home Children Regulation of the action of enzymes. How is enzyme activity regulated? The concept of the active center of the enzyme

Regulation of the action of enzymes. How is enzyme activity regulated? The concept of the active center of the enzyme

Their role in the regulation of enzyme activity.

General concepts of hormones and

LECTURE #3

Topic: Regulation of enzyme activity in the cell.

Faculties: medical and preventive, medical and preventive, pediatric.

One of the most important properties of living organisms is the ability to maintain homeostasis. Homeostasis in the body is maintained by regulating the rate of enzymatic reactions. This regulation is carried out at 3 independent levels:

I). Availability of substrate and coenzyme molecules;

II). Changes in the catalytic activity of enzyme molecules;

III). Change in the number of enzyme molecules.

I. Availability of substrate molecules provided by the controlled operation of transmembrane transport systems. Availability of coenzymes depends on the rate of their regeneration. The higher the concentration of the original substrate and regenerated coenzymes, the higher the rate of the metabolic pathway.

II. Regulation of the catalytic activity of enzymes. It happens:

1). Non-specific regulation. Due to the lability of all enzymes, their catalytic activity is nonspecifically affected by temperature, pH, and pressure.

2). Specific regulation. Under the action of specific activators and inhibitors, the activity of regulatory enzymes that control the intensity of metabolic processes in the body changes.

The main ways of regulating the catalytic activity of enzyme molecules include:

one). allosteric regulation;

2). Regulation by protein-protein interactions;

3). Regulation by phosphorylation/dephosphorylation of the enzyme molecule;

4). Regulation by partial proteolysis.

The last two methods refer to the covalent modification of the enzyme. Regulation occurs as a result of either covalent attachment of the fragment to the enzyme or cleavage of the fragment from the enzyme.

one). Allosteric regulation.

Allosteric enzymes are enzymes whose activity is regulated by the reversible non-covalent attachment of a modulator (activator and inhibitor) to a special allosteric center. Allosteric enzymes are oligomeric proteins or have a domain structure. These enzymes play an important role in the regulation of react extremely quickly to changes in the environment. Inhibitors of allosteric enzymes are often the end products of metabolic pathways, while activators are their initial substrates. Activation occurs according to the principle of direct positive communication, and inhibition - according to the principle of negative feedback.

For example, the end product of glucose catabolism, ATP, allosterically inhibits the glycolytic enzymes phosphofructokinase and pyruvate kinase. Fructose-1,6-f accumulated in glycolysis activates pyruvate kinase, which accelerates glycolysis reactions.

2). Regulation of the catalytic activity of enzymes through protein-protein interactions. There are 2 mechanisms:

a). Activation of enzymes as a result of the addition of regulatory proteins. For example, adenylate cyclase (Ac), which catalyzes the conversion of cAMP from ATP, is activated by the addition of the α subunit of the G protein. This regulatory mechanism is reversible.

b). Regulation of catalytic activity of enzymes by association/dissociation of protomers. For example, protein kinase A is activated upon dissociation of its tetramer into 4 subunits and is inactivated upon the reverse connection of 4 subunits into the tetramer.

3). Regulation of catalytic activity of enzymes by their phosphorylation/dephosphorylation. Phosphorylation is carried out by protein kinases, and dephosphorylation by phosphoprotein phosphatases. The introduction of a negatively charged phosphorus group leads to a reversible change in the conformation and activity of the enzyme.

For example, under the action of glucagon in cells, there is a decrease in the synthesis of fat, glycogen and an increase in its breakdown caused by phosphorylation of the key enzymes of these processes. And under the action of insulin, on the contrary, glycogen synthesis is activated and its breakdown is inhibited, since insulin causes dephosphorylation of the same key enzymes.

4). Regulation of the catalytic activity of enzymes by partial proteolysis. With the participation of activators and proteolytic enzymes, a part of the enzyme molecule is cleaved off and its irreversible activation occurs. These enzymes function little and then are destroyed. It is typical for extracellular enzymes of the gastrointestinal tract (pepsin, trypsin, chymotrypsin, etc.) and blood coagulation enzymes (thrombin, fibrin, plasmin, etc.). For example, trypsinogen synthesized in the pancreas enters the duodenum, where intestinal enteropeptidase cleaves off a hexapeptide from its N-terminus. As a result, an active center is formed in the remaining part of the enzyme molecule.

III. Mechanisms of regulation of the number of enzymes: induction, repression, derepression.

The amount of the enzyme in the cell depends on the rate of its synthesis and decay, the processes necessary for the renewal of the enzyme. Synthesis and degradation of enzymes are regulated independently. The mechanism of regulation of enzyme synthesis at the level of transcription, which is carried out by certain metabolites, hormones, and a number of biologically active substances, has been studied the most.

Induction– triggering the synthesis of enzymes under the action of specific low molecular weight compounds – inductors . For example, in bacteria, enzymes are synthesized only in the presence of substrates for them, which are inductors for these enzymes (in E. coli, lactose is an inducer of β-galactosidase).

Inducible Enzymes- enzymes, the concentration of which depends on the addition of inducers (organ-specific enzymes).

Constitutive enzymes- enzymes, the concentration of which is constant and not regulated by inducers (enzymes of glycolysis, RNA synthesis, etc.).

A basic level of is the concentration of the enzyme in the absence of the inducer. At induction a basic level of enzyme can be exceeded from 2 to 1000 times.

Repression- stop the synthesis of the enzyme under the action repressor (more precisely corepressor ).

Derepression– resumption of enzyme synthesis either after removal of the repressor from the medium or when its reserves are depleted.

