N-terminal regions. Antigen binding occurs within the region formed by the variable domains of the α- and β-chains (Fig. 5.52, B).

Fig. 5.52. Structure of major histocompatibility complex (MHC) proteins: MHC class I (A) and MHC class II (B)

Foreign proteins within human cells (such as viral proteins) undergo Limited proteolysis in Lysosomes, and small fragments of these proteins, along with MHC class I or II molecules, are displayed on the cell membrane surface.

Peptide-MHC Protein Complexes are recognized by T-lymphocyte receptors. This results in a specific interaction (Fig. 5.53), T-lymphocyte activation, and the development of an immune response. For instance, the interaction between a cytotoxic T-lymphocyte and the antigen-MHC I complex on the surface of a virus-infected cell prompts the lymphocyte to release specialized proteins that cause damage and destruction of the infected cell.

Isofunctional proteins are a family of proteins that perform virtually identical or similar functions, although subtle Structural and functional differences among certain members of this family can have significant physiological importance. Examples of such proteins include the human hemoglobin isoforms HbA, HbA₂, HbF, and others discussed above. All of them are tetramers, but they consist of different sets of α, β, γ, and δ protomers. While all Hemoglobins share the same function—binding O₂ and transporting it to tissues—each possesses distinct functional characteristics. For example, fetal hemoglobin (HbF) has a higher affinity for O₂ than HbA, which facilitates the diffusion of O₂ from maternal HbA in the blood to fetal HbF in the placental Circulation.

Fig. 5.53. Specific interaction between a cytotoxic T-lymphocyte receptor and the antigen-MHC I protein complex:

1 - virus-infected target cell; 2 - peptide antigen on the cell surface bound to MHC I;

3 - cytotoxic T-lymphocyte; TK — Tyrosine kinase

Isoproteins are Multiple Forms of a protein found within organisms of the same species. Proteins that perform identical functions in organisms of different biological species are termed homologous proteins. For example, cytochrome c, a mitochondrial protein involved in Biological Oxidation, is present in many animal species. Although the Cytochromes c of chickens and ducks differ by only two amino acid residues in their primary structure and perform the same function, they are classified as homologous proteins because they belong to different species.

The structural protein collagen also has A large number of isoforms classified as isoproteins. Many enzymes exist in multiple isoforms and are known as isozymes (or Isoenzymes).