Lecture 8-KumarRegulation of Enzyme Activity

• Regulation by modification

– Proteolytic cleavage

– Covalent modification

– Protein-protein interaction

• Allosteric regulation

– Properties of allosteric enzymes (important)

– Sigmoid kinetics (what does Km mean in this case) (important)

– Positive and negative modulators (where do they act and how do they modify activity at constant substrate concentration) (most important)

– Models of allosteric transitions (important)

Regulation of Enzyme Activity

Normal metabolic control may be exerted in a variety of ways. Examples are:1.Proteolytic Cleavage of inactive Proenzymes to active enzymes

Pepsinogen pepsin + small peptidein gastrointestinal tract for protein digestion

2.Coagulation cascade—a series of proenzymes are converted to active enzymes. The last step is

Fibrinogen Fibrin

Coagulation Cascade

Covalent Modification as Control

Chemical modification can either increase or decrease activity. Some examples are: Glycogen Synthetase Phosphatase Kinase Phosphorylated Glycogen synthetase

Glycogen phosphorylase

Covalent Phosphorylated Glycogen phosphorylase

Protein-Protein Interaction

Example is activation of Protein Kinase A

R2C2 + 4 cAMP R2C2(4cAMP)

Inactive 2R(cAMP)2 + 2C (active)

The catalytic unit (C) is able to phosphorylate and modulate the activity of other enzymes

Allosteric Regulation

Properties of Allosteric enzymes

1. Catalyze essentially irreversible reactions; are rate limiting

2. Generally contain more than one polypeptide chain

3. Do not follow Michaelis-Menten Kinetics

4. Are regulated by allosteric activators or inhibitors

5. Can be up-regulated by allosteric activators at constant [S]

6. Can be down regulated by allosteric inhibitors at constant [S]

7. Activators and Inhibitors need not have any structural resemblance to substrate structure

Sigmoid kinetics for allosteric enzymes

Effects of allosteric activators and allosteric inhibitors on enzyme activity

Effect of allosteric activators and inhibitors on rate at cellular concentration of the

substrate

Models of Allosteric ModulationSymmetry model

Sigm

oidal Curve E

ffect

Sigmoidal curve

Exaggeration of sigmoidal curveyields a drastic zigzag line that shows the On/Off point clearly

Positive effector (ATP)brings sigmoidal curveback to hyperbolic

Negative effector (CTP)keeps

Consequently, Allosteric enzyme can sense the concentration of the environment and adjust its activity

Noncooperative(Hyperbolic)

Cooperative(Sigmoidal)

CTPATP

vo

vo

[Substrate]Off On

Learning objectives for lecture 8

• Learn various methodologies that enzymes employ to control metabolism

• Know the properties of an allosteric enzyme.

• Understand the significance of sigmoid kinetics. Can one determine Km and Vmax for these enzymes from the sigmoid plot

• Understand how the activity of an allosteric enzyme is regulated by allosteric activators and inhibitors

• Understand the mechanism of allosterism and negative feedback inhibition