Often, one inducer or repressor regulates the synthesis of several enzymes involved in the same metabolic pathway. The genes for these enzymes are assembled into an operon.

Operon- a section of a DNA molecule that contains information about a group of functionally interconnected structural genes and a regulatory zone, promoter that controls the transcription of these genes. The induction and repression of the synthesis of operon enzymes is called coordinated.

Coordinated induction– all enzymes encoded by operon genes are induced by one inducer.

Coordinated repression– stopping the synthesis of operon enzymes by one repressor.

The operon promoter has a region operator required for the attachment of the repressor. The repressor is synthesized from a regulator gene.

1). Operon induction . The repressor attaches to the operator and prevents attachment to the RNA polymerase promoter - there is no mRNA transcription. The inductor attaches to the repressor and causes separation of the latter from the operator, RNA polymerase attaches to the promoter, and mRNA transcription from the operon begins. Several enzymes are synthesized from this mRNA at once.

2). Operon repression . The repressor does not attach itself to the operator. Attachment to the operator of the repressor occurs only in combination with the corepressor. The repressor-corepressor complex, when attached to the operator, prevents the attachment of RNA polymerase to the promoter, which blocks the transcription of mRNA from the operon and the synthesis of enzymes.

catabolic repression- a more favorable oxidation substrate inhibits the oxidation enzymes of a less favorable substrate.

The breakdown of enzymes occurs under the action of hydrolytic enzymes; the regulation of this process is poorly understood.

Biological chemistry Lelevich Vladimir Valeryanovich

Chapter 4. Regulation of enzyme activity. Medical Enzymology

Ways to regulate enzyme activity:

1. Change in the amount of enzymes.

2. Change in the catalytic efficiency of the enzyme.

3. Changing the reaction conditions.

Enzyme regulation

The number of enzyme molecules in a cell is determined by the ratio of two processes - the rates of synthesis and decay of the protein molecule of the enzyme.

There are two types of enzymes in cells:

1. Constitutive enzymes- are essential components of the cell, synthesized at a constant rate in constant quantities.

2. Adaptive enzymes- their formation depends on certain conditions. Among them, inducible and repressible enzymes are distinguished.

Inducible, as a rule, are enzymes with a catabolic function. Their formation can be caused or accelerated by the substrate of this enzyme. Anabolic enzymes are usually repressed. The inhibitor (repressor) of the synthesis of these enzymes can be the final product of this enzymatic reaction.

Change in catalytic efficiency of enzymes

This type of regulation can be carried out by several mechanisms.

Effect of Activators and Inhibitors on Enzyme Activity

Activators can increase enzymatic activity in different ways:

1. form the active center of the enzyme;

2. facilitate the formation of an enzyme-substrate complex;

3. stabilize the native structure of the enzyme;

4. protect the functional groups of the active site.

Classification of enzyme inhibitors:

1. Non-specific.

2. Specific:

irreversible

Reversible:

§ competitive

§ non-competitive.

Nonspecific inhibitors cause denaturation of the enzyme molecule - these are acids, alkalis, salts of heavy metals. Their action is not associated with the mechanism of enzymatic catalysis.

irreversible inhibition

Irreversible inhibition is observed in the case of the formation of covalent stable bonds between the inhibitor molecule and the enzyme. Most often, the active site of the enzyme undergoes modification. As a result, the enzyme cannot perform its catalytic function.

Irreversible inhibitors include heavy metal ions, such as mercury (Hg 2+), silver (Ag +) and arsenic (As 3+), which block the sulfhydryl groups of the active center in low concentrations. In this case, the substrate cannot undergo chemical transformation.

Diisopropylfluorophosphate (DPP) reacts specifically with only one of the many serine residues in the active site of the enzyme. The Ser residue capable of reacting with DPP has an identical or very similar amino acid environment. The high reactivity of this residue compared to other Ser residues is due to amino acid residues that are also included in the active center of enzymes.

DPP is classified as a specific irreversible inhibitor of "serine" enzymes, since it forms covalent bond with the hydroxyl group of serine, which is located in the active center and plays a key role in the process of catalysis.

Monoiodoacetic acid, p-chloromercuribenzoate easily react with SH-groups of protein cysteine residues. These inhibitors are not classified as specific, since they react with any free SH-groups of proteins and are called non-specific inhibitors. If the SH groups are directly involved in catalysis, then with the help of these inhibitors it seems possible to reveal the role of the SH groups of the enzyme in catalysis.

Irreversible enzyme inhibitors as drugs

An example of a drug whose action is based on irreversible inhibition enzymes, a widely used drug is aspirin. The anti-inflammatory nonsteroidal drug aspirin provides a pharmacological effect by inhibiting the cyclooxygenase enzyme, which catalyzes the formation of prostaglandins from arachidonic acid. As a result of a chemical reaction, the acetyl residue of aspirin is attached to the free terminal OH group of the cyclooxygenase serine.

This causes a decrease in the formation of prostaglandin reaction products, which have a wide spectrum biological functions, including mediators of inflammation.

Reversible inhibition

Reversible inhibitors bind to the enzyme by weak non-covalent bonds and, under certain conditions, are easily separated from the enzyme. Reversible inhibitors are either competitive or non-competitive.

