Aulani " Biokimia Enzim" Presentasi 11 Enzyme Phosphorylation Aulanni’am Biochemistry Laboratory...

Aulani " Biokimia Enzim" Presentasi 11

Enzyme Phosphorylation

Aulanni’amBiochemistry LaboratoryChemistry DepartmentBrawijaya University


Enzyme regulation: How to control the activity of a pathway in response to

the needs of the cell or body


Multiple ways of controllingenzyme activity

• Control of enzyme availability

–Amount of enzyme in the cell

•Control of enzyme synthesis

•Control of enzyme degradation

–Location of enzyme in the cell

•On/off membrane

•Inside/outside organelle



• Control of enzyme activity– Allosteric activation or inactivation: interaction of

small molecule with enzyme, but not at active site


“Feedback” control of metabolic pathways is often accomplished by allosteric

mechanisms

A B C D E F G1 2 3 4 5 6

(Excess G)


Carbamoylphosphate

+aspartate

N-carbamoylaspartate

ATCaseCTP

Allosteric modulation of aspartate transcarbamylase, the first unique step in the biosynthesis of

pyrimidines (C, T, U)

ATP


The reaction catalyzed by aspartate transcarbamoylase

carbamoyl phosphateaspartate

N-carbamoyl aspartate

H2PO4-

C

NH2

O OPO 3=

CO-O

CH2

CHCOO -N

H

C

NH2

O


Activity of aspartate transcarbamoylase (and many allosteric enzymes is cooperative with

respect to aspartate

[aspartate]

v0Sigmoid (S-shaped) curve


Cooperativity in enzymatic activity

• Multimeric enzymes

• Binding of S to one protomer causes change in adjacent protomer, increasing the affinity of both protomers for the substrate


Cooperativity of enzymatic activity: multiple states of the enzyme

ELA EMA

(T state)Less activeLower affinity for S

(R state)More activeHigher affinity for S


What good is cooperativity in enzymatic activity?

v0

[S]

(Same Vmax and v0 at 1/2 Vmax)

Steeper slope ensures greater sensitivity to variations in substrate concentration


ATP and CTP are positive and negative modulators of aspartate transcarbamoylase

[substate]

v0

positive

negative

Effects of positive and negative modulators on cooperative enzymes


Allosteric activators and inhibitors stabilize different states of the enzyme

(ATCase)

ELA EMA

(T state)Less activeLower affinity for SStabilized by CTP

(R state)More activeHigher affinity for SStabilized by ATP


Allostery is accomplished by conformational changes in ATCase and other enzymes

Red: upper catalytictrimer

Blue: lower catalytictrimer

Green: regulatorydimers (bind ATP and CTP)


Other about allosterism

• Allosteric modulators can change Km or Vmax or both

• Not all allosteric enzymes are cooperative

• Allosteric modulation renders an enzyme sensitive to metabolites other than its own substrates and products (e.g., ATCase is sensitive to endproducts of purine and pyrimidine biosynthesis).



• Control of enzyme activity

– Allosteric activation or inactivation: interaction of small molecule with enzyme, but not at active site

– Interaction with a large molecule, usually another protein

Example: Thrombin and thrombomodulin


Modification of thrombin activity by thrombomodulin

• Thrombomodulin (on cell surface of vascular endothelium)

– Decreases activity of thrombin toward fibrinogen

– Increases ability of thrombin to activate Protein C, an inhibitor of clotting


Modification of thrombin activity by thrombomodulin

clot

Thrombin

Protein C

>

Thrombomodulin

Protein C zymogen(contains Gla)




– Allosteric activation or inactivation

– Interaction with another protein

– Reversible covalent modification

Example: phosphorylation


Modulation by phosphorylation

Enzyme X(inactive)

PO32-

Enzyme X(active)

Phosphorylation may also result in inactivation,depending on the enzyme

kinaseATP ADP

phosphataseH2OPi




– Allosteric activation or inactivation

– Interaction with another protein

– Reversible covalent modification

– Irreversible modification

Example: zymogen activation


Multiple ways of controlling a single enzyme: Glycogen phosphorylase

• Gene expression– Multiple genes, tissue-specific expression– Rates transcription vary

• Protein activity–Allosteric control–Reversible phosphorylation

Aulani " Biokimia Enzim" Presentasi 11 Enzyme Phosphorylation Aulanni’am Biochemistry Laboratory...

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