Enzyme kinetics

-- Michaelis Menten kinetics

Two approaches:
1. Rapid equilibrium approach
2. Quasi steady state approach

Assumptions:

Total enzyme concentration remains constant

during the reaction
Amount of enzyme is very small compared to
amount of substrate
The product concentration is so low that the
product inhibition is negligible.

E S ES E P

k
k1

k3

where
k1= forward rate constant for formation of ES
complex
k2= backward rate constant for formation of
ES complex
k3= rate constant for formation of product P

some notations..
e = concentration of enzyme
s = concentration of substrate
p = concentration of product
(es) = concentration of enzyme substrate
complex
t = time
v = reaction rate or velocity

Michaelis Menten kinetics

--Rapid equilibrium approach
It is assumed that ES complex is established
very rapidly (since this equilibrium step is only
the formation of weak interaction between E &
S)
The product releasing step (k3) is very
slow.which determines the rate
The rate of reverse reaction of the second step
is negligible

E S ES

k1

k3

E P

k2

dp
v kv (es)
dt

k
es
2
k

e sK
2
( es )
K

(es )
k
k
1
1

The equilibrium constant

The rate of product formation,

dp
v k (es)
dt
3

(mol/ l.s)
2

The total enzyme concentration

e0 e (es)

Get an expression for (es) in known quantities

Sub. eqn (1) in (3)

(3)

e e (es)
0

K
(es ) (es)
s
K
(es) 1
s

(es )

e
K
1
s

 Now sub. the value of (es) in eqn 2.

ke
(2) v k (es)
K

s
kes

K s ( v k e Maximal velocity
3

max

k
( K K Michaelis cons tan t )
k
2

v s
v
K s
max

Michaelis Menten Equation

 Therefore..

max

is a function of enzyme concentration only

A low value of means

the substrate

that the enzyme has high affinity for

Three special cases..

Case I

(s=KM)

Case II

(s>>KM)

Case III

(s<<KM)

Case I

(s=KM)
Eqn. (5) =>

v s
v
ss
max

v
v
2

max

So when s=KM, the rate of reaction is one half of

its maximal value.
i.e. at which 50% of enzyme active sites are
occupied by substrate

Case II

(s>>KM)
Eqn. (5) =>

v s
K

1
s

max

vv

max

--------- ZERO ORDER

Case III

(s<<KM)
Eqn. (5) =>

v s
v
s

K
1
K

max

v
v
s ---------FIRST ORDER
K
max

Michaelis Menten kinetics

--Quasi steady state approach
This approach is assumed that the change in
the intermediate (transition complex)
concentration with respect to time is negligible.
(pseudo steady state/quasi steady state)--Briggs-Haldane approach
i.e.

d (es )
0
dt

E S ES E P

k3

k1

k2

dp
v k (es )
dt

ds
k es k (es)
dt
1

d (es)
k es k (es) k (es)
dt
1

d (es)
0
dt

By p.s.s assumption,

Eqn 4==>

d (es)
k es (es)(k k ) 0
dt
1

k es (k k )(es)
1

k k
e
(es )

ks

We know,

e e (es)
0

k k
(es) 1

k
s

e
(es )
k k
1
ks
0

eks
(es)
k sk k
0

 sub the value of

(es) in (1)

dp
v k (es)
dt
3

kkes

k sk k
3

kkes
k k
s
k
3

k
1

k k e s
k s k
k
3

kes

k k k

s
k

Michaelis Menten Equation

v s
v
K s

max v

k k e s
k
k
s
2
k

k e s
k

k
k

s
3

v
K

s
max
s

max

ke

 Controversy of Equilibrium approach:

by Equilibrium approach,

eks
(es)
k sk
0

ds
k es k (es)
dt
1

but e e (es )

k [ e (es)]s k (es)
1

k e s k (es) s k (es)
1

k e s (es)[k s k ]
1

kes
k e s
[k s k ]
[k s k ]
1

ds
0
dt

Means that S is costant..which is not

correct

 This has been rectified by PSS approach..

eks
by PSS assumption (es)
0

k sk k
1

ds
k es k (es)
dt
1

k [ e (es)]s k (es)
1

k e s (es )[k s k ]
1

kes
k e s
[k s k ]
k sk k
1

k e s k k e s
k e s
k sk k
2

k e s k k e sk k e sk e s k k e s

k sk k
2

k k e s

k sk k
1

k k e s

k k
k s

v s
ds
dp

dt
K s
dt
m

which is correct and holds good

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