Competitive inhibition

Competitive inhibition refers to a reversible decrease in the rate of an enzymatic reaction caused by an inhibitor that binds to the active site of the enzyme and prevents the formation of the enzyme-substrate complex. This type of inhibition is observed when the inhibitor is a structural analogue of the substrate; as a result, there is competition between the substrate and inhibitor molecules for a place in the active site of the enzyme. In this case, either the substrate or the inhibitor interacts with the enzyme, forming enzyme-substrate (ES) or enzyme-inhibitor (EI) complexes. When the complex of the enzyme and the inhibitor (EI) is formed, the reaction product is not formed.

A classic example of competitive inhibition is the inhibition of the succinate dehydrogenase reaction by malonic acid. Malonic acid is a structural analogue of succinate (the presence of two carboxyl groups) and can also interact with the active site of succinate dehydrogenase. However, the elimination of two hydrogen atoms from malonic acid is impossible; therefore, the rate of reaction is reduced.

Drugs as competitive inhibitors

Many drugs exert their therapeutic effect through the mechanism of competitive inhibition. For example, quaternary ammonium bases inhibit acetylcholinesterase, which catalyzes the hydrolysis of acetylcholine to choline and acetic acid.

When inhibitors are added, the activity of acetylcholinesterase decreases, the concentration of acetylcholine (substrate) increases, which is accompanied by an increase in the conduction of a nerve impulse. Cholinesterase inhibitors are used in the treatment of muscular dystrophies. Effective anticholinesterase drugs - prozerin, endrophonium, etc.

Antimetabolites as drugs

As inhibitors of enzymes by a competitive mechanism in medical practice, substances called antimetabolites are used. These compounds, being structural analogues of natural substrates, cause competitive inhibition of enzymes, on the one hand, and, on the other hand, can be used by the same enzymes as pseudosubstrates, which leads to the synthesis of abnormal products. Abnormal products do not have functional activity; as a result, a decrease in the speed of certain metabolic pathways is observed.

The following antimetabolites are used as drugs: sulfanilamide preparations (analogues of para-aminobenzoic acid) used to treat infectious diseases, nucleotide analogs for the treatment of oncological diseases.

Noncompetitive inhibition

Such inhibition of an enzymatic reaction is called non-competitive, in which the inhibitor interacts with the enzyme in a site other than the active site. Non-competitive inhibitors are not structural analogues of the substrate.

A non-competitive inhibitor can bind to either the enzyme or the enzyme-substrate complex to form an inactive complex. Attachment of a non-competitive inhibitor causes a change in the conformation of the enzyme molecule in such a way that the interaction of the substrate with the active site of the enzyme is disrupted, which leads to a decrease in the rate of the enzymatic reaction.

Allosteric regulation

Allosteric enzymes are enzymes whose activity is regulated not only by the number of substrate molecules, but also by other substances called effectors. The effectors involved in allosteric regulation are often cellular metabolites of the very pathway they regulate.

The role of allosteric enzymes in cell metabolism. Allosteric enzymes play an important role in metabolism, as they react extremely quickly to the slightest changes in the internal state of the cell.

Allosteric regulation is of great importance in the following situations:

1. with anabolic processes. Inhibition by the end product of the metabolic pathway and activation by the initial metabolites allow regulation of the synthesis of these compounds;

2. during catabolic processes. In the case of ATP accumulation in the cell, the metabolic pathways that provide energy synthesis are inhibited. In this case, the substrates are spent on the reactions of storage of reserve nutrients;

3. to coordinate anabolic and catabolic pathways. ATP and ADP are allosteric effectors acting as antagonists;

4. to coordinate parallel flowing and interconnected metabolic pathways (for example, the synthesis of purine and pyrimidine nucleotides used for the synthesis nucleic acids). Thus, the end products of one metabolic pathway may be allosteric effectors of another metabolic pathway.

Features of the structure and functioning of allosteric enzymes:

1. usually these are oligomeric proteins, consisting of several protomers or having a domain structure;

2. they have an allosteric center spatially remote from the catalytic active center;

3. effectors attach to the enzyme non-covalently in allosteric (regulatory) centers;

4. allosteric centers, just like catalytic ones, can exhibit different specificity with respect to ligands: it can be absolute and group specific.

Some enzymes have several allosteric centers, some of which are specific to activators, others to inhibitors;

1. the protomer on which the allosteric center is located is a regulatory protomer, in contrast to the catalytic protomer containing the active center, in which chemical reaction;

2. allosteric enzymes have the property of cooperativity: the interaction of the allosteric effector with the allosteric center causes a consistent cooperative change in the conformation of all subunits, leading to a change in the conformation of the active center and a change in the affinity of the enzyme to the substrate, which reduces or increases the catalytic activity of the enzyme;

3. regulation of allosteric enzymes is reversible: detachment of the effector from the regulatory subunit restores the initial catalytic activity of the enzyme;

4. allosteric enzymes catalyze the key reactions of this metabolic pathway.

Regulation of the catalytic activity of enzymes by protein-protein interactions.

Some enzymes change their catalytic activity as a result of protein-protein interactions.

There are 2 mechanisms of enzyme activation using protein-protein interactions:

1. activation of enzymes as a result of the attachment of regulatory proteins;

2. a change in the catalytic activity of enzymes due to the association or dissociation of enzyme protomers.

Regulation of the catalytic activity of enzymes by phosphorylation/dephosphorylation.

V biological systems often there is a mechanism for regulating the activity of enzymes with the help of covalent modification of amino acid residues. A fast and widespread method of chemical modification of enzymes is phosphorylation/dephosphorylation. The OH groups of the enzyme undergo modifications. Phosphorylation is carried out by protein kinases, and dephosphorylation by phosphoprotein phosphatases. Attaching the remainder phosphoric acid leads to a change in the conformation of the active center and its catalytic activity. In this case, the result can be twofold: some enzymes are activated during phosphorylation, while others, on the contrary, become less active.

