BBCCT-107
ENZYMES
Indira Gandhi National
Open University
School of Sciences
Block
2
ENZYME KINETICS
UNIT 4
Enzyme Kinetics 45
UNIT 5
Bisubstrate Reactions 57
UNIT 6
Enzyme Inhibition 69
, Block 2 : Enzyme Knetics
How enzyme catalytic activity is controlled? Enzyme kinetics deals with the
measurement of rate of reaction. Our understanding about the kinetics of
an enzyme will help to understand the functional information about its
catalytic mechanism, role of enzyme in metabolism and the factors that
impact its activity as well as the mechanisms of inhibition.
This block on enzyme kinetics studies—among many things will help the
learner to determine how the rate of enzyme-catalyzed reactions depends
on the concentration of the compounds directly interacting with the enzyme
and what is the highest rate achievable by the enzyme? Further questions
of interest include the catalytic mechanism in presence of two or more
substrates. How the rate of the catalyzed reaction is affected in presence of
different types of inhibitor? All related studies provide pieces of information
that serve as input data to establish a mechanistic model of the enzymatic
reaction.
Unit 4 of the block discuss the basis of the derivation of Michelis-Menten
equation for mono substrate enzyme catalyzed reaction. Since most of the
biochemical reactions involve two or more substrates, Unit 5 dwells on
bisubstrate enzyme catalyzed reactions. Enzyme activity needs to be firmly
regulated to guarantee that levels of the product do not increase to
undesired levels. This is accomplished by enzyme inhibition. Unit 6 of the
block discusses different types of enzyme inhibition-reversible and
irreversible.
Objectives:
After studying this block, you should be able to:
determine the rate of enzyme catalyzed reaction,
illustrate bisubstrate enzyme catalyzed reactions,
explain the mechanism of enzyme catalysis,
describe different types of inhibitors regulating enzyme activity.
, 4
Unit 4 Enzyme Kinetics
..........................................................................................................................................................................
UNIT
ENZYME KINETICS
Structure
4.1 Introduction 4.3 Summary
Objectives 4.4 Terminal Questions
4.2 Enzyme Kinetics, 4.5 Answers
Michaelis-Menten Equation 4.6 Suggested Readings
Lineweaver-Burk Plot
Significance of Km and Vmax
kcat and Turnover number
4.1 INTRODUCTION
The oldest and the important approach to understand enzyme mechanisms is
the discipline known as enzyme kinetics. Kinetics is the study of reaction
rates, their quantitative measurement and a systematic study of the factors
influencing the activity of enzymes. In this unit, you will gain an insight of the
enzymatic mechanisms as well as role played by enzyme activity in regulating
metabolic pathways. Enzymes convert substrates to products through a
series of steps known as enzymatic mechanism. Therefore the effect of
substrate concentration on enzyme activity is one of key concepts in enzyme
kinetics. Several models have been proposed to explain the kinetics of enzyme
catalyzed reactions. Classical experimental work for single enzyme catalysed
reactions is Henri-Michaelis-Menten plot, Briggs Haldane equation,
Lineweaver-Burk plot, etc.
Objectives
After studying this unit, you should be able to:
derive Michaelis-Menten equation;
explain the mechanisms of enzyme catalysis;
draw Lineweaver-Burk plot; and
describe Km and Vmax. 45
, Block 2 Enzyme Kinetics
..........................................................................................................................................................................
4.2 ENZYME KINETICS
Kinetic analysis helps to disclose the number and order of the individual steps
involved in the transformation of substrates to products. In the past, data
generated from the experiments of enzyme catalyzed reactions was collected
and analyzed to determine the rate of a reaction. It was found out that at low
concentrations of substrate, the reaction was of first-order with respect to the
substrate. However, at the higher concentrations of substrate, the reaction
became zero-order. Please recall from your chemistry books regarding the
zero order or first order enzyme catalyzed reactions. Generally all single
substrate enzyme catalyzed reactions and even multi-substrate reactions
where concentrations of all but one were kept constant follows the same order.
At constant enzyme concentration, graph of initial velocity [vo] (on y-axis)
against substrate [S] concentration (on x-axis) exhibit a hyperbolic curve
(Fig. 4.1).
Vmax
V0
zero-order
reaction
first-order
reaction
[S0]
Fig.4.1: Graph of initial velocity against substrate concentration for a single
substrate enzyme catalyzed reaction.
