The structure of collagen fibrils gradually changes. The number of hydrogen and ether bonds between tropocollagen molecules increases, fibrils thicken (their diameter increases from 500 to 600 A), due to this, the relative proportion of the main substance decreases. The mechanism of damage to the valvular apparatus of the heart during the development of rheumatism is similar to this.
If in a young child's body the hexosamine / hydroxyproline ratio is shifted in favor of hexosamine, then with age this ratio gradually changes in the opposite direction.
The binding base substance decreases, and instead of this, collagen fibers thicken, the ratio between reticulin and collagen fibers changes.
All this leads to significant changes in rheological properties. connective tissue.
A relatively larger percentage of fibroblasts in ST, i.e., biologically active elements in the OB of a child and growing organisms, determines a more intense metabolism, and at the same time, a lighter vulnerability under the influence of various harmful agents.
Rheumatism in childhood, Stefan Kolarov
The location of proline in the primary structure of collagen is determined by DNA and, accordingly, is transmitted from messenger RNA. So far, the process of formation of hydroxyproline has not been fully elucidated. Intracellularly synthesized polypeptides mature outside the cell and form collagen fibrils and fibers. However, the transverse striation of collagen is visible already at the stage of their formation inside the cells. In the process of its isolation, collagen goes through the following stages: ...
Plasminogen activators activate plasminogen, turning it into plasmin. It should be emphasized that the proteolytic effect of plasmin does not extend only to fibrinogen and fibrin, but also to a number of other proteins. Accepted in the past, the specificity is due to the faster breakdown of fibrinogen. It is believed that plasminogen is activated by this peptidase, which cleaves several peptide bonds. The presence of tissue fibrinokinases has been proven in a number of organs and tissues - ...
Structural elements of the collagen substance The insoluble collagen fibril created in this way is the main constituent unit of collagen. The primary structure of collagen (the sequence of amino acids in a protein molecule) is determined primarily by the frequently repeated tripeptide glycyl-prolyl-hydroxyproline, which is of decisive importance for both secondary and tertiary structures. The secondary structure of collagen is determined by the spatial arrangement of the constituent 1000 amino acids, which give the polypeptide chain a helical character. Two…
Bridge bonds that increase the stability of collagen fibrils Designations: a — intrachain and intramolecular, providing a longitudinal bond; b - interchain and intramolecular, providing a cross-link; c - interchain, intermolecular, providing a cross-link. A characteristic feature is the location of glycine residues in the side chains inside the collagen molecule, as well as proline, hydroxyproline and other amino acid residues on its outer side ....
Structural diagram of a collagen fiber (according to Reed) In essence, the sequence of distribution of molecules during the formation of fibrils is ultimately determined by the sequence of alkaline (basic) and acidic amino acids in the collagen molecule, i.e., in its primary structure. The association of individual fibrils into larger bundles before the isolation of collagen fibers is probably carried out with the participation of acid mucopolysaccharides (glucose noglycans). Collagen formation takes place...
Collagen is the main structural protein of the extracellular matrix. This is a fibrillar protein that differs from other proteins in a number of features of its composition and structure.
A typical collagen molecule consists of three polypeptide targets of different types (α-helices), twisted in the form of a right triple helix. In turn, polypeptide chains are built from frequently repeated fragments that have a characteristic sequence -Gly-X-Y-. Every third amino acid residue is glycine. Proline (Pro) is often found in the X positions, the Υ position can be occupied by both proline and 4-hydroxyproline ( 4Nur). In addition, the collagen molecule contains residues of 3-hydroxyproline ( ZNur) and 5-hydroxylysine ( 5Nul).
The presence of hydroxyamino acid residues in the polypeptide chain is characteristic feature collagen. Proline and lysine residues are hydroxylated post-translationally, i.e., after incorporation into the polypeptide chain. At one end, the collagen molecule is cross-linked by cross-links formed by side chains of lysine residues. The number of cross-links increases as the body ages.
When forming secondary structure the polypeptide chain of collagen fits into a more extended left-handed a-helix (there are 3 amino acid residues per turn);
The tertiary structure of collagen is a right-handed superhelix of 3 a-chains, during the formation of which the glycine residue is in its center, which contributes to the formation of a linear tropocollagen molecule with its subsequent incorporation into the fiber.
types of collagen. 19 types of collagen are known, which differ from each other in the primary structure of peptide chains, functions and localization in the body. Collagens are part of fibrils (fibril-forming collagens - types I, II, III, V and XI), myofibrils (type VI collagen), are a structural component of basement membranes (type IV collagen), etc.
Fibril-forming collagens(I, II, III, V and XI types). The basis of fibrils is stepped parallel rows of tropocollagen molecules, which are shifted by 1/4 relative to each other.
Fibrillogenesis is preceded by another modification of lysine. The extracellular copper-containing enzyme lysyl oxidase carries out oxidative deamination of lysine and hydroxylysine with the formation of reactive aldehydes. These reactions require the presence of vitamins PP and B 6 . These groups take part in the formation of transverse covalent bonds between tropocollagen molecules.
The predominant distribution of these types of collagen in tissues is as follows: Type I - bones, dentin, cornea, tendons; Type II - cartilage, intervertebral discs, vitreous body; Type III - kidneys, liver, blood vessels, lymph nodes. Collagen types V and XI are present in varying amounts in the intercellular substance of all tissues; they determine the diameter of collagen fibrils.
Synthesis and maturation of collagen (Fig.3,4)
The synthesis and maturation of collagen is a complex multi-stage process that begins in the cell and ends in the extracellular space. It includes a number of post-translational changes: hydroxylation of proline and lysine, glycosylation of hydroxylysine, cleavage of N- and C-terminal peptides. Due to these changes, additional opportunities for chain stabilization in the tropocollagen molecule appear; not only the NH and CO groups of the peptide backbone but also the OH groups of hydroxyproline are involved in the formation of hydrogen bonds; hydroxyproline and proline, being "hard" molecules, limit the rotation of the polypeptide rod.
A certain role in the synthesis of collagen is played by proteins - chaperones, which provide "quality control" of collagen: they contribute to the correct synthesis of collagen molecules and their transport along secretory pathways, and also "track" incorrectly assembled collagen molecules, which are then destroyed.
SYNTHESIS OF COLLAGEN
There are 8 stages of collagen biosynthesis: 5 intracellular and 3 extracellular.
Occurs on ribosomes, the precursor molecule is synthesized: preprocollagen.
With the help of the signal peptide “pre” transport of the molecule into the tubules of the endoplasmic reticulum. Here “pre” is split off - “procollagen” is formed.
Amino acid residues of lysine and proline in the composition of the collagen molecule undergo oxidation under the action of enzymes. prolyl hydroxylase and lysyl hydroxylase..
With a lack of vitamin "C" - ascorbic acid, scurvy is observed - a disease caused by the synthesis of defective collagen with reduced mechanical strength, which causes, in particular, loosening of the vascular wall and other adverse effects.
Post-translational modification - glycosylation of procollagen by the enzyme glycosyltransferase. This enzyme transfers glucose or galactose to the hydroxyl groups of oxylysin.
The final intracellular stage is the formation of a triple helix - tropocollagen (soluble collagen). The pro-sequence contains the amino acid cysteine, which forms disulfide bonds between chains. There is a process of spiralization.
Tropocollagen is secreted into the extracellular environment, where amino- and carboxyproteinases cleave off the (pro-)-sequence.
Covalent “crosslinking” of the tropocollagen molecule according to the “end-to-end” principle with the formation of insoluble collagen. This process involves the enzyme lysyl oxidase (flavometalloprotein, contains FAD and Cu). Oxidation and deamination of the lysine radical occurs with the formation of an aldehyde group. An aldehyde bond then forms between the two lysine radicals.
Lysyl oxidase is a Cu-dependent enzyme, therefore, with a lack of copper in the body, a decrease in the strength of the connective tissue occurs due to a significant increase in the amount of soluble collagen (tropocollagen).
Association of insoluble collagen molecules according to the “side-to-side” principle. The association of fibrils occurs in such a way that each subsequent chain is shifted by 1/4 of its length relative to the previous chain.
Only after repeated cross-linking of fibrils, collagen acquires its unique strength, becomes an inextensible fiber.
Constituting 25% to 35% of proteins in the entire body.
Encyclopedic YouTube
1 / 3
✪ Golden ointment from a pharmacy for 14 rubles treats 100 diseases. A penny forgotten remedy. Is it legal at all?
✪ AUTOPHAGY and FASTING. Nobel Prize for ETERNAL LIFE!? Cure CANCER in a DAY
✪ Actually connective tissues 1. RVST
Subtitles
Research History
Scientists for decades could not understand the molecular structure of collagen. The first proof that collagen has a permanent structure at the molecular level was presented in the mid-1930s. Since that time, many eminent scientists, including Nobel laureates such as Francis Crick, Linus Pauling, Alexander Rich, Ada Yonath, Helen Berman, Vileainur Ramachandran, have been working on the structure of the collagen monomer.
Several contradictory models (despite the known structure of each individual peptide chain) gave way to the creation of a ternary-helical model that explained the quaternary structure of the collagen molecule.
Properties
Collagen exists in several forms. The basis of the structure of all types of collagen is similar. Collagen fibers are formed by aggregation of microfibrils, they are pink when stained with hematoxylin and eosin and blue or green at various three-chromic stains, when impregnated with silver, they turn brown-yellow.
fibrillar structure
Tropocollagens (structural units of collagen) spontaneously combine, attaching to each other with ends displaced by a certain distance, forming larger structures in the intercellular substance. In fibrillar collagens, the molecules are displaced relative to each other by about 67nm (the unit, which is denoted by the letter "D" and varies depending on the state of hydration of the substance). In general, each D-period contains four whole and part of the fifth collagen molecule. The value of 300 nm divided by 67 nm (300:67) does not give an integer and the length of the collagen molecule is divided into segments D that are not constant in size. Therefore, in the context of each repeat of the D-period of the microfibril, there is a part consisting of five molecules, called ”, and a part consisting of four molecules - “gap”. The tropocollagens are also arranged in a hexagonal or pseudo-hexagonal (in cross section) design, in each "overlap" and "gap" region.