Regulation of the catalytic activity of enzymes by partial (limited) proteolysis.

Some enzymes that function outside cells (in the gastrointestinal tract or in blood plasma) are synthesized as inactive precursors and are activated only as a result of hydrolysis of one or more specific peptide bonds, which leads to the cleavage of part of the precursor protein molecule. As a result, a conformational rearrangement occurs in the remaining part of the protein molecule and the active center of the enzyme (trypsinogen - trypsin) is formed.

Plasma enzymes

By origin, plasma enzymes can be divided into 3 groups.

1. Own blood plasma enzymes (secretory). They are formed in the liver, but show their action in the blood. These include enzymes of the blood coagulation system - prothrombin, proaccelerin, proconvertin, as well as ceruloplasmin, cholinesterase.

2. Excretory enzymes - enter the blood from various secrets - duodenal juice, saliva, etc. These include amylase, lipase.

3. Cellular enzymes - enter the blood when cells or tissues are damaged or destroyed.

Table 4.1. Organ-specific enzymes (isoenzymes)

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Chapter 27. Regulation and Interrelationship of Metabolism For the normal functioning of the body, there must be a precise regulation of the flow of metabolites along anabolic and catabolic pathways. All related chemical processes should flow at speeds

Enzymes are regulated catalysts. Metabolites, poisons can act as regulators. Distinguish:

- activators- substances that increase the rate of a reaction;

- inhibitors- Substances that slow down the rate of a reaction.

Enzyme activation. Various activators can bind either to the active site of the enzyme or outside of it. The group of activators that affect the active center includes: metal ions, coenzymes, the substrates themselves.

Activation with the help of metals proceeds through various mechanisms:

The metal is part of the catalytic site of the active center;

The metal with the substrate form a complex;

Due to the metal, a bridge is formed between the substrate and the active site of the enzyme.

Substrates are also activators. As the substrate concentration increases, the reaction rate increases. upon reaching the saturation concentration of the substrate, this rate does not change.

If the activator binds outside the active site of the enzyme, then covalent modification of an enzyme:

1) partial proteolysis (limited proteolysis). Thus, the digestive canal enzymes are activated: pepsin, trypsin, chymotrypsin. Trypsin has the state of trypsinogen proenzyme, consisting of 229 AA residues. Under the action of the enterokinase enzyme and with the addition of water, it is converted to trypsin, and the hexapeptide is cleaved off. The tertiary structure of the protein changes, the active center of the enzyme is formed and it passes into the active form.

2) phosphorylation - dephosphorylation. Ex: lipase + ATP = (protein kinase) phosphorylated lipase + ADP. This is a transfer reaction using ATP phosphate. In this case, a group of atoms is transferred from one molecule to another. Phosphorylated lipase is the active form of the enzyme.

Phosphorylase is activated in the same way: phosphorylase B+ 4ATP = phosphorylase A+ 4ADP

Also, when the activator is bound outside the active site, dissociation of the inactive complex protein-active enzyme. For example, protein kinase is an enzyme that performs phosphorylation (cAMP-dependent). Protein kinase is a protein having a quaternary structure and consisting of 2 regulatory and 2 catalytic subunits. R 2 C 2 + 2cAMP \u003d R 2 cAMP 2 + 2C. This type of regulation is called allosteric regulation (activation).

Enzyme inhibition. An inhibitor is a substance that causes specific decrease in enzyme activity. A distinction should be made between inhibition and inactivation. Inactivation is, for example, the denaturation of a protein as a result of the action of denaturing agents.

By bonding strength Inhibitors with enzyme Inhibitors are divided into reversible and irreversible.

irreversible inhibitors are strongly bound and destroy the functional groups of the enzyme molecule, which are necessary for the manifestation of its catalytic activity. All protein purification procedures do not affect the binding of the inhibitor and the enzyme. Ex: the action of organophosphorus compounds on the enzyme - cholinesterase. Chlorophos, sarin, soman and other phosphorus organic compounds bind to the active site of cholinesterase. As a result, the catalytic groups of the active center of the enzyme are phosphorylated. As a result, the enzyme molecules associated with the inhibitor cannot bind to the substrate and severe poisoning occurs.

Also allocate reversible inhibitors, such as proserin for cholinesterase. Reversible inhibition depends on the concentration of substrate and inhibitor and is removed by an excess of substrate.

According to the mechanism of action allocate:

Competitive inhibition;

Noncompetitive inhibition;

Substrate inhibition;

Allosteric.

1) Competitive (isosteric) inhibition- this is the inhibition of the enzymatic reaction caused by the binding of the inhibitor to the active site of the enzyme. In this case, the inhibitor is similar to the substrate. In the process, there is competition for the active center: enzyme-substrate and inhibitor-enzyme complexes are formed. E+S®ES® EP® E+P; E+I® E. Ex: succinate dehydrogenase reaction [Fig. COOH-CH 2 -CH 2 -COOH® (above the arrow LDH, under FAD®FADN 2) COOH-CH=CH-COOH]. The true substrate of this reaction is succinate (amber to-ta). Inhibitors: malonic acid (COOH-CH 2 -COOH) and oxaloacetate (COOH-CO-CH 2 -COOH). [rice. 3-hole enzyme + substrate + inhibitor = inhibitor-enzyme complex]

Ex: the cholinesterase enzyme catalyzes the conversion of acetylcholine to choline: (CH 3) 3 -N-CH 2 -CH 2 -O-CO-CH 3 ® (above the ChE arrow, under - water) CH 3 COOH + (CH 3) 3 - N-CH 2 -CH 2 -OH. Competitive inhibitors are prozerin, sevin.

