ENZYMES

Enzymes biological catalysts of protein

nature, that catalyze metabolic reactions in the body under the mild conditions required to life.

STRUCTURAL ORGANIZATION OF ENZYME’S MOLECULE

Active site is the part of the enzyme that binds with the substrate.

Binding site is the R groups in the active site that are involved in substrate binding.

Catalytic site is the R groups in the active site that are involved in catalysis.

Allosteric site - site to which bind some substances which called as “Effectors”. They can modify structure of enzymes, and change activity of enzymes.

Formation of enzyme-substrate complex

E + S → ES → E + P

Models of enzyme-substrate complex formation

The lock-and key model of Fisher

According to this model, the structure of the enzyme is a rigid. The substrate fits neatly into the active site just as a key into the proper lock. Thus the active site of an enzyme is a rigid and pre-shaped template where only a specific substrate can bind.

Induced fit theory of Koshland

As the enzyme binds to the substrate, the shape of the active site conforms precisely to the shape of the substrate. The interaction of the substrate with the enzyme induces a fit or conformational change in the enzyme, resulting in the formation of a strong substrate binding site. The shape of the substrate may also change.

Specificity of enzyme is the ability of the enzyme to

bind only, or a very few substrates and thus catalyze only a single reaction.

Stereochemical specificity an enzyme that catalyze conversation only one

stereoisomer of the substrate.

Reaction specificityEach reaction is catalyzed by the different

enzyme

Substrate specificity• Absolute specificity.

Relative substrate specificity

Group depended specificity

• Bond (linkage) specificity an enzyme

catalyses the formation or breakage of only certain bonds in a molecule.

TYPES OF ENZYMES

Proenzymes. Some enzymes are synthesize as proenzymes or zymogens which undergo irreversible covalent activation by the breakdown of one or more peptide bonds.

Isoenzymes are the multiple forms of an enzymes catalyzing the same reaction.

Isoenzymes of lactate dehydrogenase (LDH) –

Lactate dehydrogenase (LDH) isoenzymes and their characteristic

Isoenzymes of creatine kinase (CK)

Creatine kinase (CK) isoenzymes and their characteristic

Multienzyme complexes

PDH complexes

Enzymes of PDH complex: 1. Pyruvate dehydrogenase (PDH)

2. Dihydrolipoyl transacetylase dihydrogenase

3. Dihydrolipoyl dеhydrogenase

Coenzymes of PDH

1. TPP (vitamin B1)

2. Lipoamide (lipoic acid)

3. FAD (vitamin B2)

4. Coenzyme A (pantotenic acid)

5. NAD (vitamin PP or niacin).

FACTORS AFFECTING ENZYME ACTIVITY

Concentration of enzyme

As the concentration of the enzyme is increased, the rate of the reaction

proportionately increases.

Effect of substrate concentration.

At first with increasing of substrate concentration, rate of reaction

increases, further rate of reaction gets high level (Vmax) when enzyme

saturated by substrate and not changes.

Effect of substrate concentration to the rate of reaction shows next Michaelis-Menten’s equation:

V - velocity of the reaction.Vmax - is the maximal rate of the reaction.[S] - is the concentration of the substrate.Km - is the Michaelis-Menten constant which shows the concentration of the substrate when the reaction velocity is equal to one half of the maximal velocity for the reaction.

Effect of temperature

At low temperatures, enzymes have low activity.

At temperature around 37oC an enzyme is maximally active, it is

optimum temperature. Higher than this temperature the enzymes are

denatured and activity ceases.

Optimum temperature for most of enzymes is

Effect of pH

This peak – pH=7 is represents the optimum pH for most of enzymes. But some of enzymes actively at

acidic (Pepsin) and alkaline (Arginase) pH

INHIBITORS

• Enzyme inhibitors can bind to enzymes and either eliminate or drastically reduce their catalytic activity.

• Enzyme inhibitors are classified on the basis of the inhibition.

Irreversible inhibitors • The inhibitors bind covalently

with enzyme and inactivate them (usually toxic substances), or destruct native structure of enzymes (chemical and physical factors: acids, temperature, x-rays others).

