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COMPLEX ENZYME KINETICS
Inhibition
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Types of inhibition
Inhibition
Reversible Irreversible
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Inhibited enzyme reactions
There are two distinct types of inhibitors:
- Irreversible inhibitors form a stable complex with enzymes
and reduce enzyme activity (e.g. lead, cadmium,
organophosphorous pesticide)
- Reversible inhibitors interact more loosely with enzymes
and can be displaced.
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competes with the
substrate for the active
site of an enzyme.
inhibitor (I) occupies
the active site and it
prevents binding of the
substrate to the enzyme.
compounds that
resemble the substrate
and combine with the
enzyme to form an EI
complex.
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Assuming rapid equilibrium and with the definition of
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Km,app > Km
Maximum rate of reaction is unchanged
Increases Km by a factor (1+[I]/KI) i. e. the affinity of the
enzyme to the substrate decreases and also reduces reaction
rate.
The net effect of competitive inhibition is an increased value of
Km app and, therefore it can be overcome by high
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concentration of substrate.
Non-competitive inhibition
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Non-competitive inhibition
K-1 K-3
= KM =
k1 k3
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Non-competitive inhibition
In the presence of a non-competitive inhibitor,
the maximal rate of the reaction (Vmax) is lower
but the Michaelis constant (KM) is unchanged.
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The net effect of noncompetitive inhibition is in
reduction in Vm, therefore, high concentration of
substrate would not overcome noncompetetitive
inhibition.
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Uncompetitive inhibition
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k1
k-1 k2
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The net effect of uncompetitive inhibition is in reduction
of both Vm and Km, therefore, net result in reduction of
reaction rate.
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Substrate / Product inhibition
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Substrate / Product inhibition
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UNCOMPETITIVE SUBSTRATES
Uncompetitive Substrates Inhibition
Inhibition
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At high substrate concentrations, Km[S] 1, and inhibition is
dominant. The rate in this case is
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Optimum Substrate Concentration
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Competitive versus Uncompetitive inhibition
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Allosteric Enzymes
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Allosteric Enzymes Kinetics
Kinetic properties cannot be studied by using the Michaelis-
Menten equation.
The Hill
equation
Hill constant Hill or cooperative
coefficient
n = 1 gives Michaelis-Menten kinetics
n > 1 gives positive cooperativity
n < 1 gives negative cooperativity
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http://chemwiki.ucdavis.edu/Biological_Chemistry/Catalysts/Enzymatic_Kinetics/Sigmoid_Kinetics
Allosteric Enzymes Kinetics
Examples of the S-shaped sigmoidal/Hill curve, which is
different from the hyberbolic curve of M-M kinetics.
[S]=Km [S]n=Km
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Allosteric Enzymes Kinetics
For an alternative formulation of Hill equation, we could
rewrite it in a linear form as follows:
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Example 1
An inhibitor (I) is added to the enzymatic reaction at a level of 1.0
g/l. The following data were obtained for Km=9.2 g S/l.
a. Is the inhibitor competitive or noncompetitive?
b. Find KI
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Effects of pH on enzyme activity:
Three dimensional structure of proteins is based on the interaction
between R groups
R groups can carry a charge: Asp (-), Glu (-), Cys (-), Tyr (-),
His (-), Arg (+), Lys (+)
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Effects of pH on enzyme activity:
pH
Prof. R. Shanthini 32
Updated: 23 Nov 2012 https://wikispaces.psu.edu/display/230/Enzyme+Kinetics+and+Catalysis
Effects of pH on enzyme activity:
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
Effects of pH on enzyme activity:
Catalase enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
Effects of pH on enzyme activity:
Invertase enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
EFFECT of pH on ENZYME KINETICS
Cause:
Some of the Enzyme has ionic groups on their active sites and this
ionic group must be in suitable form (acid/base) to function.
Variation of the pH indicates the results in changes in the ionic
forms.
Changes of pH also alter the shape of 3-D structure of enzyme.
Results:
Enzymes are only actives over certain pH ranges and medium may
affect the maximum reaction rate, Km and the stability of the
enzyme.
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Effect of pH on Enzyme Kinetics
(Ionizing Enzyme)
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We can derive the following rate expression:
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For the case of ionizing substrate, the following scheme and rate
expression can be developed:
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Effects of temperature on enzyme activity:
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http://academic.brooklyn.cuny.edu/biology/bio4fv/page/enz_act.htm
Effects of temperature on enzyme activity:
Denaturation of the enzyme:
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Effects of temperature on enzyme activity:
Denaturation for most human enzymes:
The optimum
Optimal for most
temperature for most
human enzymes
human enzymes to
work at is around
37C which is why
this temperature is
body temperature.
Enzymes start to
denature at about
40C.
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http://www.woisd.net/moodle/mod/resource/view.php?id=44
Effects of temperature on enzyme activity:
enzymes
Temperature (deg C) 43
Prof. R. Shanthini
Updated: 23 Nov 2012 https://wikispaces.psu.edu/display/230/Enzyme+Kinetics+and+Catalysis
Effects of temperature on enzyme activity:
The rate of enzyme-catalyzed reactions increases with
temperature to a certain limit.
Above a certain temperature, enzyme activity decreases
with temperature because of enzyme denaturation.
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temperature inactivation or thermal
denaturation. The kinetics of thermal
denaturation can be expressed as
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The activation energies of enzyme-catalyzed
reactions are within the 4 to 20 kcal/g mol range
(mostly about 11 kcal/g mol).
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Notes
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Effects of pH on enzyme activity:
Maltase enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
Effects of pH on enzyme activity:
Pepsin enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
Effects of pH on enzyme activity:
Trypsin enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm
Effects of pH on enzyme activity:
Urease enzyme
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www.docbrown.info/page01/ExIndChem/ExIndChema.htm