2) Noncompetitive inhibition– inhibition associated with the effect of the inhibitor on the catalytic conversion, but not on the binding of the enzyme to the substrate. In this case, the inhibitor can bind both to the active center (catalytic site) and outside it.

Attachment of an inhibitor outside the active center leads to a change in the conformation (tertiary structure) of the protein, as a result of which the conformation of the active center changes. This affects the catalytic site and interferes with the interaction of the substrate with the active site. In this case, the inhibitor is not similar to the substrate, and this inhibition cannot be removed by an excess of the substrate. The formation of triple enzyme-inhibitor-substrate complexes is possible. The speed of such a reaction will not be maximum.

Non-competitive inhibitors include:

cyanides. They bind to the iron atom in cytochrome oxidase and as a result, the enzyme loses its activity, and since. is an enzyme of the respiratory chain, then the respiration of cells is disturbed and they die.

Ions of heavy metals and their organic compounds (Hg, Pb, etc.). The mechanism of their action is associated with their connection with various SH-groups. [rice. enzyme with SH-groups, mercury ion, substrate. All this combines into a triple complex]

A number of pharmacological agents that should affect the enzymes of malignant cells. This includes inhibitors used in agriculture, household poisonous substances.

3) substrate inhibition- inhibition of the enzymatic reaction caused by an excess of the substrate. Occurs as a result of the formation of an enzyme-substrate complex that is unable to undergo catalytic transformation. It can be removed and the substrate concentration reduced. [rice. enzyme binding to 2 substrates at once]

4) Allosteric inhibition - inhibition of the enzymatic reaction caused by the addition of an allosteric inhibitor in the allosteric center of the allosteric enzyme. This type of inhibition is characteristic of allosteric enzymes having a quaternary structure. Metabolites, hormones, metal ions, coenzymes can act as inhibitors.

Mechanism of action:

a) attaching an inhibitor to an allosteric center;

b) the conformation of the enzyme changes;

c) the conformation of the active center changes;

d) the complementarity of the active center of the enzyme to the substrate is disturbed;

e) the number of ES molecules decreases;

e) the rate of the enzymatic reaction decreases.

[rice. enzyme with 2 holes, to one is an allosteric inhibitor and the second changes shape]

The features of allosteric enzymes include inhibition by negative feedback. A®(E 1)B®(E 2) C®(E 3) D (from D arrow to arrow between A and B). D is a metabolite that acts as an allosteric inhibitor on the E 1 enzyme.

Metabolism

Metabolism (metabolism) is a set of physiological and biochemical processes that ensure the vital activity of the organism in interrelations with the external environment, aimed at self-reproduction and self-preservation.

Physiological processes include digestion, absorption, external respiration, excretion, etc.; to biochemical - chemical transformations of proteins, fats, carbohydrates entering the body in the form of nutrients. A feature of biochemical processes is that they are carried out in the course of a series of enzymatic reactions. It is enzymes that provide a certain sequence, places and speed of reactions.

According to the direction, all chemical transformations are divided into:

a) dissimilation(catabolism) - the breakdown of substances to simpler ones with the transition of the energy of the bonds of the substance into the energy of high-energy bonds (ATP, NADH, etc.);

b) assimilation(anabolism) - synthesis of more complex substances from simpler ones with energy expenditure.

biological significance These two processes lies in the fact that during the breakdown of substances, the energy contained in them is released, which provides all the functional capabilities of the body. At the same time, during the decay of substances, "building materials" (monosaccharides, AA, glycerin, etc.) are formed, which are then used in the synthesis of substances specific to the body (proteins, fats, carbohydrates, etc.).

[SCHEME] Above the horizontal line (in the external environment) - "proteins, fats, carbohydrates", from them an arrow down under the line (inside the body) to the inscription "dissimilation", from the last four arrows: two up to the inscriptions above the line "warmth" and "final products"; one arrow to the right to the inscription "intermediate substances (metabolites)", from them to "assimilation", then to "own proteins, fats, carbohydrates"; one arrow down to the inscription "ATP energy", from it - to "muscle contraction, conduction of a nerve impulse, secretion, etc." and also up to "warmth" and "assimilation".

The dissimilation of proteins, fats and carbohydrates proceeds in different ways, but there are a number of common steps in the destruction of these substances:

1) Digestion stage. In the gastrointestinal tract, proteins break down to AA, fats - to glycerol and VFA, carbohydrates - to monosaccharides. Is being developed a large number of non-specific substances from specific, coming from outside. Due to digestion, about 1% of the chemical energy of substances is released in the gastrointestinal tract. This stage is necessary so that the substances that come with food can be absorbed.

2) The stage of interstitial metabolism (tissue metabolism, metabolism). At the cellular level, it is divided into anabolism and catabolism. Metabolic intermediates are formed and converted - metabolites. At the same time, the monomers formed during the digestion stage decompose with the formation of a small (up to five) key intermediates: PAA, alpha-KG, acetyl-CoA, PVC, alpha-glycerophosphate. Up to 20% of the energy of substances is released. As a rule, interstitial exchange occurs in the cytoplasm of cells.

3) Final disintegration substances containing oxygen final products(CO 2 , N 2 Oh, nitrogen-containing substances). Approximately 80% of the energy of substances is released.

