Enzyme

Kinetics
Definition

 It is the study of rate of enzyme-catalyzed

reactions and the factors affecting these
reaction rates
Thermodynamic properties of the
reaction
 Free energy difference between products
and reactants.

 The energy required to initiate conversion

of reactants to products
Free energy (∆G)
 The thermodynamic driving force that determines
the direction of chemical reaction and the
concentration of reactants and products at
equilibrium

 Standard free energy (∆GO)

Free energy measured under standard conditions
∆GO = - RT ln Keq
 Exergonic reaction
If ∆G is negative: reaction is spontaneous
Urea ammonia + CO2

 Endergonic reaction
If ∆G is positive: energy is required to drive the
reaction
Glucose + ATP Glu-6-p + ADP
Equilibrium constant (Keq)

 Ratio of reaction rate constants

 Equilibrium –
rate of forward reaction = rate of backward reaction
no net change in the concentration of substrates and
products
enzymes do not alter the equilibrium
Kinetic (collision) theory

 Only molecules that collide can react

 For each chemical reaction, there is an energy

barrier that must be overcome in order for a
reaction to occur
Factors affecting the reaction rate

 Temperature
 pH
 Substrate concentration
 Enzyme concentration
 Activators and inhibitors
Temperature

Temp: increases kinetic energy of molecules

increases collision frequency
lowers energy barrier
 Optimum temperature
The temperature at which maximum amount of
substrate is converted to the product per unit time
350 - 400 C

 Q10 (temperature coefficient)

The factor by which the rate of reaction increases
for every 100 C rise in temperature.
Q10 = 2
 pH
a) It alters the ionization of active site and
favors ES complex formation

b) It may influence the separation of

coenzyme from holoenzyme complex

 Optimum pH = 4 – 9
(pepsin pH= 1-2, ALP pH= 9-10)
Enzyme concentration

 The rate of reaction increases proportionately with

the enzyme concentration
Substrate concentration

 Rate of reaction is proportionate to substrate concentration in the

initial phase
Excess substrate

saturation of enzymes

no increase in reaction rates

 Vi = initial velocity (the velocity
measured when very little substrate
has reacted)

 Vmax = maximal velocity (the maximum

reaction rate attainable in presence of
excess substrate)
Order of reaction
 Zero order
reaction rate is independent of substrate
concentration
 First order
reaction rate is proportional to substrate
concentration { v = k (A)}
 Second order
reaction rate is proportional to square of substrate
concentration { v = k (A)2}
Mechanism of enzyme action
 Lowering of activation energy

 Sucrose hydrolysis by H+ = 26,000 cal/mol

Sucrose hydrolysis by sucrase = 9000 cal/mol
 Transition state
transient intermediate state with higher
free energy than substrate / product

A+B–L A- - B- -L A-B + L

 Activation energy
The difference in free energy between the
transition state and the substrate
 Enzymes facilitate the formation of
transition state
 Enzymes stabilizes the transition state
proper alignment of acid-base groups

transfer of protons to / from transition state

positioning of metal ions

stabilizes the developing charges

Transition state analogs

 Compounds that resemble the transition state of

a catalyzed reaction
 Importance-
● act as inhibitors of enzymes
● provide an insight into catalytic mechanisms
● act as antigens which can generate novel
catalysts

eg: Pyrrole 2-carboxylate (transition state analog)

inhibit proline racemase
 Abzymes
catalytic antibodies produced by using
transition state analogs as antigens
Abzymes catalyze
amide bond formation
trans-esterification
decarboxylation
photo-induced cleavage
 Clinical applications
Drug designing: protease inhibitors in HIV
Abzymes: treatment of cocaine addiction