Enzymes accelerate chemical reactions by lowering the activation energy of the reaction. They do this by providing an alternative pathway for the conversion of substrates to products, involving a transition state with lower energy. The Michaelis-Menten model describes the enzyme-catalyzed reaction, where the enzyme reversibly binds to the substrate to form an enzyme-substrate complex, before yielding products and regenerating the free enzyme. The Michaelis-Menten equation relates the initial reaction velocity to the substrate concentration.

4. MECHANISM OF
ENZYME
ACTION

5. MECHANISM OF ENZYME ACTION
 The catalytic efficiency of enzymes is explained by two
perspectives:
Thermodynamic
(Energy)
changes
Processes at the
active site

6. THERMODYNAMIC/ENERGY CHANGES
 All chemical reactions have energy barriers separating the
reactants and the products.This barrier called Activation
energy (Ea) is energy difference b/w that of reactants and high
energy intermediate,the Transition state(T) which is formed
during the conversion of reactant to products i.e. ES/EP-
complex,(no enz free,but bound to Reactant/Product).
 Because of high energy of activation,rates of uncatalysed
reactions are slow.
 Enzymes provide alternate pathway,by lowering Ea,the lower
the Ea,the more molecules have sufficient energy to pass
through the transition state,the faster the rate of reaction

7. Substrate Concentration and Reaction Rate
Rate• substrate concentration increases (at
constant enzyme concentration)
• Maximum activity occurs when the
enzyme is saturated (when all enzymes are
binding substrate)
• The relationship between reaction rate and
ial,
e
subst
and a
enzym
rate concentration is exponent
symptotes (levels off) when th
e is saturated

8. THERMODYNAMIC CHANGES
 Only a few substances cross the activation barrier and change
into products.
 That is why rate of uncatalyzed reactions is much slow.
 Enzymes provide an alternate pathway for conversion of
substrate into products.
 Enzymes accelerate reaction rates by forming transitional
state having low activational energy.
 Hence, the reaction rate is increased many folds in the
presence of enzymes.
 The total energy (ΛG) of the system remainsthe same.
 The equilibrium state (when forward reaction =backward
reaction) is not disturbed.

11. THERMO-DYNAMIC CHANGES
OVERVIEW

12. PROCESSES AT THE ACTIVE SITE
Covalent
catalysis
Acid base
catalysisCatalysis
by strain
Catalysis
by
proximity

13. COVALENT CATALYSIS
o Enzymes form covalent linkages with substrate forming
transient enzyme-substrate complex with very low activation
energy.
o Enzyme is released unaltered after completion of reaction.

14. Synthase
Acetyle choline + CoA-SH--------Acetyle CoA + Choline

15. ACID-BASE CATALYSIS
 Mostly undertaken by oxido- reductases enzyme.
 Mostly at the active site, histdine is present which act as both
proton donor and proton acceptor.

17. CATALYSIS BYPROXIMITY
forming In this catalysis molecules must come in bond
distance (mostly by ligases).
 When enzyme binds:
 A region of high substrate concentration is produced at active
site.
 This will orient substrate molecules especially in a position
ideal for them.

18. Carbonic Anhydrase
H2O+CO2---------------------------H2CO3

19. CATALYSIS BY BOND STRAIN
 Mostly undertaken by lyases.
 The enzyme-substrate binding causes reorientation of the
structure of site due to in a strain condition.
 Thus transitional state is required and here bond is unstable
and eventually broken.
 In this way bond between substrate is broken and converted
into products.

20. Fumarase
Fumaric acid + H2O--------------L.Malic acid

21. ENZYMES
KINETICS

22. INTRODUCTION
“It is a branch of biochemistry in which we study the rate of
enzyme catalyzed reactions.”
 Kinetic analysis reveals the number and order of the individual
steps by which enzymes transform substrate into products
 Studying an enzyme's kinetics in this way can reveal the
catalytic mechanism of that enzyme, its role in metabolism,
how its activity is controlled, and how a drug or an antagonist
might inhibit the enzyme

23. RATES OF REACTIONS AND THEIR DEPENDENCE ON
ACTIVATION ENERGY
 Activation Energy (Ea):
“The least amount of energy needed for a chemical reaction to
take place.”
 Enzyme (as a catalyst) acts on substrate in such a way that
they lower the activation energy by changing the route of the
reaction.
 The reduction of activation energy (Ea) increases the amount
of reactant molecules that achieve a sufficient level of energy,
so that they reach the activation energy and form the product.
Example:
 Carbonic anhydrase catalyses the hydration of 10⁶ CO₂
molecules per second which is 10⁷x faster than spontaneous
hydration.