All the considered stages reflect only the main forms of metabolic processes. Both in the second and in the third stages, the released energy is accumulated in the form of energy chemical bonds high-energy compounds (these are substances that have at least one high-energy bond, for example, ATP, CTP, TTP, GTP, UTP, ADP, CDP, ..., creatine phosphate, 1,3-diphosphoglyceric acid). Thus, the binding energy of the last phosphate in ATP molecule is about 10-12 kcal/mol.

Biological role metabolism:

1. energy accumulation during decay chemical compounds;

2. the use of energy for the synthesis of the body's own substances;

3. disintegration of the renewed structural components of the cell;

4. there is a synthesis and disintegration of biomolecules for special purposes.

Protein metabolism

Regulation of enzyme activity. Medical enzymology (biochemistry)

Ways to regulate enzyme activity:

1. Change in the amount of enzymes.

2. Change in the catalytic efficiency of the enzyme.

3. Changing the reaction conditions.

Enzyme regulation

The number of enzyme molecules in a cell is determined by the ratio of two processes - the rates of synthesis and decay of the protein molecule of the enzyme.

There are two types of enzymes in cells:

1. Constitutive enzymes- are essential components of the cell, synthesized at a constant rate in constant quantities.

2. Adaptive enzymes- their formation depends on certain conditions. Among them, inducible and repressible enzymes are distinguished.

Change in catalytic efficiency of enzymes

This type of regulation can be carried out by several mechanisms.

Effect of Activators and Inhibitors on Enzyme Activity

Activators can increase enzymatic activity in different ways:

1. form the active center of the enzyme;

2. facilitate the formation of an enzyme-substrate complex;

3. stabilize the native structure of the enzyme;

4. protect the functional groups of the active site.

Classification of enzyme inhibitors:

1. Non-specific.

2. Specific:

irreversible

Reversible:

§ competitive

§ non-competitive.

Nonspecific inhibitors cause denaturation of the enzyme molecule - these are acids, alkalis, salts of heavy metals. Their action is not associated with the mechanism of enzymatic catalysis.

irreversible inhibition

DPP is classified as a specific irreversible inhibitor of "serine" enzymes, since it forms a covalent bond with the hydroxyl group of serine, which is located in the active center and plays a key role in the catalysis process.

Irreversible enzyme inhibitors as drugs

An example of a drug whose action is based on irreversible enzyme inhibition is the widely used drug aspirin. The anti-inflammatory non-steroidal drug aspirin provides a pharmacological effect by inhibiting the cyclooxygenase enzyme, which catalyzes the formation of prostaglandins from arachidonic acid. As a result of a chemical reaction, the acetyl residue of aspirin is attached to the free terminal OH group of the cyclooxygenase serine.

This causes a decrease in the formation of prostaglandin reaction products, which have a wide range of biological functions, including mediators of inflammation.

Reversible inhibition

Competitive inhibition

Drugs as competitive inhibitors

Antimetabolites as drugs

Noncompetitive inhibition

A non-competitive inhibitor can bind to either the enzyme or the enzyme-substrate complex to form an inactive complex. The addition of a non-competitive inhibitor causes a change in the conformation of the enzyme molecule in such a way that the interaction of the substrate with the active site of the enzyme is disrupted, which leads to a decrease in the rate of the enzymatic reaction.

Allosteric regulation

The role of allosteric enzymes in cell metabolism. Allosteric enzymes play an important role in metabolism, as they react extremely quickly to the slightest changes in the internal state of the cell.

Allosteric regulation is of great importance in the following situations:

1. with anabolic processes. Inhibition by the end product of the metabolic pathway and activation by the initial metabolites allow regulation of the synthesis of these compounds;

3. to coordinate anabolic and catabolic pathways. ATP and ADP are allosteric effectors acting as antagonists;

4. to coordinate parallel flowing and interconnected metabolic pathways (for example, the synthesis of purine and pyrimidine nucleotides used for the synthesis of nucleic acids). Thus, the end products of one metabolic pathway may be allosteric effectors of another metabolic pathway.

Features of the structure and functioning of allosteric enzymes:

1. usually these are oligomeric proteins, consisting of several protomers or having a domain structure;

2. they have an allosteric center spatially remote from the catalytic active center;

3. effectors attach to the enzyme non-covalently in allosteric (regulatory) centers;

4. allosteric centers, just like catalytic ones, can exhibit different specificity with respect to ligands: it can be absolute and group specific.

Some enzymes have several allosteric centers, some of which are specific to activators, others to inhibitors;

1. the protomer on which the allosteric center is located is a regulatory protomer, in contrast to the catalytic protomer containing the active center in which the chemical reaction takes place;

3. regulation of allosteric enzymes is reversible: detachment of the effector from the regulatory subunit restores the initial catalytic activity of the enzyme;

4. allosteric enzymes catalyze the key reactions of this metabolic pathway.

Regulation of the catalytic activity of enzymes by protein-protein interactions.

Some enzymes change their catalytic activity as a result of protein-protein interactions.

There are 2 mechanisms of enzyme activation using protein-protein interactions:

1. activation of enzymes as a result of the attachment of regulatory proteins;

2. a change in the catalytic activity of enzymes due to the association or dissociation of enzyme protomers.

Regulation of the catalytic activity of enzymes by phosphorylation/dephosphorylation.

In biological systems, there is often a mechanism for regulating the activity of enzymes with the help of covalent modification of amino acid residues. A fast and widespread method of chemical modification of enzymes is phosphorylation/dephosphorylation. The OH groups of the enzyme undergo modifications. Phosphorylation is carried out by protein kinases, and dephosphorylation by phosphoprotein phosphatases. The addition of a phosphoric acid residue leads to a change in the conformation of the active center and its catalytic activity. In this case, the result can be twofold: some enzymes are activated during phosphorylation, while others, on the contrary, become less active.