Within tropocollagen, there is a covalent bond between the strands, as well as a variable amount of these bonds between the tropocollagen helices themselves, forming well-organized structures (eg, fibrils). Thicker fibril bundles are formed by several other classes of proteins, including other types of collagens, glycoproteins, proteoglycans, used to form different types of tissues from different combinations of the same basic proteins. The insolubility of collagen was an obstacle to the study of the collagen monomer until it was found that it was possible to extract young animal tropocollagen because it had not yet formed strong bonds with other fibril subunits. However, improvements in microscopes and X-ray machines made research easier, and more and more detailed images of the structure of the collagen molecule appeared. These late discoveries are very important for a better understanding of how the structure of collagen affects the connections between cells and intercellular substance, how tissues change during growth and regeneration, how they change during embryonic development and pathology.
A collagen fibril is a semi-crystalline structural unit of collagen. Collagen fibers are bundles of fibrils.
Usage
food industry
From a nutritional point of view, collagen and gelatin are low quality proteins, since they do not contain all the essential amino acids needed by a person - these are incomplete proteins. Manufacturers of collagen-based supplements claim their products can improve skin and nail quality, as well as joint health [ ] .
Relatively cheap, often offered on the market today under the guise of a source of free amino acids, collagen hydrolysates are not always able to satisfy human needs for free amino acids, since these products do not contain amino acids ready for assimilation, but are only partially “digested” extracts of articular tissues of mammals, birds or sea dwellers. For example, collagen hydrolysates are almost completely devoid of the amino acid L-glutamine, which is not resistant to thermal effects and long-term storage of raw materials, most of glutamine is destroyed already at the first stages of storage and processing of raw materials, the existing small residue is almost completely decomposed during the thermal extraction of cartilage tissue.
The best quality sources of amino acids are preparations containing the so-called "free amino acids". Since it is free amino acids that are almost ready for absorption, the body does not need to spend digestive enzymes, time and energy on their digestion. They are able to enter the blood in the shortest possible time and, being delivered by it to places in need of additional collagen synthesis, are immediately included in its formation [ ] .
Cosmetical tools
- The formation of a breathable, moisture-retaining layer on the surface of the skin, which has plasticizing (smoothing) properties, with the properties of a wet compress;
- Prolongation of the action of extracts, oils, etc. in the composition of cosmetic compositions;
- Giving shine to hair, creating a collagen (protective) layer on the surface of the hair.
Scientific research
In 2005, scientists were able to isolate collagen from the preserved soft tissues of a Tyrannosaurus rex and use it chemical composition as another proof of the relationship of dinosaurs with modern birds.
Scientific research in medicine
Collagen synthesis is a complex enzymatic multi-stage process that must be provided with a sufficient amount of vitamins and mineral elements. Synthesis takes place in the fibroblast and a number of stages outside the fibroblast. An important point in the synthesis is the hydroxylation reactions, which open the way for further modifications necessary for the maturation of collagen. Specific enzymes catalyze hydroxylation reactions. Thus, the formation of 4-hydroxyproline is catalyzed by proline hydroxylase, in the active center of which there is iron. The enzyme is active if the iron is in the divalent form, which is provided by ascorbic acid (vitamin C). Ascorbic acid deficiency disrupts the hydroxylation process, which affects the further stages of collagen synthesis: glycosylation, cleavage of N- and C-terminal peptides, etc. As a result, abnormal collagen is synthesized, more loose. These changes underlie the development of scurvy. Collagen and elastin form a kind of "basis" of the skin, which prevents it from sagging, ensures its elasticity and firmness. Elastin as a protein stops the production of enzymes in human body at the age of 14, and collagen - at 21-25, after which the skin is not restored and the skin ages. Also, the most important component of connective tissue is keratin - a family of fibrillar proteins with mechanical strength, which among materials of biological origin is second only to chitin. Keratins mainly consist of horny derivatives of the epidermis of the skin - structures such as hair, nails, horns, feathers, etc.
The photo
A characteristic manifestation of these diseases is damage to the ligamentous apparatus, cartilage, bone system, the presence of heart valve defects.
Diseases caused by defects in collagen biosynthesis, including so-called collagenoses, arise from a variety of causes. This may be due to a mutation in the gene encoding the amino acid sequence of collagen-producing enzymes, resulting in a change in the shape of the collagen molecule, or an error in the post-translational modification of collagen. Also, diseases can be caused by a lack or "malfunction" of enzymes involved in collagen biosynthesis - a deficiency of hydroxylation enzymes (proline-, lysine hydroxylase), glycosyltransferases, N-procollagen and C-procollagen peptidases, lysyl oxidase, followed by a violation of cross-links, deficiency of copper, vitamins B6, B13 (orotic acid), . In acquired diseases such as scurvy, restoring the enzyme balance to normal can lead to a complete cure.
Almost any gene mutation leads to the loss or change in the functions of collagen, which, in turn, affects the properties of tissues and organs. Gene mutations in the collagen domain can lead to a change in the shape of the triple helix by insertion/deletion of an amino acid from the polypeptide chain or replacement of Gly with another base. Mutations in non-collagenous domains can lead to misassembly of α-chains into supramolecular structures (fibrils or networks), which also leads to loss of function. Mutant α-chains are able to form a three-stranded complex with normal α-chains. In most cases, such complexes are unstable and rapidly decompose, however, such a molecule can normally fulfill its role if not functionally affected. important areas. Most diseases caused by mutations in "collagen" genes are
The structure of simple proteins is represented only a polypeptide chain(albumin, insulin). However, it must be understood that many simple proteins (for example, albumin) do not exist in a "pure" form, they are always associated with some non-protein substances. They are referred to simple proteins only for the reason that the bonds with the non-protein group weak and when highlighting in vitro they are free from other molecules - a simple protein.
Albumins
In nature, albumins are part of not only blood plasma (serum albumin), but also egg protein (ovalbumin), milk (lactalbumin), and are storage proteins in the seeds of higher plants.
Globulins
A group of diverse blood plasma proteins with molecular weight up to 100 kDa, subacid or neutral. They are poorly hydrated, less stable in solution and easier to precipitate than albumins, which is used in clinical diagnostics in "sedimentary" samples (thymol, Veltman). Although they are usually classified as simple, many globulins contain carbohydrate or other non-protein components.
At electrophoresis Serum globulins are divided into at least 4 fractions - α 1-globulins, α 2-globulins, β-globulins and γ-globulins.
Electrophoregram pattern (top) of blood serum proteins
and the resulting proteinogram (below)
Since globulins include a variety of proteins, their functions are varied:
Part of α-globulins has antiprotease activity, which protects blood proteins and extracellular matrix from premature destruction, for example, α 1 -antitrypsin, α 1 -antichymotrypsin, α 2 -macroglobulin.
Some globulins are capable of binding certain substances: transferrin (carries iron ions), ceruloplasmin (contains copper ions), haptoglobin (hemoglobin carrier), hemopexin (heme transport).
γ-Globulins are antibodies and provide immune defense of the body.
Histones
Histones are intranuclear proteins weighing about 24 kDa. They have pronounced basic properties, therefore, at physiological pH values, they are positively charged and bind to deoxyribo-nucleic acid (DNA), forming deoxyribo-nucleoproteins. There are 5 types of histones - very rich in lysine (29%) histone H1, other histones H2a, H2b, H3, H4 are rich in lysine and arginine (up to 25% in total).
Amino acid radicals in histones can be methylated, acetylated, or phosphorylated. This changes the net charge and other properties of proteins.
There are two functions of histones:
1. regulation of genome activity, namely, they interfere with transcription.
2. Structural - stabilize spatial structure DNA.
Histones in complex with DNA form nucleosomes - octahedral structures composed of histones H2a, H2b, H3, H4. Histone H1 is bound to the DNA molecule, preventing it from slipping off the histone octamer. DNA wraps around the nucleosome 2.5 times, after which it wraps around the next nucleosome. Thanks to this stacking, a 7-fold reduction in DNA size is achieved.
Thanks to histones and the formation of more complex structures, the size of DNA eventually decreases thousands of times: in fact DNA length reaches 6-9 cm (10 -1), and the size of chromosomes is only a few micrometers (10–6).
Protamines
These are proteins weighing from 4 kDa to 12 kDa, they are found in the nuclei of the spermatozoa of many organisms, in the sperm of fish (milk) they make up the bulk of the protein. Protamines are histone substitutes and serve to organize chromatin in sperm. Compared with histones, protamines have a sharply increased content of arginine (up to 80%). Also, unlike histones, protamines have only a structural function, they do not have a regulatory function, chromatin in spermatozoa is inactive.
Collagen
Collagen is a fibrillar protein with a unique structure that forms the basis of the intercellular substance of the connective tissue of tendons, bones, cartilage, skin, but, of course, it is also found in other tissues.
The polypeptide chain of collagen consists of 1000 amino acids and is called the α-chain. There are about 30 variants of the α-chain of collagen, but they all have one common feature - to a greater or lesser extent include a repeating triplet [ Gly-X-Y], where X and Y are any amino acids except glycine. Pregnant X more often located proline or, much less often, 3-hydroxyproline, pregnant Y meets proline And 4-hydroxyproline. Also in position Y often located alanine, lysine And 5-oxylysin. Other amino acids account for about a third of the total number of amino acids.
The rigid cyclic structure of proline and hydroxyproline does not allow the formation of a right-handed α-helix, but forms a so-called. "proline fracture". Due to this break, a left-handed helix is formed, where there are 3 amino acid residues per turn.
Hydroxylation plays an important role in collagen synthesis. lysine And proline included in the primary chain, carried out with the participation of ascorbic acid. Also, collagen usually contains monosaccharide (galactose) and disaccharide (glucose-galactose) molecules associated with the OH groups of some oxylysin residues.
Stages of collagen molecule synthesis
synthesized molecule collagen built of 3 polypeptide chains woven together into a tight bundle - tropocollagen(length 300 nm, diameter 1.6 nm). Polypeptide chains are firmly linked to each other through the ε-amino groups of lysine residues. Tropocollagen forms large collagen fibrils with a diameter of 10-300 nm. The transverse striation of the fibril is due to the displacement of tropocollagen molecules relative to each other by 1/4 of their length.
Collagen fibrils are very strong, they are stronger than steel wire of equal cross section. In the skin, fibrils form an irregularly woven and very dense network. For example, dressed leather is almost pure collagen.
Hydroxylation of proline is carried out iron-containing enzyme prolyl hydroxylase which requires vitamin C (ascorbic acid). Ascorbic acid protects prolyl hydroxylase from inactivation, maintaining the reduced state iron atom in the enzyme. Collagen synthesized in the absence of ascorbic acid is insufficiently hydroxylated and cannot form fibers that are normal in structure, which leads to skin damage and vascular fragility, and manifests itself as scurvy.