The general equation from the graph is
V max [ So ]
vo
[ So ] b
Vmax = Maximum velocity = maximum value of vo
b = constant = value of [ So ] where vo = ½ Vmax
In general terms, in a mono substrate enzyme catalyzed reaction and
considering just one substrate binding site per enzyme molecule, substrate [S]
comes in physical contact with enzyme [E] to form an enzyme substrate
complex [ES] complex which eventually undergoes a further reaction and leads
46 to the formation of product [P]. It can be represented as:
ENZYMES
Indira Gandhi National
Open University
School of Sciences
Block
2
ENZYME KINETICS
UNIT 4
Enzyme Kinetics 45
UNIT 5
Bisubstrate Reactions 57
UNIT 6
Enzyme Inhibition 69
, Block 2 : Enzyme Knetics
How enzyme catalytic activity is controlled? Enzyme kinetics deals with the
measurement of rate of reaction. Our understanding about the kinetics of
an enzyme will help to understand the functional information about its
catalytic mechanism, role of enzyme in metabolism and the factors that
impact its activity as well as the mechanisms of inhibition.
This block on enzyme kinetics studies—among many things will help the
learner to determine how the rate of enzyme-catalyzed reactions depends
on the concentration of the compounds directly interacting with the enzyme
and what is the highest rate achievable by the enzyme? Further questions
of interest include the catalytic mechanism in presence of two or more
substrates. How the rate of the catalyzed reaction is affected in presence of
different types of inhibitor? All related studies provide pieces of information
that serve as input data to establish a mechanistic model of the enzymatic
reaction.
Unit 4 of the block discuss the basis of the derivation of Michelis-Menten
equation for mono substrate enzyme catalyzed reaction. Since most of the
biochemical reactions involve two or more substrates, Unit 5 dwells on
bisubstrate enzyme catalyzed reactions. Enzyme activity needs to be firmly
regulated to guarantee that levels of the product do not increase to
undesired levels. This is accomplished by enzyme inhibition. Unit 6 of the
block discusses different types of enzyme inhibition-reversible and
irreversible.
Objectives:
After studying this block, you should be able to:
determine the rate of enzyme catalyzed reaction,
illustrate bisubstrate enzyme catalyzed reactions,
explain the mechanism of enzyme catalysis,
describe different types of inhibitors regulating enzyme activity.
, 4
Unit 4 Enzyme Kinetics
..........................................................................................................................................................................
UNIT
ENZYME KINETICS
Structure
4.1 Introduction 4.3 Summary
Objectives 4.4 Terminal Questions
4.2 Enzyme Kinetics, 4.5 Answers
Michaelis-Menten Equation 4.6 Suggested Readings
Lineweaver-Burk Plot
Significance of Km and Vmax
kcat and Turnover number
4.1 INTRODUCTION
The oldest and the important approach to understand enzyme mechanisms is
the discipline known as enzyme kinetics. Kinetics is the study of reaction
rates, their quantitative measurement and a systematic study of the factors
influencing the activity of enzymes. In this unit, you will gain an insight of the
enzymatic mechanisms as well as role played by enzyme activity in regulating
metabolic pathways. Enzymes convert substrates to products through a
series of steps known as enzymatic mechanism. Therefore the effect of
substrate concentration on enzyme activity is one of key concepts in enzyme
kinetics. Several models have been proposed to explain the kinetics of enzyme
catalyzed reactions. Classical experimental work for single enzyme catalysed
reactions is Henri-Michaelis-Menten plot, Briggs Haldane equation,
Lineweaver-Burk plot, etc.
Objectives
After studying this unit, you should be able to:
derive Michaelis-Menten equation;
explain the mechanisms of enzyme catalysis;
draw Lineweaver-Burk plot; and
describe Km and Vmax. 45
, Block 2 Enzyme Kinetics
..........................................................................................................................................................................
4.2 ENZYME KINETICS
Kinetic analysis helps to disclose the number and order of the individual steps
involved in the transformation of substrates to products. In the past, data
generated from the experiments of enzyme catalyzed reactions was collected
and analyzed to determine the rate of a reaction. It was found out that at low
concentrations of substrate, the reaction was of first-order with respect to the
substrate. However, at the higher concentrations of substrate, the reaction
became zero-order. Please recall from your chemistry books regarding the
zero order or first order enzyme catalyzed reactions. Generally all single
substrate enzyme catalyzed reactions and even multi-substrate reactions
where concentrations of all but one were kept constant follows the same order.
At constant enzyme concentration, graph of initial velocity [vo] (on y-axis)
against substrate [S] concentration (on x-axis) exhibit a hyperbolic curve
(Fig. 4.1).
Vmax
V0
zero-order
reaction
first-order
reaction
[S0]
Fig.4.1: Graph of initial velocity against substrate concentration for a single
substrate enzyme catalyzed reaction.
The general equation from the graph is
V max [ So ]
vo
[ So ] b
Vmax = Maximum velocity = maximum value of vo
b = constant = value of [ So ] where vo = ½ Vmax
In general terms, in a mono substrate enzyme catalyzed reaction and
considering just one substrate binding site per enzyme molecule, substrate [S]
comes in physical contact with enzyme [E] to form an enzyme substrate
complex [ES] complex which eventually undergoes a further reaction and leads
46 to the formation of product [P]. It can be represented as:
