KKHYBER MEDICAL UNIVERSITY

Course Title: Enzymology

Module 2; Lecture 1: Enzymes Kinetics

Presented by

Dr. Muhammad Shahzad

Assistant Prof IBMS - KMU
 Enzymes Kinetics

• Quantitative measurement of the rates of enzyme catalyzed

reactions &

• The systematic study of factors that affect these rates

• Proposed by Adrina Brown in 1902 when studying the rate of

hydrolysis of sucrose by yeast enzyme invertase.

• When sucrose concentration is much higher than enzyme

concentration, the rate of reaction becomes independent of
sucrose concentration
 Brown’s Proposal

• Overall reaction is compose of two elementary reaction in which

(1) The substrate form a complex with enzyme

E+S ES
(2) ES subsequently decomposes into product and enzyme

2
ES E+P
• According to this model, When the substrate concentration
becomes high enough to entirely convert the enzyme to the ES
form, the second step of the reaction becomes rate limiting step
• So basically, we have two reactions going on at the same time
1 2
E+S ES E+P

Rate 1 = k [ E ] [ S ] Rate2 = k [ ES]

Speed = V = d [ P] / d [ t] = Change in concentration of product

per unit time

To increase the rate of reaction

Either we have to increase [ E ] or [S]
where k = Constant
Lets assume that [ E ] is constant

E E E E

Each enzyme can catalyze 10 reactions/second

Maximum speed of reaction “Vmax” = 40 reactions/second

At high [ S ] , the enzymes will be saturated

Even if [ S ] is much higher, there will still be a Vmax

We have made some assumptions

1. Our solutions are behaving ideally

1 2
E +S ES E+P

Rate 1 = k [ E ] [ S ] Rate2 = k [ ES]

2. Our constants are indeed constants

[ E ] is not changing due to protein degradation or synthesis
K = Environmental factor such pH and temperature have no effect

3. Conversion of substrate into product without enzyme is

negligible
 The steady state assumption

• We have two reactions going on at the same time

1 2
E+S ES E+P
-1 -2

Steady state assumption state that formation of ES is

constant

Formation of ES = Loss of ES

Rate1 + Rate -2 = Rate-1 + Rate 2

1 2
E+S ES E+P
-1

Basis of mechaelis Menton equation

 Mechaelis Menton equation

V0 = Initial velocity of the reaction

Vmax = Maximum velocity

[Substrate] 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.
Km indicate how fast a substrate will bind to enzyme to ES (binding affinity)

low Km value indicates a large binding affinity, as the reaction will

approach Vmax more rapidly and vice versa
 Regulation of enzyme activity
Either by change in activity of the pre-existing enzyme or change in
amount of enzyme
A. Changing the activity of the pre-existing enzyme

1. Substrate availability: If the actual concentration of substrate is

lower than Km (kinetic parameter of enzyme), activity of
enzyme is low.

2. Product inhibition: Product resemble the starting substrate and

bind to the active site of enzyme thus lowering the rate of
reaction.
S

ES E +
E P
3. Allosteric regulation
 Feedback loops

Downstream products regulating upstream reaction

An example is phosphofructokinase (PFK)

4. pH effect on enzyme conformation and active site

Changes in pH can alter shape of enzyme of conformation of

active site thus decreasing rate of reaction

5. Covalent modification

• Post translational modification involving covalent bonds

• Reversible (most of the time).

• phosphorylation, acetylation, methylation, sulfation etc

• Thus regulating enzyme activity

 B. Chang in the amount of enzymes

1. Alteration in transcription of enzyme gene

2. Degradation of messenger RNA for the enzymes

3. Postranslational changes