Hydroxylation of lysine is carried out by the enzyme lysylhydroxylase. It is sensitive to the influence of homogentisic acid (tyrosine metabolite), with the accumulation of which (diseases alkaptonuria) collagen synthesis is disrupted, and arthrosis develops.
The half-life of collagen is calculated in weeks and months. Plays a key role in its exchange collagenase cleaving tropocollagen 1/4 of the distance from the C-terminus between glycine and leucine.
As the body ages, an increasing number of cross-links are formed in tropocollagen, which makes the collagen fibrils in the connective tissue more rigid and brittle. This leads to increased bone fragility and a decrease in the transparency of the cornea of the eye in old age.
As a result of the breakdown of collagen, hydroxyproline. With damage to the connective tissue (Paget's disease, hyperparathyroidism), the excretion of hydroxyproline increases and has diagnostic value.
Elastin
|
|
According to the structure in in general terms elastin is similar to collagen. It is located in the ligaments, the elastic layer of blood vessels. The structural unit is tropoelastin with a molecular weight of 72 kDa and a length of 800 amino acid residues. It has much more lysine, valine, alanine and less hydroxyproline. The absence of proline causes the presence of helical elastic regions.
A characteristic feature of elastin is the presence of a peculiar structure - desmosine, which, with its 4 groups, combines protein chains into systems that can stretch in all directions.
α-Amino groups and α-carboxyl groups of desmosine are included in peptide bonds one or more protein chains.
Intercellular matrix - a complex of organic and inorganic components that fill the space between cells. Different tissues have their own intercellular matrix. Epithelial cells are predominantly bound by glycoproteins, calcium-binding proteins. The special structure of the intercellular matrix is inherent in tissues of mesenchymal origin, which perform mechanical, protective and trophic functions. They are divided into:
Connective tissue proper - loose, unformed,
dense decorated and unformed; tissues with special properties - adipose, pigmented,
reticular and mucous; skeletal tissues - bone and cartilage.
All these types of connective tissue are widely represented throughout the body, and in particular in the head and neck.
1.1. ORGANIZATION OF THE INTERCELLULAR MATRIX
Connective tissue is characterized by the presence of a large amount of intercellular substance (extracellular matrix), consisting of collagen proteins, proteoglycans and glycoproteins, and a small number of cells located at a considerable distance from each other. Fibroblasts, chondroblasts, osteoblasts, odontoblasts, cementoblasts and other blast cells participate in the formation of the intercellular substance. A feature of mineralized tissues is the presence in the intercellular substance of inorganic ions that form salts and crystals.
The extracellular matrix contains molecules capable of self-assembly to form complexes. Due to a certain location of binding centers on molecules and the specificity of their interactions, a highly ordered three-dimensional structure of the extracellular matrix is formed, which determines its functional properties (Fig. 1.1).
Rice. 1.1.Structural organization of the intercellular matrix and its relationship with
cells:
BUT -basement membrane; B - supramolecular organization of the matrix in
connective tissue [according to Campbell N. A., Reece J. B., 2002, with changes].
A specialized form of the extracellular matrix of normal tissue is the basement membrane, which forms a discrete structure that separates one cell layer from another. It is responsible not only for the differentiation of various structures and maintenance of tissue architectonics, but also affects their differentiation, migration, and cell phenotyping. The basement membrane serves as a barrier for macromolecules.
The main components of the extracellular matrix are various types of collagen and non-collagen proteins.
1.2. STRUCTURE AND PROPERTIES OF COLLAGEN PROTEINS
The basis of the extracellular matrix is the family of collagen proteins, related to glycoproteins and containing a large number of residues glycine, proline And hydroxyproline. Collagens are represented by 20 proteins, of which some are actually
collagens, while others contain only collagen-like domains. All types of collagens, depending on the structure, are divided into several groups: fibril-forming, associated with collagen fibrils, reticular, microfibrils, anchored fibrils, etc. To designate each type of collagen, a certain formula is used, in which a-chains are written in Arabic numerals, and the type collagen - Roman.
The bulk of the collagens present in the tissues of the oral cavity are fibril-forming. Localization of the main types of collagen proteins in the tissues of the oral cavity is presented in Table. 1.1.
Table 1.1
Types of collagen proteins in oral tissues
The tissues of the oral cavity are characterized by the presence of collagen types I, III, V and VI. It should be noted the diversity of collagen in the cement of the tooth, in which, in addition to collagen types I, III and V, collagen types II, IX, XII, XIV, characteristic of cartilage tissue, are determined.
Fibril-forming collagens
All fibril-forming collagens differ in amino acid composition and carbohydrate content.
Molecules of collagen types I, II, III, V, XI have the form of fibrils and are built from structural units called tropocollagens. Tropocollagen molecules (M r 300 kDa) are 1.5 nm thick and 300 nm long. They are formed by three polypeptide chains, referred to as a-chains. Each chain contains about 1000 amino acid residues and is a tight left-handed helix containing three amino acid residues per turn. One third of the amino acid residues in collagen are glycine.
(30%), one fifth of proline in total with 3- and 4-hydroxyproline (21%), so the primary structure of collagen can be represented as a scheme of gly - x - y -, where x is most often proline or hydroxyproline, and y - other amino acids (Fig. 1.2). In total, about 330 such repeats are found in the a-chain.
Rice. 1.2.Fragment primary structure but - collagen chains. In the area where proline and hydroxyproline are located, a “proline break” occurs.
The glycine of the repeating sequence gly - x - y - is necessary for the formation of the fibrillar structure, since the radical of any other amino acid does not fit between the three peptide chains in the center of the triple helix. Proline and hydroxyproline limit the rotation of the polypeptide chain. The amino acid radicals located in the -x- and -y- positions are located on the surface of the triple helix. The distribution of radical clusters along the length of the collagen molecule provides self-assembly of multimolecular collagen structures. Three a-chains form a structure, slightly twisted into a helix. Forming fibrils, tropocollagen molecules (trimers) are arranged in steps, shifting relative to each other by one quarter of the length, which gives the fibrils a characteristic striation. Being deposited in tissues, the formed collagen fibrils are stabilized through the formation of covalent cross-links (Fig. 1.3).
type I collagen 2 but 2 contains 33% glycine, 13% proline, 1% hydroxylysine and a low amount of carbohydrates. It is determined in the composition of bones, dentin, dental pulp, cement, periodontal fibers. This type of collagen fibers is involved in mineralization processes.
type II collagen[α 1 (II)] 3 is present in cartilage and is formed in non-cartilaginous tissues early in development. This type contains no collagen a large number of 5-hydroxylysine (less than 1%) and has a high carbohydrate content (more than 10%).
type III collagen[α 1 (III)] 3 is present in the walls blood vessels. A distinctive feature of this collagen is the presence of a large amount of hydroxyproline. The α-chains contain cysteine, and the collagen molecule itself is weakly glycosylated.
Type V collagen [α(V)α 2 (V)α 3 (V)] is a hybrid molecule consisting of different chains, namely: α 1 (V), α 2 (V) and α 3 (V).
Fibrillar collagens can also contain 2 or more different types of collagens. So, in some tissues there are hybrid molecules containing chains of collagen types V and XI.
Rice. 1.3.Structure of collagen fibrils: BUT - tropocollagen, consisting of three α - chains ; B - collagen microfibrils from 5 rows of tropocollagen; IN - collagen fibrils containing 9-12 tropocollagen microfibrils.
Fibril-associated collagens
Collagen types IX, XII, XIV are involved in the organization of the intercellular matrix of the mucous membrane, cartilage and cementum of the tooth root. Collagen proteins of this class are not capable of forming fibrils, but, by binding to fibrillar collagens, they limit the length, thickness, and orientation of collagen types I and II fibrils. A specific feature of collagens associated with fibrils is the presence of both globular and fibrillar domains in their structure.
α-chains of type IX collagen [α(IX)α 2 ( IX )α 3 (IX)] consist of 3 fibrillar and 4 globular domains. They are connected across covalent bonds with type II collagen fibrils. The type IX collagen molecule also contains a glycosaminoglycan side chain and a large number of positively charged groups, so negatively charged molecules of hyaluronic acid and chondroitin sulfate can join it. Type XII collagens enter into similar interactions with type I fibrillar collagens. This type of collagen is localized in cartilage, cementum, and also in the oral mucosa at the junctions of the epithelium with the subepithelial layers. Type IX collagen is a transmembrane protein that lamina densa(dark plate of the basement membrane, located on the border with the papillary dermis) is fixed to the collagen fibrils of the papillary dermis.
Non-fibrillar (network) types of collagen
The group of non-fibrillar collagens includes collagen proteins IV, VIII and X types, which differ in length and size and are capable of forming reticular structures. The most common, including in the tissues of the oral cavity, type IV collagen, which is the main structural protein of basement membranes. Type IV collagen contains 1 α 1 (IV) and 2 α 2 (IV) chains. The peptide chains of type IV collagen do not undergo proteolytic modification after secretion and therefore retain the structure of the N- and C-terminal globular domains (NC 1 , 7S and NC 2) (Fig. 1.4).
Rice. 1.4.The structure of type IV collagen is a triple helix of the collagen monomer. The N- and C-terminal regions contain globular domains 7S, NC 1 and NC 2 .
Unlike fibrillar collagens, α-chains of type IV collagen molecules contain “non-collagenous” amino acid regions not only in the N- and C-terminal sections, but throughout the entire molecule. The terminal domains NC 1 , 7S of collagen monomers interact with each other in the process of self-aggregation and form end-to-end bonds, which leads to the formation of dimers and trimers. Supercoiling is provided by lateral interactions and end-to-end connections. As a result, three-dimensional structures are formed, similar to a grid with hexagonal cells 170 nm in size.
Type X collagen consists of 3 identical chains with a pier. weighing 59 kDa.
Collagen forming microfibrils
Type VI collagen is referred to as microfibril-forming collagen. Being a short chain protein, it forms microfibrils located between fibrils of interstitial collagens. This type of collagen is characterized by the presence of large globular domains in the α-chains in the N- and C-terminal regions and a short three-helix domain between them. In the process of synthesis inside the cell, 2 molecules of this collagen are combined antiparallel to form a dimer, and tetramers are formed from the dimers, which are secreted from the cell. Outside the cell, tetramers bind end-to-end to form microfibrils. The molecules of this collagen contain numerous sequences arg-gli-asp(RGD), which provide cell adhesion by attaching to membrane adhesive proteins - integrins α 1 β 1 and α 2 β 1 . In addition, type VI collagen is able to bind to interstitial collagen fibrils, proteoglycans, and glycosaminoglycans.
Synthesis of collagen
Collagen is synthesized and supplied to the extracellular matrix by almost all cells (fibroblasts, chondroblasts, osteoblasts, odontoblasts, cementoblasts, keratoblasts, etc.). The synthesis and maturation of collagen is a complex multi-stage process that begins in the cell and ends in the extracellular matrix. Disturbances in collagen synthesis caused by mutations in genes, as well as in the process of translation and post-translational modification, are accompanied by the appearance of defective collagens. Since about 50% of all collagen proteins are found in the tissues of the skeleton, and the remaining 40% in the dermis and 10% in the stroma internal organs, then violations of collagen synthesis are accompanied by pathology as
musculoskeletal system and internal organs. This inevitably affects the state of the tissues of the maxillofacial region.
Synthesis of collagen includes two stages. On the intracellular stage translation and post-translational modification of polypeptide chains occurs, and on extracellular - protein modification, culminating in the formation of collagen fibers (Fig. 1.5).
Rice. 1.5.Synthesis of collagen. Scheme of collagen synthesis: BUT - intracellular stage, B - extracellular protein modification. The numbers indicate the synthesis reactions. 1a - transcription, 1b- translation of procollagen chains, 2 - cleavage of the signal peptide, 3 - hydroxylation of proline and lysine residues, 4 - glycosylation of 5-hydroxylysine and asparagine, 5 - formation of disulfide bonds in N- and C-terminal peptides, 6 - formation of the procollagen triple helix, 7 - exocytosis of a protein molecule, 8 - cleavage of N- and -terminal peptides, 9 - adjustable fibril assembly, 10 - oxidation of lysine and 5-hydroxylysine to allisins, 11 - the formation of cross-links with the formation of polymeric peptides [according to Kolman Ya., Rem K.-G., 2000, with changes]. Enzymes:
1 - procollagenproline-4-dioxygenase;
2 - procollagenlysin-5-dioxygenase;
3 - protein-lysine-6-oxidase.
Intracellular stage of collagen synthesis . Peptide α-chains of collagen are synthesized on polyribosomes associated with the membranes of the endoplasmic reticulum. Its synthesized peptide chains in the tanks undergo post-translational modification, which includes:
Removal of the signal peptide of the procollagen chain with the participation of a specific proteinase;
Hydroxylation of proline and lysine residues, which begins during the translation of the polypeptide chain up to its separation from the ribosomes.
Hydroxylation reactions are catalyzed by oxygenases: procollagenprolyl-4-dioxygenase (prolyl-4-hydroxylase), procollagenprolyl-3-dioxygenase (prolyl-3-hydroxylase), and procollagenlysyl-5-dioxygenase (lysyl-5-hydroxylase). O 2 and 2-oxoglutarate are used in the hydroxylation reaction, and ascorbic acid is involved as a cofactor. Proline and lysine hydroxylases contain Fe 2+ in the active center, and ascorbic acid, which is easily oxidized to dehydroascorbic acid, is necessary to preserve the iron atom in the ferroform (Fig. 1.6).
Rice. 1.6.Structural formula of ascorbic acid.
In hydroxylation reactions, one oxygen atom is attached to the fourth carbon atom in the proline residue, and the second oxygen atom is included in succinic acid, which is formed during the decarboxylation of 2-oxoglutarate (Fig. 1.7).
Along with the hydroxylation of proline, hydroxylation of lysine residues occurs with the formation of 5-hydroxylysine (Fig. 1.8).
Subsequently, hydroxylated lysine residues undergo glycosylation.
With the participation of glycosyltransferases, covalent O-glycosidic bonds are formed between the 5-OH group of hydroxylysine and the galactose residue or the disaccharide galactosylglucose. Molecules of N-acetylglucosamine or mannose are attached to the amide group of asparagine. Simultaneously with the hydroxylation of proline, a stable three-helix structure of collagen is formed (Fig. 1.9).
Rice. 1.7.Hydroxylation of proline residues in the procollagen α-chain with the formation of 4-hydroxyproline.
Rice. 1.8.Hydroxylation of lysine residues in the pro-collagen a-chain with the formation of 5-hydroxylysine.
Hydroxyproline is essential for the stabilization of this collagen triple helix, since its hydroxyl groups are involved in the formation of hydrogen bonds between α-chains. Upon completion of hydroxylation and glycosylation, all pro-α-chains are linked by hydrogen bonds, and disulfide bridges are formed in the region of the C-terminal propeptides.
Rice. 1.9.Glycosylated regions of the α-chain of the procollagen molecule.
From the endoplasmic reticulum, procollagen molecules move to the Golgi apparatus, where they are included in the secretory vesicles and, in their composition, are secreted into the extracellular space.
Extracellular stage - modification of procollagen molecules . In the intercellular space, with the participation of proteolytic enzymes, N- and C-terminal peptides are cleaved from the procollagen molecule and the triple helix of collagen (tropocollagen) is released. Next, the process of self-assembly of collagen fibrils, fixed by intermolecular covalent bonds (crosslinks), occurs. The formation of these bonds involves lysine and 5-hydroxylysine residues and their aldehyde derivatives, which are formed as a result of oxidative deamination. Oxidative deamination of lysine and 5-hydroxylysine occurs with the participation of lysyl oxidase. A feature of this enzyme is the presence of Cu 2+ in the active center. Lysyl oxidase molecules are synthesized in the cell as proenzymes and, after binding to Cu 2+ ions, are packaged into vesicles that leave the cell. On the cell surface, the prolysyl oxidase molecule undergoes limited proteolysis, and in the formed active center, with the participation of Cu 2+ ions, the tyrosine residue is oxidized to tyrosine quinone. The quinoid structure formed in the active center binds lysine residues in the procollagen molecule to form an enzyme-substrate complex. Further deamination of lysine occurs in accordance with the reactions presented in Fig. 1.10.
In the next step, allisin and 5-hydroxyallysine are condensed together with lysyl and hydroxylysyl residues; intra- and intermolecular cross-links are formed. In reaction
Rice. 1.10.Lysine oxidation in the collagen structure:
1 - formation of an enzyme-substrate complex; 2 - NH 3+ is transferred to tyrosinequinone (LTQ) and lysine is oxidized with subsequent displacement of allisin from the active center; 3 - O 2 and H 2 O molecules enter the active center of the enzyme and NH 3 and H 2 O 2 are released. In this case, LTQ returns to its original state (Enz - enzyme).
condensation of allisin with a lysine residue of another chain, a Schiff base is formed. In the case of aldol condensation of two allisine residues, aldol intermolecular bonds (lysinnorleucine) are formed. The formation of aldol intermolecular bonds is shown in fig. 1.11.
Aldol condensation is characteristic of collagen bone tissue and dentine, while Schiff bases are most commonly found in tendon collagens.
About 25% of tropocollagen molecules decompose without forming fibrils. The resulting fragments perform signaling functions and stimulate collagenogenesis. The spatial organization of fibrils is completed with the participation of fibronectin, proteoglycans, and collagens associated with fibrils.
Aldol intermolecular bonds
Rice. 1.11.Oxidation of lysine and formation of an aldol intermolecular bond in the reactions of aldol condensation of two allisin residues.
Violation of the synthesis of collagen proteins in humans
Any violations in the synthesis of collagen proteins are clinically manifested, first of all, by a change in the dentoalveolar system in the form of bleeding gums, mobility and loss of teeth, multiple caries. The reasons leading to the violation of the synthesis of collagen proteins are different - a lack of ascorbic acid in the body, Cu 2+ ions, genetic defects and autoimmune conditions.
Hydroxylation of lysine and proline is very milestone for the subsequent formation of covalent bonds between collagen molecules and the assembly of collagen fibrils, depending on the amount of ascorbic acid. In scurvy, a disease resulting from a lack of ascorbic acid, the hydroxylation of proline and lysine residues in the structure of procollagen suffers. As a result, fragile and brittle vessels are formed. Violation of collagen synthesis in the pulp and dentin leads to the development of multiple caries, periodontal ligaments suffer.
Similar phenomena occur in congenital lysylhydroxylase deficiency (Ehlers-Danlo-Rusakov syndrome, type IV). The high solubility of collagen molecules is manifested in a congenital defect of lysyl oxidase (Ehlers-Danlos syndrome, type V) or a violation of copper metabolism (Menkes disease), which is associated with a violation of the formation of cross-links between collagen microfibrils. This leads to a deterioration in the mechanical properties of periodontal ligaments, the condition of periodontal tissues, flaccidity of the skin and the occurrence of defects in the development of the skeleton in people suffering from this disease.
In diabetes mellitus, due to the inability of cells to capture glucose from blood plasma, the process of intracellular glycosylation of procollagen α-chains is disrupted. When procollagen enters the intracellular space, carbohydrates are attached in a non-enzymatic way, which also disrupts the structure of collagen fibrils and non-collagen proteins. A severe form of periodontitis develops, which is difficult to treat. In children born to mothers suffering from insulin-dependent diabetes mellitus, systemic hypoplasia of hard tooth tissues is detected.
Violation of the structure of the basement membrane occurs when antibodies to proteins that form the architectonics of the basement membranes appear (Goodpasture's syndrome), or mutations in the gene encoding the α-chains of type IV collagen (Alport's syndrome). In these forms of pathology, along with damage to the kidneys and other organs, non-carious lesions of the hard tissues of the tooth (enamel hypoplasia, a decrease in the volume and violation of the structure of the dentin) and degenerative changes in the soft tissues of the oral cavity are observed.
To study the metabolism of collagen in urine and blood plasma, the concentration of hydroxyproline, proline, the amount of degradation products of type I collagen - N- and C-telopeptides are determined. A characteristic indicator of collagen breakdown is an increase in the amount of hydroxyproline in the blood plasma and urine, as well as an increase in the amount of N- and C-telopeptides in the blood plasma and the calcium content determined in the urine in the morning before meals. An increase in the amount of proline in the blood plasma indicates a violation of collagen maturation.
In addition to collagen proteins, the extracellular matrix also contains non-collagen proteins - elastin, proteoglycans, glycoproteins, etc.
