Enzyme inhibition.ppt

enyme

1. Inhibitionofenzymeactivity
Somesubstancesreduceorevenstopthecatalyticactivityof
enzymesinbiochemicalreactions.Theyblockordistorttheactive
site.Thesechemicalsarecalledinhibitors,becausetheyinhibit
reaction.
Inhibitorsthatoccupytheactivesiteandpreventasubstrate
moleculefrombindingtotheenzymearesaidtobeactivesite-
directed(orcompetitive,asthey'compete'withthesubstrateforthe
activesite).
Inhibitorsthatattachtootherpartsoftheenzymemolecule,perhaps
distortingitsshape,aresaidtobenon-activesite-directed(ornon
competitive).
1

2. i) inhibitor competes with substrate for the enzyme active site
ii) non-covalent interactions
iii) structures similar to substrate, products or transition state
Reaction scheme for a reversible enzyme inhibitor and the definition of the
dissociation/inhibition constant.
Competitive inhibitors (Michaelis-Menton kinetics)
2

3. P + E
S
+
E
S
E
I S
E
I
SS
S
I
S
E
X
I
I
S
Competitive Inhibition
P + E
FormationoftheE-Icomplexprevents
bindingofsubstrate,thereforethereaction
cannotproceedtothenormalphysiological
product,P.
High[S]overcome
inhibition
3

4. FormationoftheE-Icomplexpreventsbindingofsubstrate,
thereforethereactioncannotproceedtothenormalphysiological
product,P.
Larger[I]leadstolarger[E-I],leavinglessfreeenzymetowhich
thesubstratecanbind.
Limitation:mustmaintainhigh[I]tomaintaininhibition,requiring
multipleadministrationsorextendedreleaseformulations.
Notethathigh[S]canovercomeinhibitionbyfavoringequilibrium
totheE-Scomplex.
Competitive inhibitors
4

5. ACompetitiveInhibitorhasachemicalsimilaritytothesubstrateandcompeteswith
thesubstrateforbindingtotheactivesiteoftheenzyme.Agoodexampleto
describecompetitiveinhibitionisthemitochondrialenzyme,succinate
dehydrogenase:
(A)Thereactioncatalyzedbysuccinatedehydrogenaseistheoxidationofsuccinate
tofumarate.(B)Malonateandoxaloacetatearecompetitiveinhibitorsofsuccinate
dehydrogenase.
Boththesecompetitiveinhibitors,malonateandoxaloacetate,looklikesuccinatein
theirchemicalcharacter.Bothinhibitorscanbindinthesameplacesintheactive
siteofsuccinatedehydrogenaseasthesubstrate.However,neitherinhibitorhasthe
capacitytoundergothereactionandsotheenzymeisinhibited.Thehallmarkof
competitiveinhibitionisthatitcanbeovercomebyasufficientlyhighconcentration
ofsubstrate.
C
CH
2
CH
2
O
-
O
O
-
OC
2H
-
Succinate
dehydrogenase
C
CH
C
O
-
O
O
-
OC
H
C
C O
CH
2
O
-
O
O
-
OC
C
CH
2
O
-
O
O
-
OC
A B
5

6. 6

7. 7

8. Competitiveinhibitionoccurswhenbothsubstrateand
inhibitorwillfittheactivesite,andcompetewitheach
othertooccpyit.
y = mx + c
1
[S]
1
V
V
maxunaltered
K
mincreased
E
I
E-I P + EX
E-S P + E
s
8

9. 9

10. Non-competitive inhibition
In this case of the non-competitive inhibition, the inhibitor
reacts with the enzyme at a site other than the active site,
causing conformational change in enzyme which decreases or
stops catalytic activity.
Both the free enzyme (E) and the enzyme-substrate complex
(E-S) react with inhibitor.
Non-competitive inhibition is irreversible and the substrate can
not over come the inhibitors impact on the enzyme
E
S
E
S
P
E
+
E
S
E
S
E
+
I
EI
ES
ESI
10

11. 11

12. Non-competitiveinhibitionoccurswhentheinhibitorbinds
toasiteontheenzymeotherthantheactivesiteorbinds
irreversiblytotheactivesite.
y = mx + c
1
[S]
1
V
V
maxreduced
K
munaltered
1
K
m
12

13. Non-competitiveinhibitionoccurswhentheinhibitorbinds
toasiteontheenzymeotherthantheactivesiteorbinds
irreversiblytotheactivesite.
E ES
EI ESI
E + P
S
S
I II I
k
i
k’
i
13
TwoK
i
'shavethesamevalue,i.e.the
inhibitorbindsequallywelltoEandES,
thenamixedinhibitorisusuallycalleda
noncompetitiveinhibitor.

14. 14

15. Mixed Inhibitor.
y = mx + c
1
[S]
1
V
+ I
1
K
m
1
K
m
app
1
V
max
1
V
maxapp
E ES
EI ESI
E + P
S
S
I II I
k
i
k’
i
A mixed inhibitor binds to both E and ES,
not at the substrate binding site:
Note that there are 2 K
i
's. If they have the same
value, i.e. the inhibitor binds equaly well to E and ES,
then a mixed inhibitor is usually called a
noncompetitive inhibitor.
In the presence of a mixed inhibitor, the
following Lineweaver-Burk plot is obtained:
Both the apparent Km and the apparent Vmax
are changed by the inhibitor
k
ik’
i
15

16. Uncompetitiveinhibitionoccurswhentheinhibitorbindsafter
thesubstratehasboundtotheenzyme,andthenstopsthe
reactionoccurring.
Uncompetitive inhibition
occurs when the inhibitor
binds only to the ES
complex:
S+
E
E
S
P+
E
E
S
I
16