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CHAPTER 1:
ENZYME KINETICS AND
APPLICATIONS
(Part Ib : Kinetics of Enzyme Catalyzed
Reactions)
ERT 317 Biochemical Engineering
Sem 1, 2015/2016
Models for More Complex
Enzyme Kinetics
Allosteric Enzymes
Allostery @ cooperative binding
• The binding of one substrate to enzyme
facilitates binding of other substrate molecules
• Rate expression:
Where,
n = cooperativity coefficient :
n>1 indicates positive cooperativity (activator; n<1=inhibitor)
The cooperativity coefficient can be determined by rearranging above
equation
Graphical Determination of the
Cooperativity Coefficient, n
The binding of one
substrate to enzyme
facilitates binding of other
substrate molecules
Comparison of Michaelis-Menten and Allosteric
Enzyme kinetics
Inhibited Enzyme Kinetics
ENZYME INHIBITORS :
• Certain compound that bind to enzyme and reduce
enzyme activity
• May be IRREVERSIBLE or REVERSIBLE
• IRREVERSIBLE ( like heavy metal)form a stable
complex with enzyme and reduce enzyme activity
• Such enzyme inhibition may be REVERSED only by
using chelating agents such as EDTA and citrate. It is
easily DISSOCIATED from the enzyme after binding.
3 Major Classes REVERSIBLE
Enzyme Inhibition
1. COMPETITIVE
2. NONCOMPETITIVE
3. UNCOMPETITIVE
1. Competitive Inhibition
Substrate analogs and compete with substrate for the active site of the enzyme
Equation for
the rate of
enzymatic
conversion
K’m,app
The effect of such inhibitors can be overcome by
increasing the substrate concentration.
2. Noncompetitive Inhibition
•Noncompetitive inhibitors bind on site other than the
active site, reduce enzyme affinity to the substrate
• Can be described by:
Rate equation:
Overcome by adding reagents to block binding of inhibitor
3. Uncompetitive Inhibition
Inhibitors binds to the ES complex only and
have no affinity for the enzyme itself
Rate of reaction,
v
Vm, app [ S ]
K ' m ,app  [ S ]
The net effect is reduction in both Vm and K’m
values.
Substrate inhibition
• High substrate concentrations may cause
inhibition in some enzymatic reaction
The Effects of Enzyme
Inhibitors
Substrate
analogs and
compete with
substrate for the
active site of the
enzyme
•Noncompetitive inhibitors bind
on site other than the active
site, reduce enzyme affinity to the
substrate
Effect of pH and temperature
Effect of pH on Enzyme
Kinetics
-Enzyme are active only over small range of pH due to:
 the active site functional group charges (ionic form)
 the three dimensional shape of enzyme are pHdependent
-these ionic group on active sites must be in a suitable form
(acid or base) to function.
-Variation in pH of medium result in changes of:
Ionic form of the active site
Activity of enzyme, hence the reaction rate
Affect the maximum reaction rate, Km and enzyme stability
-Scheme to describe pH dependence of the enzymatic
reaction rate for ionizing enzymes.
-
Ionic groups at active site
Variation of enzyme activity with
pH for 2 different enzymes (A) and (B)
-Optimum pH is usually determined experimentally
Effect of Temperature on
Enzyme Kinetics
ascending
descending
The rate varies according to
Arrhenius equation
Thermal
denaturation
occurred
Variation of reaction rate with temperature
The rate,
v  k2 [ E ]
k2  Ae
 Ea / RT
 kd is the denaturation constant,
k d  Ad e
 Ea / RT
Enzyme denaturation is much faster than enzyme
activation.
Variation in T affect both Vm and Km
Thank You
MID TERM EXAM 1
week 6
FRIDAY, 16 Oct 2015 (DKD1)
9-11 am