1.3. STRUCTURE AND PROPERTIES OF NON-COLLAGEN PROTEINS
Elastin
In the intercellular substance of the walls of blood vessels, periodontal tissues, the root of the tongue, in the submucosal layer of the lips and cheeks, lungs, and skin, elastin fibers are present in large quantities. These fabrics have very important properties: they can stretch several times their original length, while maintaining high tensile strength, and return to their original state after the load is removed. The rubber-like properties of these tissues are provided by the main protein elastin - a glycoprotein with a mol. weighing 70 kDa.
Elastin contains about 27% glycine, 19% alanine, 10% valine, 4.7% leucine. The presence of a large number of hydrophobic radicals prevents the creation of a stable globule; as a result, elastin polypeptide chains do not form regular secondary and tertiary structures, but take different conformations in the intercellular matrix with approximately equal free energy (Fig. 1.12).
Rice. 1.12.Fragment of the elastin polypeptide chain.
Native elastin fibers are built from relatively small, almost spherical molecules connected into fibrous strands with the help of rigid cross-links - desmosine and isodesmosine, as well as lysinorleucine. 4 lysine residues are involved in the formation of cross-links, 3 of which are pre-oxidized to the corresponding aldehydes with the participation of lysyl oxidase. Desmosine and isodesmosine are formed by residues belonging to at least two chains, but they can also be formed by residues located in three and four chains. 2 lysine residues are involved in the formation of lysinnorleucine (Fig. 1.13).
Rice. 1.13.Cross-links in the structure of elastin: BUT- desmosine, formed by four lysine residues; B- lysinnorleucine, formed by two lysine residues.
The formation of covalent links between peptide chains of elastin with a random conformation allows the network of elastin fibers to stretch and contract in all directions, which gives them the property of elasticity (Fig. 1.14).
Synthesis and breakdown of elastin . Synthesis of elastin begins in fibroblasts with the formation of a precursor of elastin, the tropoelastin protein. Tropoelastin is a soluble monomer whose hydrophilic regions are enriched in lysine residues. In the extracellular matrix, with the participation of copper-dependent lysyl oxidase, lysine residues are oxidized to allisin, which form cross-links that stabilize the elastin molecule. After cross-linking, elastin assumes its final extracellular form, which is characterized by insolubility, high stability, and low metabolic rate.
In the breakdown of elastin, elastase of polymorphonuclear leukocytes is involved, which, being an endopeptidase, mainly dis-
Rice. 1.14.Structural model of elastin:
BUT- a state of relaxation; B- the state of stretching.
cleaves bonds formed by carboxyl groups of aliphatic amino acids. It is active in a slightly alkaline environment (pH 7.5-8.5) and hydrolyzes in the extracellular space not only elastin, but also other proteins - proteoglycans, hemoglobin, collagen, immunoglobulins. Elastase activity inhibits protein α 1 -antitrypsin (α 1 -AT). The largest amount of α 1 -AT is synthesized by the liver and is found in the blood. In tissues, α 1 -AT is synthesized by macrophages.
Changes in the structure of elastin during pathological processes
In violation of the formation of desmosines, isodesmosines and lysine-norleucine, the tensile strength of elastic tissues decreases, such violations as thinness, lethargy, extensibility appear, that is, their plastic properties are lost. Changes in the structure of elastin may be due to a decrease in the activity of lysyl oxidase in hereditary and acquired diseases, copper deficiency. Violations of the structure of elastin can be manifested by cardiovascular changes in the form of aneurysms and ruptures of the aorta, heart valve defects, frequent pneumonia and emphysema.
Elastase is not active in gum tissues. With the development of inflammation, the number of polymorphonuclear leukocytes increases and they become a source of elastase. The increase in the amount of the latter occurs against the background of unchanged or reduced content of α 1 -AT in the gum tissue. The resulting imbalance between the enzyme and its inhibitor leads to the destruction of elastic fibers in gingivitis and periodontitis.
Proteoglycans and glycosaminoglycans
Proteoglycans- a class of complex proteins of the extracellular matrix. They consist of various core (core) proteins, to which oligosaccharides associated with glycosaminoglycan chains are attached through N- and O-glycosidic bonds (Fig. 1.15).
Rice. 1.15.The structure of the proteoglycan.
Different proteoglycans differ in molecular size, relative protein content and set of glycosaminoglycans. Proteoglycans are present in large quantities in dentin, pulp, cement, periodontal tissues, mucous membranes of the oral cavity (Table 1.2).
Some proteoglycans - serglycin, cartilage matrix proteoglycan, decorin, versican, etc. are in a soluble state and are localized in the extracellular matrix. Other proteoglycans, such as syndecan, are represented by transmembrane integrals. Table 1.2
Proteoglycans and glycosaminoglycans in oral tissues
ny proteins. Syndecan has extracellular transmembrane and cytoplasmic domains and interacts with the actin cytoskeleton. Outside, on the cell surface, syndecan binds to fibronectin and other components of the extracellular matrix.
Molecules of glycosaminoglycans are involved in the binding of proteoglycans to specific proteins. Their negatively charged groups interact with the positively charged radicals of the amino acids lysine and arginine located in certain areas of the protein molecule. In this way, highly sulfated glycosaminoglycans bind to fibronectin.
Proteoglycans act as receptors in the assembly of the extracellular matrix, facilitate cell attachment, and regulate cell growth processes. They can also form complexes with certain proteins, such as growth factors. In the resulting complexes, proteins are protected from proteolytic enzymes. These complexes act as reservoirs, and only if necessary, the growth factor is released from them and acquires the ability to display its biological activity.
Glycosaminoglycans belong to heteropolysaccharides. These are linear structures built from repeating disaccharide units. The disaccharide molecule consists of an uronic acid and an amino sugar, the amino group of which is usually acetylated. The presence of sulfate and carboxyl groups in glycosaminoglycans gives them a large negative charge and the ability to bind water. Thanks to high density negative charge on their surface, they bind cations Ca 2+ , Na + , K + and thus take part in mineral metabolism.
All glycosaminoglycans are divided into 2 groups: sulfated and non-sulfated. Non-sulfated glycosaminoglycans include hyaluronic acid. Sulfated glycosaminoglycans do not occur in free form; when associated with a small amount of protein, they form proteoglycans. The structure of disaccharide units that make up glycosaminoglycans is shown in fig. 1.16.
Hyaluronic acid found in many organs and tissues. It is built from disaccharide residues connected by β-(1->4)-glycosidic bonds. Disaccharide fragments, in turn, consist of β-D-glucuronic acid and N-acetyl-(3-D-glucosamine) residues linked by β-(1-3)-glycosidic bonds. Hyaluronic acid has
Rice. 1.16.Structure of repeating disaccharide units in glycosaminoglycans.
high pier. mass (Mr 10 5 -10 7 Yes). In a number of organs (vitreous body of the eye, umbilical cord, articular fluid), it is in a free form, and in the cartilage it forms proteoglycan aggregates. In the joint fluid, hyaluronic acid plays the role of a lubricant, reducing friction between the joint surfaces. In the process of embryo development, it fills the intercellular spaces, facilitating the movement of cells. In large quantities, hyaluronic acid is synthesized during wound healing. By binding water, it provides a barrier function.
Hyaluronic acid chains are able to coagulate, binding large amounts of water and forming a domain. To this domain (defined
space) have access to small molecules or ions, but large molecules (albumin, immunoglobulins) are not able to penetrate into it. The domains are able to contact, shrink, and penetrate each other, which determines the high viscosity of the solution.
Chondroitin sulfates contain repeating disaccharide units connected by β-(1->4)-glycosidic bonds. Disaccharides are built from glucuronic acid and sulfated N-acetylgalactosamine, interconnected by (3-(1-3)-glycosidic bonds. Depending on the position of the sulfate group, chondroitin-4-sulfate and chondroitin-6-sulfate are distinguished. One the polysaccharide chain of chondroitin sulfate contains about 40 repeating disaccharide units.The molecular weight of chondroitin sulfates ranges from 10 to 600 kDa.Despite minimal differences in chemical structure, chondroitin sulfates differ significantly in physicochemical properties and distribution in various types connective tissue. Chondroitin-4-sulfate is predominantly found in cartilage and bone tissues, the cornea of the eye, and chondroitin-6-sulfate is present in tendons, ligaments, umbilical cord, and also in bones.
Dermatan sulfate - heteropolysaccharide similar in structure to chondroitin sulfate. In contrast to the latter, the disaccharide fragment of dermatan sulfate contains a L-iduronic acid residue instead of D-glucuronic acid. Dermatan sulfate is present in the skin, cartilage, tendons and intervertebral discs, blood vessels and heart valves. As part of small proteoglycans (biglycacan and decorin), it is found in the intercellular substance of bones, cartilage, intervertebral discs and menisci, where it participates in the stabilization of collagen fibers.
Keratan sulfates - the most heterogeneous glycosaminoglycans; differ from each other in the total content of carbohydrates and distribution in different tissues. Unlike all other glycosaminoglycans, keratan sulfates contain a D-galactose residue instead of uronic acid. The D-galactose residues in the disaccharide fragments of keratan sulfate are linked by β-(1->4)-glycosidic bonds to the residues of N-acetyl-D-glucosamine-6-sulfate. The disaccharide fragments are interconnected by β-(1->3)-glycosidic bonds.
Corneal keratan sulfate-1 contains, in addition to the repeating disaccharide unit, L-fucose, D-mannose and sialic acid. Keratan sulfate-2 is found in cartilage, bones, and intervertebral discs. Keratan sulfates are part of a large proteoglycan - agrecan and some small proteoglycans of the cartilage matrix.
Heparan sulfate is a heteropolysaccharide built from glucuronic acid and N-acetylglucosamine; contains more N-acyl groups and less sulfate. It is part of the basement membrane proteoglycans and is a constant component of the cell surface.
Large proteoglycans
TO large proteoglycans include proteins with a large mol. weighing more than 100 chains of glycosaminoglycans. This group includes agrecan, versican, neurocan, brevican, etc. Their feature is the ability to bind to collagens, hyaluronic acid and form proteoglycan aggregates.
There is a large chondroitin sulfate-containing proteoglycan in the cementum, dental pulp, mucous membrane, bone tissue and skin - versican , they say. whose mass is about 1000 kDa. The core protein of versican consists of amino acid sequences containing glu-glyphene residues. Due to the high content of sulfate, glutamic acid and the connection with hyaluronic acid, versican in the hydrated state occupies a significant amount of space.
The cartilage matrix is characterized by the presence of another large proteoglycan - agrecana (see cartilage).
