' k 1 [ E ][ S ]
Km
k1 [ ES ]
[ E ][ I ] [E0 ] [E] [ES ] [EI ]
KI
[ EI ]
Inhibited Enzyme Kinetics
Competitive inhibitors (I),
we can obtain,
Vm [ S ]
v
' (1 [ I ] / K ) [ S ]
Km I
Vm [ S ]
v
'
Km , app [ S ]
' ' (1 [ I ] / K ) ' '
Km , app K m I When [I] =0, Km , app K m
Vm Remains that of Michaelis-Menten equation.
K m,app is an indication of the net effect of competitive
inhibition, ????? .
Inhibited Enzyme Kinetics
Noncompetitive inhibitors (I)
- not substrate analogs
- bind on site other than active sites
Km
k2
E+S ES P E
+ +
I I
KI KI
Km
EI+S ESI
Inhibited Enzyme Kinetics
Noncompetitive inhibitors (I)
Assume:
- rapid equilibrium
- same equilibrium constants of inhibitor
binding to E and ES KI
- same equilibrium constants of substrate
binding to E and EI Km
Inhibited Enzyme Kinetics
Noncompetitive inhibitors (I)
d [ P]
v k [ ES ]
dt 2
' k 1 [ E ][ S ] [ EI ][ S ]
Km
k1 [ ES ] [ ESI ]
[ E ][ I ] [ ES ][ I ]
KI
[ EI ] [ ESI ]
' k 1 [ E ][ S ]
Km
k1 [ ES ]
[ ES ][ I ]
KI
[ ESI ]
' k 1 [ E ][ S ]
Km
k1 [ ES ]
[ ES ][ S ]
K SI
[ ES 2]
Vm [S ]
dv / d [S ] d ( ) / d[S ] 0
' [ S ] [ S ]2 / K
Km SI
' K )1/ 2
[S ]max ( K m SI
Uncompetitive substrate inhibitors (I)
Determine [S]max:
Vm [ S ]
dv / d [ S ] d ( ) / d[S ] 0
' [ S ] [ S ]2 / K
Km SI
Vm [ S ](1 2[ S ] / K SI )
Vm
( )0
' [ S ] [ S ]2 / K
Km ( K ' [ S ] [ S ]2 / K ) 2
SI m SI
' [ S ] [ S ]2 / K ) V [ S ](1 2[ S ] / K )
Vm ( K m SI m SI
( )0
(K m' [ S ] [ S ]2 / K ) 2
SI
' V [ S ]2 / K
Vm K m m SI
)0
' [ S ] [ S ]2 / K ) 2
(K m SI
' V [ S ]2 / K 0
Vm K m m SI
' K )1 / 2
[ S ]max ( K m SI
Inhibition Estimation
Product formation rate v ~ [S]: v has a peak?
If yes, then its substrate inhibition.
- get [S]max from the plot of v~[s].
- at low substrate concentration, obtain Vm and Km
graphically or through direct calculation.
- calculate KI through
' K )1/ 2
[S ]max ( K m SI