Reversible inhibitorsReversible, competitive enzyme inhibitors are structural analogs, molecules that look like the structure of the natural substrate. They can bind to enzyme active site, enzyme-inhibitor complex (Ei) formed and reaction will be stopped. If the inhibitor is removed reactions can occur.

E + i Ei

• Reversible, noncompetitive inhibitors.

• These inhibitors don’t bind to the active site. They bind to other part of enzymes, formed enzyme-substrate-inhibitor complex (ESi), the binding is weak and the enzyme activity is restored when the inhibitor dissociates from the enzyme-subs trate- inhibitor complex.

Enzymology

There are three directions of enzymology:

• enzymodiagnosis

• enzymopathology

• enzymotherapy

Enzymopatology Enzymopatology study diseases related with

deficiency and high activity of enzymes.

For example disease fructosuria due to lack of enzymes fructokinase and galactozemia due to deficiency of galactose -1-phosphate uridyl transferase enzyme; gout due to high activity of hypoxanthine-oxidase.

Enzymotherapy is using of enzymes as therapeutic agents

Streptokinase prepared from streptococcus is useful for clearing the blood clots. Streptokinase activates plasma plasminogen to plasmin which, in turn, attacks fibrin to convert into soluble products.

GIT enzymes for treating of GIT diseases.

Diagnostic importance of enzymes

Some enzymes are totally absent or present at a low concentration in plasma compared to their levels found in the tissues.

The raised enzyme levels could be due to cellular damage, increased rate of cell turnover, proliferation of cells, increased synthesis of enzymes. Serum enzymes are conveniently used as markers to detect the cellular damage which ultimately help in the diagnosis of diseases.

Examples: LDH1, and 2, CC2 increase in heart diseases, Amylase is increased in

pancreas diseases etc.

Nomenclature of enzymes – is derived from their substrates or catalyzed

chemical reactions, and “ase” is usially added as suffix.

Ex: Urease, hexokinase, lactate dehydrogenase etc. .

Classification of enzymes According to the types of reactions

that enzymes catalyze, enzymes classified to6 classes:

- Oxidoreductases,- transferases,- hydrolyses,

- lyases,-isomerases,

- ligases.

Oxidoreductases – catalyze oxidation-reduction reactions.

Transferases – group of enzymes catalyze transport of group

between substanses

Hydrolyses – group of enzymes which catalyze removing of group by the acting of water

Lyases – group of enzymes which catalyze removing of group without

acting of water

Isomerase – group of enzymes which catalyze isomerization

reactions.

Ligases – catalyze synthesis of new substance using ATP hydrolysis.

CH3-CO-SCoA

CO2 ATP

COOH-CH2CO-CoAAcetyl-CoA MalonylCoA

ADP+Pi

Faculty for Foreign Citizens the 1st year studentsBiochemistryLesson 3Theme: Structure and properties of enzymes.

Theoretical issues:

1. Definition, chemical nature and nomenclature of enzymes.2. Structure of enzymes: coenzyme, co-factor, apoenzyme, substrate.3. Active site of enzymes, it’s sites and functions. Allosteric center.4. Mechanism of action of enzymes.5. Main properties of enzymes: thermolability, dependence of activity of enzymes from pH.6. Dependence of rate of enzymatic reaction from substrate concentration. Michaelis-Menten equation.7. Specificity of enzymes. Types of specificity, examples.8. Isoenzymes and multienzyme systems, their localization in various organs:a) lactate degydrogenase (LDH);b) transaminases: alanin-aminotransferase (ALT), aspartate aminotransferase (AST);c) creatine kinase: CK1, CK2, CK3.

Faculty for Foreign Citizens the 1st year studentsBiochemistry

Lesson 4Theme: Influence of activators and inhibitors on enzyme activity. Organ-specific enzymes. Classification of Enzymes

THEORETICAL ISSUES:9. Activators of enzymes, mechanism of their action.10. Inhibitors of enzymes: reversible and irreversible, specific and non-specific.11. Competitive and non-competitive types of inhibition, mechanism of action, examples.12. Nomenclature and classification of enzymes, examples of reactions.14. Medical applications of enzymes: enzymopathies, enzymodiagnostics, and enzymotherapy.15. Control work on enzymes.