Small proteoglycans
Small proteoglycans have a small core protein to which 1 or 2 chains of glycosaminoglycans are attached. Small proteoglycans include leucine-rich proteoglycans, cell-associated proteoglycans, and basement membrane proteoglycans.
Proteoglycans rich in leucine . A feature of small proteoglycans of this family is the presence of 9-12 leucine-rich domains in the C-terminal region of the core protein. These domains have the property to bind collagen. The N-terminal region is highly variable in its amino acid sequence, which is associated with glycosaminoglycans. Domains of the N-terminal region are involved in the interactions of proteins with each other and with cells.
The proteins of this family are represented by decorin, biglycan, fibromodulin, lumican, osteoaderin, osteoglycin, oculoglycan, opticin, and asporin.
Small proteoglycans - fibromodulin, lumican and osteoaderin in the N-terminal region contain chains keratan sulfate who are joining asparagine, as well as sulfated residues tyrosine.
Fibromodulin - proteoglycan with a mol. weighing about 40 kDa. It has been shown that fibromodulin attaches to type II collagen fibrils and limits their diameter.
Lumican its structure is very similar to fibromodulin. Present in the intercellular matrix of muscle and cartilage tissues, lungs, small intestine, cornea. It is supposed to be involved in the regulation of the formation of a reticulated collagen structure.
Osteoaderin - protein with mol. weighing 49.1 kDa. A feature of this protein is the presence of four sulfated tyrosine residues, three of which are located in the N-terminal region. The C-terminal region contains a large number of negatively charged amino acids. The osteoaderin molecule is synthesized by mature osteoblasts and also by odontoblasts. It is determined in the ameloblastic layer at the stage of enamel maturation and is involved in mineralization processes.
Decorin and Biglycan similar in size and structure, but their synthesis is under the control of different genes. Mol. the mass of decorin is about 130 kDa, and that of biglycan is about 270 kDa. Their core proteins contain a peculiar 24-amino acid sequence rich in leucine, which is repeated in tandem 10 times in decorin and 12 times in biglycan. Biglycan contains serine at positions 5 and 11, and decorin at position 4, which allows biglycan to attach 2 polysaccharide chains, and decorin only one (Fig. 1.17). These proteoglycans have polysaccharide chains dermatan sulfate. Decorin and biglycan are involved in intercellular interactions that can be facilitated by the (3-structure in the core protein. Decorin and probably biglycan have been shown to interact with β -transforming growth factor (TGF-(3).
The localization of decorin coincides with the location of collagen. If the purpose of biglycan is unknown, then decorin is involved in binding to collagen types I and II, and also inhibits fibrinolysis. In addition, biglycan and decorin provide interactions between cells, elastin and adhesive proteins - fibronectin and laminin.
Cell-associated proteoglycans
During cell development, small proteoglycans appear, which are called cell-associated proteoglycans. This family of proteins includes serglycins, syndecans, betaglycins, thrombomodulin, phosphatidylinositol - anchored proteoglycans.
Rice. 1.17.Domain structure of small proteoglycans: BUT - decorin ; B - biglycan ; IN - fibromodulin .
Syndecans include 4 types of different proteins. They are integral proteoglycans and contain intracellular, transmembrane and extracellular domains. The extracellular domain of these proteins is similar to the domain of proteinases and is able to open the cell membrane, and also contains varying chains of glycosaminoglycans connected to syndecan. Thus, syndecans 1 and 3 contain heparan sulfate and chondroitin sulfate. Syndecan-1 appears in epithelial cells during development, syndecan-2 (fibroglycan) is synthesized by fibroblast cells and hepatocytes; syndecan-3 (N-syndecan) is present in nervous tissue and developing cartilage, and syndecan-4 (ryudokan, amphiglycan) is present in endothelium, epithelium, smooth muscle cells, and skin fibroblasts. Syndecans bind collagens, fibronectin, thrombospondin, tenascin, and fibroblast growth factor through extracellular domains. The intracellular domains of syndecan bind to the cytoskeleton via actin.
Serglycins isolated from secretory vesicles. Their composition depends on cell type and cell differentiation. Chains of chondroitin- and heparan sulfate are associated with the core protein. A feature of serglycin molecules is the high content of sulfate residues, which makes them resistant to proteolysis. Mol. the mass of serglycins varies widely (60-750 kDa), and they say. the mass of the core protein is practically and constant (16-18 kDa).
It is believed that serglycins are involved in the regulation of the enzymatic activity of secretory granules and the differentiation of hematopoietic cells.
Some serglycins are synthesized by endothelial cells, and their synthesis is increased under the influence of tumor necrosis factor and interleukin 1α (IL-1α). Serglycine may be involved in the migration of leukocytes during inflammatory processes. It has recently been established that, together with other proteoglycans, they are involved in the adhesion and activation of lymphoid cells.
basement membrane proteoglycans
A whole group of heterogeneous proteoglycans containing heparan sulfate has been identified in the basement membranes. The structure of the core protein contains globular domains separated by rod fragments. Globular domains provide the connection of these proteoglycans with type IV collagen, laminin and other glycoproteins, as well as with cells located on the basement membrane.
Main heparan sulfate-containing basement membrane proteoglycan is perlecan . The polypeptide chain, consisting of 3500 amino acid residues, is linked to three heparan sulfate chains through hydroxyl groups. serine in the N-terminal region. Each polysaccharide chain contains up to 200 monomers. About three dozen globular domains are defined in the perlecan molecule, separated by short rod-like fragments, providing a connection between cells and components of the extracellular matrix.
The preservation of the biomechanical and physiological characteristics of the connective tissue is largely determined by maintaining a balance between the processes of biosynthesis and degradation of collagens and proteoglycans. The breakdown and synthesis of proteoglycans are regulated by: 1) hormones - somatotropin, thyroxine, insulin; 2) cytokines - IL-1, cachectins; 3) vitamins of groups A and C; 4) trace elements; 5) growth factors.
Synthesis of proteoglycans
The synthesis of proteoglycans begins with the biosynthesis of the core protein on polyribosomes. Already in the process of protein translation in the rough endoplasmic reticulum, trisaccharides are bound through the amide groups of asparagine residues. Dolichol-linked oligosaccharides with a high mannose content act as an oligosaccharide donor. After the addition of N-linked oligosaccharides, the core protein undergoes xylosylation and phosphorylation. UDP-xylosate-
ransferase, which carries out the transfer of xylose residues to the hydroxyl group of the core protein, is one of the key enzymes in the biosynthesis of proteoglycans. Further processes of formation of chains of glycosaminoglycans occur in the Golgi apparatus. Polysaccharide chains of glycosaminoglycans are synthesized by sequential addition of monosaccharides, the donors of which are usually the corresponding UDP sugars. Glycosyltransferases are localized on the membranes of the Golgi apparatus, with the participation of which the protein molecule undergoes glycosylation (Fig. 1.18).
Rice. 1.18.Attachment of glycosaminoglycan to the core protein through a binding trisaccharide. A binding oligosaccharide consisting of two galactose residues and one xylose residue is attached to serine, threonine, or asparagine through an O- or N-glycosidic bond.
UDP-galactosyltransferase I transfers the first galactose residue to xylose, UDP-galactosyltransferase II transfers the second galactose residue, and the formation of the binding tripeptide is completed by attaching a glucuronic acid residue to it. This reaction is catalyzed by UDP-glucuronyltransferase I. Further synthesis of the polysaccharide chain is carried out by the sequential addition of N-acetylgalactosamine (or N-acetylglucosamine, galactose) and glucuronic (or iduronic) acid (Fig. 1.19).
Modification of glycosaminoglycan chains is sulfation, that is, the addition of sulfate to C-4 and (or) to C-6 N-acetylga-
Rice. 1.19.Synthesis of chondroitin sulfate as part of proteoglycan. Enzymes: 1 - UDP-xylosyltransferase; 2 - UDP-galactosyltransferase I; 3 - UDPgalactosyltransferase II; 4 - UDP-glucuronyltransferase I; 5 - UDP-N-acetylgalactosamine transferase I; 6 - UDP-glucuronyltransferase II; 7 - UDP-N-acetylgalactosamine transferase II; 8 - sulfotransferase.
Rice. 1.20.The sulfation reaction of the N-acetylgalactosamine residue during the synthesis of the chondroitin sulfate chain.
lactosamine. The sulfate is transferred to the acceptor molecule by specific sulfotransferases (Fig. 1.20). The donor of the sulfate group is 3"-phosphoadenosine-5"-phosphosulfate (FAPS).
Amino sugars and hexuronic acids are synthesized from glucose. The immediate precursor of N-acetylglucosamine and N-acetylgalactosamine is fructose-6-phosphate. The source of the NH2 group for sugars is glutamine. The resulting amino sugar is further acetylated with acetyl-CoA (Fig. 1.21).
Rice. 1.21.Synthesis of glycosaminoglycans.
Enzymes: 1 - hexokinase; 2 - phosphoglucoisomerase; 3 - aminotransferase; 4 - acetyltransferase; 5 - N-acetylglucosamine phosphomutase; 6 - UDP-N-acetylglucosamine pyrophosphorylase; 7 - epimerase; 8 - UDP - glucosamine pyrophosphorylase; 9 - UDP-glucopyrophosphorylase; 10 - UDPglucose dehydrogenase.
In epimerization reactions, after the incorporation of glucuronate into the carbohydrate chain, L-iduronic acid is formed from D-glucuronic acid.
The synthesis of glycosaminoglycans is influenced by somatotropin and retinoic acid, which activate the incorporation of sulfate into the molecules. On the contrary, the synthesis of hyaluronic acid and sulfated glycosaminoglycans is inhibited by glucocorticoids and sex hormones.
The breakdown of proteoglycans
The breakdown of proteoglycans is a physiological process consisting in the regular renewal of extracellular and intracellular macromolecules. Proteinases and glycosidases are involved in the degradation of proteoglycans. Initially, core and binding proteins are exposed to free radicals and are hydrolyzed in the extracellular matrix by matrix metalloproteinases - collagenase, gelatinase, stromelysin. Proteinases cleave the core protein, and glycosidases hydrolyze the chains of glycosaminoglycans and oligosaccharides. All proteoglycans containing chondroitin sulfate, dermatan sulfate, heparan sulfate, and keratan sulfate chains are initially cleaved into fragments. Then fragments of proteoglycans are taken up by blast cells and subjected to intracellular degradation. These fragments can also be transported with lymph and blood to the liver. In hepatocytes, their further hydrolysis occurs, in which aspartyl, serine and other proteinases participate.
