Regulation of enzyme activity

• The most important factors for enzyme regulation.• Inhibitors• Types of regulatory enzymes.• Allosteric enzymes, their kinetics and allosteric

regulation.• Covalent modification of enzymes.• Mechanism of reversible phosphorylation• Isoenzymes (isozymes)• Cofactors and coenzymes.

Factors, that contribute to regulation of enzyme activity

• pH• Changing amount of enzyme (synthesis of

new molecules of an enzyme/ activation of zymogens)

• Inhibitors and activators (their effect is exerted through regulatory enzymes)

Inhibition of enzymes

Irreversible inhibition

Reversible inhibition

Competitive inhibition

Recognition of competitive inhibition

Noncompetitive inhibition

Recognition of noncompetitive inhibition

Regulatory enzymes

• These are the enzymes (2 large groups) whose activity can be changed in response to cell needs.

• Allosteric enzymes.• Enzymes regulated by reversible covalent

modification (reversible phosphorylation, reversible acetylation and so on).

• There are enzymes which have properties of both groups.

Allosteric enzymes• Contain two different sites:

the active site for catalysisallosteric site for regulation.

• The sites are located in distinct places of enzyme molecule.

• Molecules that affect activity of an allosteric enzyme bind to the allosteric site. These molecules are called allosteric effectors.

Monomericenzyme

Multimericenzyme

Kinetics of allosteric enzymes. Effects of allosteric effectors

• Cooperative effect in multimeric enzymes.

• In a case of allosteric enzyme, the dependence of the reaction velocity agains substrate concentration is S-shaped curve.

• Homotropic allosteric effectors (activators) are substrate molecules.

• Heterotropic allosteric effectors are moluecules different than substrates.

Allosteric effectors

• Allosteric activators/ positive effectors:result in increasing the reaction velocity and in increasing the affinity of an allostericenzyme to its substrate.

• Allosteric inhibitors/ negative effectors:result in decreasing the reaction velocity and in decreasing the affinity of an allosteric enzyme to its substrate

Feed-back inhibition

• A sequence of enzymatic reaction with a particular goal is considered as a system/pathway of enzymes.

• Feed-back inhibition is a way of regulation of enzymatic system activity exerted via initial enzymes of the system. Initial enzymes usually are allosteric ones.

Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4

Regulation of enzyme activity by reversible phosphorylation

• Phosphoryl group is added by an enzyme (phosphotransferase /kinase) to OH-group of Ser/Thr in the regulatory site of an enzyme. The regulatory site and the AS are different in both thefunction and the location in enzyme molecule.

• Phosphoryl group is removed from the regulatory site by another enzyme (phosphatase).

• 3-D structure of an enzyme and its active site is being changed inresponse to phosphorylation. Therefore the activity of an enzyme can increase or decrease.

• Phosphorylation and dephosphorylation is swiched on/off in response to hormones (glucagon, epinephrine, insulin).

• Storage of glycogen and fats as well as cell cycle is controlled by enzymes subjected to reversible phosphorylation.

H-O-P=O⎪O-H

O-H⎪

Phosphoryl group

H-O-P=O⎪O-

O-

⎪

Phosphoryl group (ionized)

Regulation of glycogen phosphorylase activity by allosteric mechanism and by reversible

phosphorylation

Regulatory siteRegulatory site

Allosteric site (in blue)

Allosteric site (in blue)

Regulatory site(phosphorylated)

Regulatory site(phosphorylated)

Regulatory site(phosphorylated)

Regulatory site

Isoenzymes

• Different proteins which catalyze the same reaction are called isoenzymes/isozymes.

• Ussually, isoenzymes are multimeric proteins containing two or more different polypeptide chains.

• In isoenzymes, polypeptide chains are products of different genes.

• Isoenzymes which catalyse the same reaction can be located either in different places of one cell or in different tissues . It is said, isoenzymes have tissue specificity.

Maturation of enzymes (irreversible covalent modification)

• Some enzymes are produced as catalytically inactive proteins –zymogens, e.g. digestive enzymes, cellular enzymes of cell-death pathway.

• Activation of those enzymes is step-like breakdown of their primary structure until active enzyme is being formed.

A single step activationzymogen trypsinogen

Isozymes of lactate dehydrogenase (LDH)

Heart type

Muscle type

CH3-CH-COOH

CH3-C-COOH

OH

O

=

Changes in the pattern of LDH isoenzymes in blood serum after myocardial infarction

Cofactors and coenzymes

• Metal ions which activate an enzyme are cofactors. They are present in the AS of an enzyme.

• Cofactors also are organic molecules which do not take part in catalysis but activate an enzyme. They are located in any place of enzyme molecule except the AS.

• Coenzymes are organic molecules which take part in catalysis. They are present in the AS.

• Coenzymes are classified according the type of reaction in which they take part

oxidoreduction, group transfer, isomerisationthe structure

nucleotides, peptides and heterocyclic • A number of coenzymes com from water soluble vitamins.