Enzymes Pt 3: Regulation of enzyme activity

Relicardo M. Coloso, Ph. D.College of MedicineCentral Philippine University

Regulate – to control or direct according to a rule, principle or law

Enzyme regulation- is the control of the rate of a reaction catalyzed by an enzyme by some effector (e.g., inhibitors or activators) or by alteration of some condition (e.g., pH or ionic strength).

Five ways by which enzyme activity in the cell can be changed:

1. Enzyme production – synthesis or degradation

2. Compartmentation – different metabolic pathways occur in different cellcompartments

3. Activation and inhibition – by activators or inhibitors, for examplefeed back inhibition by one of products of the reaction

4. Post-translational modification – for example by phosphorylation,methylation, glycosylation

5. Localization to a different environment – from a reducing (cytoplasm) toan oxidizing (periplasm) environment,of high pH to a low pH, or low salinity to high salinity, high to low energy charge

Enzyme Regulation

• Constitutive enzymes– Enzymes needed at the same level all of the time

• Regulated enzymes– Enzymes needed under some conditions but not

others• For example enzymes of the Lac Operon

– Enzymes are made to break down lactose only if lactose is present

2 Types of Regulation

• Regulation of amount of enzyme made– At the level of transcription = is RNA made?

– At the level of translation = is protein made?

– Slower process (minutes)

• Regulation of enzyme activity– After the protein is synthesized

– Posttranslational modification– Very rapid process (seconds or less than a

second)

Mechanisms of Regulation

Cooperativity shown by an allosteric enzyme

Postranslational regulation of activity byfeed back inhibition

• Feedback inhibition• Pathways with many

intermediates

• The end product of a pathway feeds back and inhibits the activity of the first step in a pathway

• If there are enough end products available, more does not need to be made

• By inhibiting the enzymes in the pathway, the end product will not be made

Activation of cAMP-dependent protein kinase (cAPK) by cyclic AMP (b) At low concentrations of cAMP, the enzyme exists as an inactive tetramer composed of two

regulatory (R) and two catalytic (C) subunits. The tetrameric protein is inactive because the pseudosubstrate sequences on the R subunits block the active sites on the C subunits. Binding of cAMP to the regulatory subunits causes release of the active monomeric catalytic subunits. (b) Structure of cAMP. This unusual nucleotide, which acts as a “second messenger” in many intracellular signaling pathways, controls the activity of many proteins.

Copyright © 2000, W. H. Freeman and Company

Allosteric release of catalytic subunits

Allosteric regulation of aspartate transcarbamoylase (ATCase), the initial enzyme in synthesis of pyrimidines This enzyme comprises a pair of trimeric catalytic subunits (orange) connected by three pairs of dimeric regulatory subunits (green). Binding of cytidine triphosphate (CTP; the blue dot) to the regulatory subunits causes a conformational transition from the active R state to the inactive T state. The more open conformation of the R state permits substrate binding. Thus an increase in the concentration of CTP, an end product in the pyrimidine pathway, shuts off ATCase, an example of feedback inhibition.

Copyright © 2000, W. H. Freeman and Company

Allosteric transition betweenactive and inactive states

Cooperativity shown by an allosteric enzyme

•binding of one ligand molecule affects the binding of subsequent ligand molecules

•positive cooperativity, sequential binding is enhanced; in

•negative cooperativity, sequential binding is inhibited.

Binding of ligands

Cyclic phosphorylation and dephosphorylation is a common cellular mechanism for regulating protein activity In this example, the target protein R (orange) is inactive when phosphorylated and active when dephosphorylated; the opposite pattern occurs in some proteins.

Copyright © 2000, W. H. Freeman and Company

Phosphorylation and dephosphorylation

A linear representation of the conversion of chymotrypsinogen into chymotrypsin by the excision of two dipeptides

These reactions yield three separate chains (A, B, and C), which are covalently linked by disulfide bonds (yellow) in the active enzyme. In the folded, native conformation of chymotrypsin, histidine 57, aspartate 102, and serine 195 are located in the active site.

Copyright © 2000, W. H. Freeman and Company

Proteolytic activation