The breakdown of glycosaminoglycans
Glycosaminoglycans are distinguished by a high metabolic rate: the half-life (T 1/2) of many of them is from 3 to 10 days, and only for keratan sulfate T 1/2 - 120 days. The destruction of polysaccharide chains involves exo- and endoglycosidases (hyaluronidase, (3-glucuronidase, (3-galactosidase, (3-iduronidase)) and sulfatases.
Glycosaminoglycans enter the cell from the extracellular space by the mechanism of endocytosis, where endocytic vesicles merge with lysosomes. Active lysosomal enzymes provide complete gradual hydrolysis of glycosaminoglycans to monomers. Cleavage of intact glycosaminoglycans in cells begins with their breakdown into fragments under the action of endohexosaminidase and endoglucuronidase. The oligosaccharides formed in the hydrolysis reactions are subjected to successive actions of exoglycosidases and sulfatases, which cleave
monomers from the non-reducing end. Thus, the hydrolysis of chondroitin sulfate fragments containing an N-acetylgalactosamine residue at the non-reducing end is initiated by sulfatase, followed by β-N-acetylgalactosaminidase, and then (3-glucuronidase. As a result, inorganic sulfate and monosaccharides are formed (Fig. 1.22).
Rice. 1.22.breakdown of chondroitin sulfate.
Enzymes: 1 - endoglycosidase ; 2 - sulfatase; 3 - β - N-acetylgalactose-
minidase; 4 - β - glucuronidase.
Hyaluronidase is involved in the breakdown of hyaluronic acid to oligosaccharides. The hydrolysis of the resulting oligosaccharides is carried out by β-N-acetylglucosaminidase and β-D-glucuronidase.
Extracellular breakdown of glycosaminoglycans is characteristic only of heparan sulfate, which is cleaved by heparanase and synthesized by platelets or T-lymphocytes.
Mucopolysaccharidoses
Mucopolysaccharidoses - heavy hereditary diseases due to defects in hydrolases involved in the catabolism of glycosaminoglycans. In the lysosomes of tissues, which are characterized by the synthesis of the largest amount of glycosaminoglycans, incompletely destroyed glycosaminoglycans accumulate and their oligosaccharide fragments are excreted in the urine. There are several types of mucopolysaccharidoses caused by defects in various enzymes involved in the breakdown of glycosaminoglycans.
Mucopolysaccharidoses are manifested by mental disorders in children, lesions of the cardiovascular system, deformities of the bone skeleton, significantly pronounced in the maxillofacial region, hypoplasia of hard dental tissues, clouding of the cornea of the eyes, and a decrease in life expectancy (Table 1.3).
Currently, these diseases are not amenable to treatment, therefore, if a carrier of defective genes is suspected, prenatal diagnosis should be carried out. In these cases, the activity of lysosomal hydrolases is determined.
1.4. NON-COLLAGEN PROTEINS WITH SPECIAL PROPERTIES
Adhesive and anti-adhesive proteins
Proteins of the intercellular matrix perform the most different functions. Some of them have the ability to glue the components of the intercellular substance and cells, and these proteins are called adhesive. Another group of proteins, on the contrary, inhibits the adhesion of cells and extracellular components, and they are called anti-adhesive. The interaction of cells with the extracellular matrix is a complex process and is manifested both by increased adhesion and its weakening. The proteins fibronectin, vitronectin, laminin, nidogen (entactin), and integrins are involved in the adhesion of mesenchymal and epithelial cells. On the contrary, antiadhesive proteins - tenascin, thrombospondin are able to change the shape of cells and partially detach them from the components of the extracellular matrix. At the same time, such a division of proteins into adhesive and antiadhesive ones is rather conditional.
Table 1.3Diseases associated with impaired metabolism of glycosaminoglycans
fibronectin - high molecular weight glycoprotein, a key protein of the extracellular matrix, synthesized by fibroblasts. Depending on the ionic strength and pH of the extracellular matrix, the shape of the fibronectin molecule can vary from globular to intermediate. Fibronectin molecules are dimers consisting of two similar polypeptide chains linked by hydrophobic interactions and two disulfide bonds. The subunits are subdivided into a number of distinct domains capable of binding to cellular receptors via integrins, as well as collagens, fibrin, and proteoglycans. Linking crosswise with each other through disulfide bridges, fibronectin molecules form fibrillar structures (Fig. 1.23).
Rice. 1.23.The structure of the fibronectin molecule (BUT). Fibronectin molecule model (B). Numbers indicate domains linking: 1 - heparin , 2 - cells , 3 - collagen , 4 - other fibronectin molecules [according to Cooper G.M., 2000, with changes].
The fibronectin molecule has a binding site for the enzyme transglutaminase. Transglutaminase catalyzes the reaction of combining glutamine residues of one polypeptide chain with lysine residues of another protein molecule. This allows cross-linking of fibronectin molecules with each other, collagen and other proteins by transverse covalent bonds. Fibronectin is involved in multiple cellular processes including tissue repair, embryogenesis, migration, and cell adhesion.
Integrins are heterodimeric proteins with a mol. weighing 100-160 kDa, located on the plasma membrane of cells and consisting of two non-covalently bound transmembrane a- and (3-subunits. For the functioning of integrins, the presence of divalent ions (Ca 2+ or Mg 2+) is necessary, since the binding of the Ca 2+ cation allows N-terminal sections α- and (3-subunits to connect with each other and attach to the extracellular matrix. They are able to recognize the RGD peptide in matrix proteins ( arg-gli-asp).
The family of integrins includes 20 types of receptors with different specificities. This diversity is provided by the difference in the structure α- and (3-chains. 9 varieties of α-chains and 14 (3-subunits) are described. Each integrin chain crosses the membrane once. Both integrin chains have large extracellular domains. These domains provide cell adhesion to cells and to the components of the extracellular matrix - collagen, fibronectin, vitronectin, laminin (Fig. 1.24).
Due to their transmembrane orientation, integrins carry signals from the extracellular matrix to the cytoskeleton. Most integrins are associated with cytoplasmic C-terminal regions with actin-binding proteins of cells. When binding the ligand, the β-subunits of the binding integrins interact with the so-called proteins.
Rice. 1.24.Interaction of integrins with actin proteins of the cytoskeleton and extracellular matrix [according to Campbell N. A., Reece J. B., 2002, with changes].
attachment kami - talin and α-actinin, which, in turn, initiate the assembly of other connecting proteins. Thus, integrins bind to actin filaments. Actin filaments, through integrins, can change the orientation of secreted fibronectin molecules in the extracellular matrix. At the same time, the extracellular matrix can influence the organization of the cytoskeleton in target cells, which ensures two-way signal transmission. The binding of integrins to ligands and the convergence of cells are necessary for the rearrangement of the basement membrane.
The interaction of integrins with extracellular matrix proteins in some cases prevents apoptosis. The loss of some integrins (in breast, prostate, colon cancer) or their excess (in melanoma, squamous cell carcinoma of the oral cavity, nasopharynx, larynx) is associated with a high degree of tumor malignancy.
Thus, the information that integrins transmit from the extracellular matrix into the cell stimulates adhesion and migration of tumor cells in some cases, and leads to their death in others. In other words, integrins play the role of a kind of "switch" that determines the further fate of the tumor cell.
Laminins - representatives of the family of adhesive glycoproteins c mol. weighing 850 kDa. The laminin molecule is a large flexible complex consisting of long α-, β 1 -, β 2 -polypeptide chains associated in the form of an asymmetric cross and held together by disulfide bonds. Each chain contains several functional domains capable of binding to type IV collagen, heparan sulfate, entactin (nidogen), and cell surface receptors. Laminins glue epithelial cells to the basement membrane (Fig. 1.25).
In the early stages of morphogenesis, the basement membrane consists mainly of a laminin network and does not contain (or contains little) type IV collagen.
Laminins in the basement membrane are in complex with the nidogen protein, which is connected by the C-terminal domain to the (3 2 -chain of laminin. The N-terminal region of nidogen contains two globular domains, one of which binds to type IV collagen and the core proteoglycan perlecan protein. Thus, laminin, together with nidogen, provides the structural organization of the basement membrane components.
Rice. 1.25.The structure of laminin [according to Cooper G.M., 2000, with changes].
Laminins ensure the migration of epithelial cells and thus participate in odontogenesis, the binding of periodontal tissues to the cementum of the tooth root, the construction of the epithelial membrane on the surface of the pulp tissue during the formation of a pulp polyp.
Vitronectin - glycoprotein found in blood plasma and extracellular matrix. Vitronectin interacts with glycosaminoglycans, collagen, plasminogen, urokinase receptor. By stabilizing the inhibitory conformation of the plasminogen activation inhibitor 1 (proteinase), it regulates the degradation of the matrix. Through the binding of vitronectin to complement, heparin, and thrombin-antithrombin III complexes, it is involved in the immune response and regulation of blood coagulation. The polypeptide chain of vitronectin contains the amino acid sequence RGD, which ensures its interaction with α V β 3-integrin receptor and participation in attachment, spreading and movement of cells.
Tenascin And thrombospondin - glycoproteins with both adhesive and anti-adhesive properties. Tenascin and thrombospondin play a certain role in embryogenesis and morphogenesis. These proteins provide a change in cell shape under conditions in vitro, which, in turn, leads to shifts in the behavior of cells in culture. They contribute to the reorganization of the actin cytoskeleton by changing the adhesive contacts with protein factors that provide cell motility. Tenascin and thrombospondin form complexes with proteoglycans, and when tenascin binds to chondroitin sulfate, the adhesive properties of proteoglycans change.
Tenascin - oligomeric glycoprotein with a mol. weighing more than 100 kDa. The molecule of this protein has a mosaic structure, and the amino acid sequence is similar to the epidermal growth factor. Tenacin contains calcium-binding domains.
Thrombospondin - a glycoprotein that exhibits its antiadhesive properties in endothelial cells and fibroblasts, since with (3-transforming and platelet growth factors they weaken the binding of matrix molecules to each other.
Thrombospondin also exhibits adhesive properties when interacting with molecules of integrins, glycoproteins, heparan sulfate and
glycolipids. The globular domains contained in the N- and C-terminal regions promote the binding of calcium to heparin, after which thrombospondin interacts with collagen, fibronectin, fibrinogen, laminin, and plasminogen.