Regulation of the catalytic activity of enzymes by partial (limited) proteolysis.

Plasma enzymes

By origin, plasma enzymes can be divided into 3 groups.

2. Excretory enzymes - enter the blood from various secrets - duodenal juice, saliva, etc. These include amylase, lipase.

3. Cellular enzymes - enter the blood when cells or tissues are damaged or destroyed.

Table 4.1. Organ-specific enzymes (isoenzymes)

Enzymopathies

At the heart of many diseases are violations of the functioning of enzymes in the cell - enzymopathies. Acquired enzymopathies, as well as proteinopathies in general, seem to be observed in all diseases.

In primary enzymopathies, defective enzymes are inherited mainly in an autosomal recessive manner. Heterozygotes, most often, do not have phenotypic abnormalities. Primary enzymopathies are usually referred to as metabolic diseases, as there is a violation of certain metabolic pathways. In this case, the development of the disease can proceed according to one of the following "scenarios". Consider the conditional scheme of the metabolic pathway:

E 1 E 2 E 3 E 4

A → B → C → D → P

Substance A, as a result of successive enzymatic reactions, turns into product P. With hereditary deficiency of any enzyme, for example, enzyme E 3, various metabolic pathway disorders are possible:

Violation of the formation of end products.

The lack of the end product of this metabolic pathway (in the absence of alternative pathways of synthesis) can lead to the development of clinical symptoms characteristic of this disease.

Clinical manifestations. An example is albinism. With albinism, the synthesis of melanin pigments in melanocytes is impaired. Melanin is found in the skin, hair, iris, retinal pigment epithelium and affects their color. With albinism, weak skin pigmentation, blond hair, a reddish color of the iris due to translucent capillaries are observed. The manifestation of albinism is associated with a deficiency of the enzyme tyrosine hydroxylase (tyrosinase) - one of the enzymes that catalyzes the metabolic pathway for the formation of melanins.

Accumulation of precursor substrates.

If the enzyme is deficient, certain substances will accumulate, and in many cases also the compounds that precede them. An increase in precursor substrates of a defective enzyme is a leading link in the development of many diseases.

Clinical manifestations. A known disease is alkaptonuria, in which the oxidation of homogentisic acid in tissues is impaired (homogentisic acid is an intermediate metabolite of tyrosine catabolism). In such patients, deficiency of the homogentisic acid oxidation enzyme, homogentisic acid dioxygenase, is observed, leading to the development of the disease. As a result, the concentration of homogentisic acid and its excretion in the urine increase. In the presence of oxygen, homogentisic acid turns into a black compound - alkaptone. Therefore, the urine of such patients in the air turns black. Alkapton is also formed in biological fluids, settling in tissues, skin, tendons, and joints. With significant deposits of alkapton in the joints, their mobility is impaired.

Violation of the formation of end products and the accumulation of precursor substrates.

Diseases are noted when both the lack of the product and the accumulation of the initial substrate cause clinical manifestations.

Clinical manifestations. For example, people with Gierke's disease (type I glycogenosis) have a decrease in blood glucose concentration (hypoglycemia) between meals. This is due to a violation of the breakdown of glycogen in the liver due to a defect in the enzyme glucose-6-phosphatase. At the same time, in such people, the size of the liver (hepatomegaly) increases due to the accumulation of unused glycogen in it.

The use of enzymes in medicine

Enzyme preparations are widely used in medicine. Enzymes in medical practice are used as diagnostic (enzymodiagnostics) and therapeutic (enzyme therapy) agents.

In addition, enzymes are used as specific reagents for the determination of a number of substances. So, glucose oxidase is used to quantify glucose in urine and blood. The enzyme urease is used to determine the amount of urea in the blood and urine. With the help of various dehydrogenases, the corresponding substrates are detected, for example, pyruvate, lactate, ethyl alcohol, etc.

Enzymodiagnostics

Enzymodiagnostics consists in making a diagnosis of a disease (or syndrome) based on determining the activity of enzymes in human biological fluids.

The principles of enzymodiagnostics are based on the following positions:

1. when cells are damaged in the blood or other biological fluids (for example, in urine), the concentration of intracellular enzymes of damaged cells increases;

2. the amount of released enzyme is sufficient for its detection;

3. the activity of enzymes in biological fluids detected when cells are damaged is stable for a sufficiently long time and differs from normal values;

4. a number of enzymes have a predominant or absolute localization in certain organs (organ specificity);

5. there are differences in the intracellular localization of a number of enzymes.

The use of enzymes as medicines

The use of enzymes as therapeutic agents has many limitations due to their high immunogenicity.

Nevertheless, enzyme therapy is being actively developed in the following areas:

1. replacement therapy - the use of enzymes in case of their insufficiency;

2. elements of complex therapy - the use of enzymes in combination with other therapy.

Enzyme replacement therapy is effective in gastrointestinal diseases associated with insufficient secretion of digestive juices. For example, pepsin is used for achilia, hypo- and anacid gastritis. Deficiency of pancreatic enzymes can also be largely compensated by oral administration of drugs containing the main pancreatic enzymes (festal, enzistal, mezim-forte, etc.).