24. ENZYMES LOWER THE ACTIVATION ENERGY OF A REACTION
Final energy state of
products
Initial energy state
of substrates
Activation energy
of uncatalysed
reactionsActivation energy
of enzyme catalysed
reaction
Progress of reaction (time)
Energylevelsofmolecules

25. KINETICS OF ENZYMES CATALYSIS
 Enzymes catalysis:
“ It is an increase in the rate of reaction with the help of
enzyme(as catalyst).”
 Catalysis by enzymes that proceed
mechanism, typically occurs when
via unique reaction
the transition state
intermediate forms a covalent bond with the enzyme(covalent
catalysis).
 During the process of catalysis enzymes always emerge
unchanged at the completion of the reaction.

26. Direction of enzyme reaction reaction
• Most enzyme reactions are bidirectional and are represented by
double arrows,e.g.
• A+B-----------C+D
• -----------
• However some reactions are favoured by thermodynamic
factors-free energy, in one direction only,and are represented by
a single arrow,e.g.
• A+B---------C+D

27. MICHAELIS-MENTEN MODEL & EFFECTSOF
SUBSTRATE CONCENTRATION
 Michaelis-Menten Model:
“According to this model the enzyme reversibly combines with
substrate to form an ES complex that subsequently yields
product, regenerating the free enzyme.”





where:
S is the substrate
E is the enzyme
ES-is the enzyme substrate complex
P is the product
K1,K-1 and K2 are rate constants
E + S ES E + P
k₁
k₋₁
k₂

28. MICHAELIS-MENTEN EQUATION
 Michaelis-Menten Equation:
“It is an equation which describes how reaction velocity varies
with substrate concentration.”
Vmax [S]
Vo=
Km+[S]
 Where
 Vo is the initial reaction velocity.
 Vmax is the maximum velocity.
 Km is the Michaelis constant =(k₋₁+k₂)/k₁.
 [S] is the substrate concentration.

29. ASSUMPTIONS FOR MICHAELIS-MENTEN EQUATION
Following assumptions are made in deriving the Michaelis-
Menten equation:
 Relative concentrations of E and S:
The substrate conc.[S] Is much greater than enzyme conc.
so that the % of total substrate bound by enzyme at any
one time is small.
 Steady-State assumptions:
The conc of ES complex does not change with time.
rate of formation of ES = rate of breakdown of ES
 Initial Velocity:
Initial velocities are used in the analysis of enzyme reactions
(bcz products are negligible at the start of the reaction).

30. SUBSTRATE CONCENTRATION

31. SUBSTRATE CONCENTRATION

32. Important Conclusions
• Km characteristics:
Small Km: shows high affinity of enzyme for
substrate,bcz low conc. of substrate is needed to
half saturate the enzyme.
Large Km: Reflect a low affinity of enzyme for
substrate bcz a high conc. Of substrate is needed to
half saturate the enzyme.
• Velocity relationship to enzyme conc.:Rate
increases by increasing enz. conc. at all sub conc.
• Reaction order: 1st order/zero order

35. LINEWEAVER-BURK PLOT
• When vo is plotted against [S], it is not always possible
to determine when Vmax has been achieved because
of the gradual upward slope of the hyperbolic curve at
high substrate concentrations.
• However, if 1/vo is plotted versus 1/[S], a straight line
is obtained . This plot is called the Lineweaver-Burk
plot.
• It can be used to calculate Km and Vmax as well as to
determine the mechanism of action of enzyme
inhibitors.

36. LINEWEAVER-BURK PLOT

37. PHARMACEUTICAL IMPORTANCE
 Enzymes are virtually involved in all physiological processes
which makes them the targets of choice for drugs that cure or
ameliorate human disease.
 Applied enzyme kinetics represents the principal tool by which
scientist identify and characterize therapeutic agents that
selectively inhibit the rates of specific enzymes catalyzed
processes.
 Enzymes kinetics thus play a critical role in drug discovery as
well as elaborating the mode of action of drugs.