In addition to adhesive proteins involved in the organization of supramolecular complexes of the intercellular matrix, tissues contain glycoproteins related to growth factors.
growth factors
Growth factors are usually small polypeptides that stimulate or inhibit the proliferation of certain cell types. As a rule, they are secreted by one cell and act on other cells, although it sometimes happens that they act on the same cells that secrete them. Growth factors bind to their specific receptors localized on the surface of the cell membranes of their target cells. Most growth factors activate tyrosine protein kinases in cells, and only TGF-(3) activates threonine protein kinases.
transforming growth factor (TFR-(3) - a family of glycoproteins, including 6 different proteins. They are dimers consisting of two identical subunits. TGF-(3) proteins are synthesized as precursors, secreted in an inactive form, and activated by limited proteolysis.
On the cell membrane blast cells revealed 3 types of receptors for TGF. Receptors of the third type are surface proteoglycans and provide TGF-(3) access to receptors of the first and second types, which, after binding to TGF-(3), form a heterodimer with protein kinase activity. The cytoplasmic domain of the receptors is autophosphorylated at serine and threonine residues. Next, cytoplasmic proteins are phosphorylated, involved in signal transmission to the nucleus, where the transcription gene is activated.Through this mechanism, the synthesis of extracellular matrix proteins, such as type I collagen and metalloproteinases, is activated.
In addition, TGF-(3) acts as a chemotaxis factor for monocytes and fibroblasts. It inhibits the proliferation and function of T- and B-lymphocytes and endothelial cells. Among the complex network of cytokines that influence the function of odontoblasts in the process of dentin regeneration, TGF plays an important role -(3, which functions
It acts as a powerful immunosuppressant and inducer of extracellular matrix protein synthesis. TFR- β maintains homeostasis in the dentin-pulp complex during inflammation.
Morphogenetic bone protein (MBK) -an acidic glycophosphoprotein rich in serine and glycine containing three disulfide bonds. Restoration of disulfide bonds causes MBC inactivation. In the dental pulp, it is secreted in response to external stimuli by odontoblasts to form replacement dentin. MBC is very active in bone tissue and induces differentiation of stem cells into osteogenic ones.
endothelial growth factor (FRE) -a glycoprotein that binds only to vascular endothelial cells and stimulates their proliferation.
In addition, EGF can activate a specific protein, including a kinase complex. The resulting phosphorylated proteins cause cell movement, therefore, when the dental pulp, bone tissue, mucous membrane, periodontium and other tissues of the oral cavity are damaged under the influence of EGF, cells move rapidly, increase and differentiate with the activation of alkaline phosphatase.
EGF causes vasodilation, which is an important condition for maintaining blood flow in tissues during inflammation. It also increases the synthesis of IL-1, tumor necrosis factor (TNF), which contribute significantly to vasodilation in pathological processes. Dysregulation of the processes of endothelial growth factors is accompanied by an increase in osmotic pressure, pain and irreversible changes in the tissue.
Insulin-like growth factor (FMI)It has autocrine and paracrine effects. Its participation in the rapid growth of cells, their differentiation and mineralization of hard tissues of the tooth is assumed.
Fibroblast growth factor (FRF) -a family of structurally related polypeptides, represented by nine proteins. Mol. the mass of various forms of FGF ranges from 168 to 250 kDa. Up to 50% of the amino acid sequence of the fibroblast growth factor molecule corresponds to the structure of the endothelial growth factor. Both of these peptides also show similar affinity for heparin and cause vasodilation. Fibroblast growth factor is involved in the growth and differentiation of fibroblasts during the formation of a fibrous capsule around the inflammatory focus.
nerve growth factor (FRN) -a family of proteins that stimulate the growth of nervous tissue cells. Almost all human cells synthesize this factor. Nerve growth factor is involved in the rapid recovery of the damaged area due to the growth of axons from the damaged nerve trunk or from nearby intact nerve fibers. Thus, NGF may play an important role in the response of nerve cells to injury. The release of NGF into the oral cavity with saliva stimulates the healing of damaged areas of the mucous membrane.
Hepatocyte growth factor (Germany)stimulates cell proliferation in various tissues. It may be involved in cell aggregation in case of tissue damage, as well as in the morphogenesis of tooth tissues.
epidermal growth factor (EFR) -protein with mol. weighing 70 kDa. Distinguish α - and β-form EGF. It has an effect on ectoderm cells: skin keratinocytes, epitheliocytes of the mucous membrane of the oral cavity, esophagus, pharynx, as well as mesoderm: chondrocytes, vascular endothelium. Epidermal growth factor stimulates the differentiation of odontoblasts and increases DNA synthesis in them at the time of maturation of dental tissues. With age, EGF inhibits the division of odontoblasts, reduces the synthesis of type I collagen and reduces the activity of alkaline phosphatase. The production of EGF is influenced by steroid hormones, thyroxine and progesterone.
Platelet growth factor (FRT)affects many cells. Induces the synthesis of alkaline phosphatase and proteoglycans in the odontoblastic cells of the dental pulp and bone tissue.
1.5. PROTEIN CATABOLISM IN THE INTERCELLULAR MATRIX
Tissue remodeling is associated with cell differentiation and migration. A cell that has entered the path of differentiation inevitably dies. The emerging new cell begins to synthesize new own proteins, some of which enter the matrix.
In the catabolism of cell proteins and the extracellular matrix, the main role is assigned to matrix metalloproteinases (MMPs, matricins). Under physiological conditions, MMPs play a central role in the processes of morphogenesis, remodeling, and tissue resorption. Matrixins manifest their action in the intercellular matrix. The active center of these enzymes contains calcium or zinc, so they are called Ca 2+ -dependent zinc matrix metalloproteinases. More known
20 different metalloproteinases differing in substrate specificity and other properties. Based structural organization and substrate specificity, four main subfamilies of MMPs have been identified:
collagenases -trigger the hydrolysis of the helical region of collagen types I, II and III;
gelatinase -hydrolyze type IV collagen basement membranes;
stromelysins -cleave core proteins of proteoglycans and a number of adhesive matrix proteins;
metalloelastase - breaks down elastin.
Collagenases related to MMP-1 and MMP-13 are involved in the breakdown of native collagen, the half-life of which is measured in weeks or months. Collagenases cut through all three peptide α - chains of a native collagen molecule in a helical region, approximately 1/4 of the distance from the C-terminus, between glycine and leucine (or isoleucine) residues. The resulting collagen fragments become soluble in water and denature, after which their peptide bonds become available for hydrolysis by other peptidases.
Hydrolysis of basement membrane collagens occurs with the participation of gelatinases (MMP-2, MMP-9). The binding of gelatins and collagens by gelatinases involves the so-called fibronectin domain, which is present in the structure of the N-terminal region of the enzyme.
Two other enzymes - stromelysin -1 (MMP-3) and stromelysin - 2 (MMP-10), cleave the core proteins of proteoglycans and a number of adhesive proteins of the extracellular matrix (Table 1.4).
The activity of matrix metalloproteinases increases with the destruction of the intercellular matrix, which is observed in a number of diseases - periodontitis, pulpitis, chronic ulcers, invasion and metastasis of tumors, etc.
Regulation of the activity of matrix metalloproteinases
The activity of matrix metalloproteinases is under constant control.
Firstly,they are synthesized as preproenzymes. The signal peptide provides directed secretion of the molecule, and after its cleavage, a proenzyme is formed. The proenzyme contains an amino acid sequence in which the cysteine residue binds the Zn 2+ molecule located in the active center. Subsequently, after the cleavage of the polypeptide, the formed active form of MMP contains two main domains. The N-terminal domain contains a zinc-bound
MMP type | Enzyme | Mol. weight, kDa | Split components |
MMP-1 | Institial collagenase | Collagen I, II, III, VII, VIII, X types, gelatin, proteoglycans |
|
MMP-2 | Gelatinase A | Gelatin, collagen IV, V, VII, X, XI types, fibronectin, elastin, proteoglycans |
|
MMP-9 | Gelatinase B | Gelatin, collagen IV, V types, elastin, proteoglycans |
|
MMP-3 | Stromelizin-1 | Elastin, proteoglycans, laminin, fibronectin, type IV, VII, IX collagen, pro MMP-1 |
|
MMP-7 | Matrilysin | Proteoglycans, laminin, gelatin, fibronectin, type IV collagen, proMMP-1, -7, -8, -9 |
|
MMP-12 | Metal lo elastase | Elastin |
|
MMP-13 | Institial collagenase-3 | Collagen I, II, III types, gelatin |
|
MMP-14 | Membrane type MMP | Collagen I, II, III types, proMMP- 2, -13 (membrane type) |
a binding site in which Zn 2+ is bound by three histidine residues and has catalytic activity. In catalysis, in addition to zinc, the residue of glutamic acid takes part. The C-terminal domain is responsible for binding to substrates and MMP inhibitors. Between the N- and C-terminal domains is a small binding domain that provides substrate specificity (Fig. 1.26, A).
Various proteinases are involved in signal peptide cleavage. Thus, trypsin-like proteinase plasmin, proMMP-2 membrane metalloproteinase, and proMMP-9 gelatinase A participate in the activation reaction of proMMP-1 and proMMP-3. formed active enzymes (Fig. 1.26, B).
Rice. 1.26.Structure of proMMP-1: BUT - enzyme activation occurs upon cleavage of the signal propeptide; B - various proteinases are involved in the limited proteolysis of proMMPs.
Secondly, the activity of enzymes depends on the level of expression of their genes. Most MMPs are referred to as “inducible” enzymes, the synthesis of which at the transcriptional level is controlled by
factors: cytokines and other factors acting on the cell surface (estrogen, progesterone, etc.). MMP promoters contain common elements responsible for the regulation of gene expression.
Thirdly, under physiological conditions, tissues contain an insignificant amount of MMPs and their activity depends on the presence of activators and inhibitors in environment. MMP activity is under the control of specific proteins - tissue inhibitors of metalloproteinases (TIMPs). Currently, four types of TIMP isolated from various human tissues are well studied: TIMP-1, TIMP-2, TIMP-3, TIMP-4. TIMPs are able to bind to both active and inactive forms of MMPs. These proteins differ in their specific action on metalloproteinases. Thus, TIMP-1 inhibits MMP-9 much better, while TIMP-2 suppresses the activity of MMP-2. TIMPs are inactivated by hydrolysis involving various proteinases - trypsin, chymotrypsin, stromelysin-3, and neutrophil elastase.