As additional therapeutic agents, enzymes are used in a number of diseases. Proteolytic enzymes (trypsin, chymotrypsin) are used locally to treat purulent wounds in order to break down the proteins of dead cells, to remove blood clots or viscous secrets in inflammatory diseases of the respiratory tract. Enzyme preparations have become widely used in thrombosis and thromboembolism. For this purpose, preparations of fibrinolysin, streptolyase, streptodecase, urokinase are used.

The enzyme hyaluronidase (lidase), which catalyzes the breakdown of hyaluronic acid, is used subcutaneously and intramuscularly to dissolve scars after burns and operations (hyaluronic acid forms crosslinks in the connective tissue).

Enzyme preparations are used in oncological diseases. Asparaginase, which catalyzes the reaction of asparagine catabolism, has found application in the treatment of leukemia.

The prerequisite for the anti-leukemic action of asparaginase was the detection in leukemic cells of a defective enzyme asparagine synthetase, which catalyzes the reaction of asparagine synthesis.

Leukemic cells cannot synthesize asparagine and receive it from blood plasma. If the asparagine present in the plasma is destroyed by the administration of asparaginase, then asparagine deficiency will occur in leukemia cells and, as a result, a violation of the cell's metabolism and a stop in the progression of the disease.

Immobilized enzymes are enzymes bound to a solid carrier or placed in a polymer capsule.

Two main approaches are used to immobilize enzymes:

1. Chemical modification of the enzyme.

2. Physical isolation of the enzyme in an inert material.

Enzymes are often immobilized in capsules made of lipids - liposomes, which easily pass through membranes and exert the necessary effects inside the cell.

Advantages of immobilized enzymes:

1. Easily separated from the reaction medium, which allows the enzyme to be reused. The product is not contaminated with enzyme.

2. The enzymatic process can be carried out continuously.

3. Increases the stability of the enzyme.

Immobilized enzymes can be used for analytical and preparative purposes. There are several types of devices where immobilized enzymes are used for analytical purposes - enzyme electrodes, automatic analyzers, test systems, etc.

Preparative use of immobilized enzymes in industry:

1. Obtaining L-amino acids using aminoacylase.

2. Preparation of high fructose syrups using glucose isomerase.

The body is a very complex system, and all processes in it are normally interconnected, without unnecessary reactions and waste. But since the body is not a closed system, and is constantly experiencing external influences, regulation mechanisms are needed that would adapt it to these changes.

Since all processes in our body are controlled by enzymes (hormones act through the enzyme), when conditions change to meet these conditions, the activity and amount of enzymes will change.

1 level. Changes in activity with a change in temperature, amount of substrate, pH of the medium, tk. under these conditions, the mobility of the molecule changes, the ionization of functional groups, and, consequently, the activity of the enzyme.

2 level. The influence of activators and inhibitors on the work of the enzyme (its quantity does not change, the conformation changes) through allosteric. and sometimes the active center.

3 level. Induction and repression of E synthesis, i.e. its quantity changes.

4 level - organismal (neuroregulation). There is a regulation of the synthesis of enzymes involved in the processes of normalization of homeostasis. 4.1. hormonal - some hormones affect the release of others (release factors: statins, liberins, and then - tropic hormones). 4.2. regulation of hormone production by the type of feedback (almost always negative). 4.3. regulation involving CNS structures. 4.4. self-regulation, depends on the parameters of homeostasis. (Parathyroid gland with

a decrease in Ca in the blood increases the production of parathyroid hormone).

Regulation of enzyme activity.

1. Partial proteolysis - activator

From an inactive enzyme

active is formed. peptide

This ensures the appearance

active enzyme at the right time

and in the right place (digestive enzymes; enzymes involved in blood clotting).

2. Protein - protein C R

Interactions in the form of C R + 4cAMP 2 R 4cAMP + 2 C

accession or

regulatory cleavage is inactive. PC active

subunits or regulators. There is a binding to AMP with regulatory.

subunit (R) and thereby release

catalytic subunit that performs

protein phosphorylation.

3. Phosphorylation and

Dephosphorylation - ATP ADR

basic mechanism of protein kinase

Speed control protein FP

Protein phosphatase

The introduction of a "-" charged phosphorus group leads to reversible changes in conformation, and to a change in the activity of the enzyme (glycogen synthase, tissue lipase).

4. Allosteric:

*activator interacts

with allosteric center a

the conformation changes.

Improved binding of S to E

and reaction speed. phosphofructokinase is inhibited by ATP

* Inhibitor interacts with isocitrate DG and is inhibited by ATP,

with E inhibition occurs +

reactions resulting from NADH H

Regulation of the action of enzymes. How is enzyme activity regulated? The concept of the active center of the enzyme

Regulation of enzyme activity. Medical enzymology (biochemistry)

Enzyme regulation

Change in catalytic efficiency of enzymes

Effect of Activators and Inhibitors on Enzyme Activity

irreversible inhibition

Irreversible enzyme inhibitors as drugs

Reversible inhibition

Competitive inhibition

Drugs as competitive inhibitors

Antimetabolites as drugs

Noncompetitive inhibition

Allosteric regulation

Regulation of the catalytic activity of enzymes by protein-protein interactions.

Regulation of the catalytic activity of enzymes by phosphorylation/dephosphorylation.

Regulation of the catalytic activity of enzymes by partial (limited) proteolysis.

Plasma enzymes

Table 4.1. Organ-specific enzymes (isoenzymes)

Enzymopathies

E 1 E 2 E 3 E 4

A → B → C → D → P

Violation of the formation of end products.

Accumulation of precursor substrates.

Violation of the formation of end products and the accumulation of precursor substrates.

The use of enzymes in medicine

Enzymodiagnostics

The use of enzymes as medicines

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