1. Introduction to enzymes, enzyme catalyzed reactions and
CP504 – Lecture 3
Enzyme kinetics and associated reactor design:
Introduction to enzymes,
enzyme catalyzed reactions and
simple enzyme kinetics
learn about enzymes
learn about enzyme catalyzed reactions
study the kinetics of simple enzyme catalyzed reactions
Prof. R. Shanthini Sept 2011

2. What is an Enzyme?
Enzymes are mostly proteins, and hence they consists of amino acids.
Enzymes are present in all living cells, where they help converting nutrients into energy and fresh cell material.
Enzymes breakdown of food materials into simpler compounds.
Examples:
- pepsin, trypsin and peptidases break down proteins into amino acids
- lipases split fats into glycerol and fatty acids
- amylases break down starch into simple sugars
Prof. R. Shanthini Sept 2011

3. What is an Enzyme? Enzymes are very efficient (biological) catalysts.
Enzyme catalytic function is very specific and effective.
Enzymes bind temporarily to one or more of the reactants of the reaction they catalyze.
By that means, they lower the amount of activation energy needed and thus speed up the reaction.
Enzymes does not get consumed in the reaction that it catalyses.
Prof. R. Shanthini Sept 2011

4. Enzyme classification
Oxidoreductase: transfer oxygen atoms or electron
Transferase: transfer a group (amine, phosphate, aldehyde, oxo, sulphur, etc)
Hydrolase: hydrolysis
Lyase: transfer non-hydrolytic group from substrate
Isomerase: isomerazion reactions
Ligase: bonds synthesis, using energy from ATPs
Prof. R. Shanthini Sept 2011

5. Examples of enzyme catalyzed reactions
Examples of Enzyme Catalysed Reactions
Example 1:
Carbonic anhydrase
CO2+ H2O H2CO3
Carbonic anhydrase is found in red blood cells.
It catalyzes the above reaction enabling red blood cells to transport carbon dioxide from the tissues (high CO2) to the lungs (low CO2).
One molecule of carbonic anhydrase can process millions of molecules of CO2 per second.
Prof. R. Shanthini Sept 2011

6. Examples of enzyme catalyzed reactions
Catalase
2H2O H2O + O2
Catalase is found abundantly in the liver and in the red blood cells.
One molecule of catalase can breakdown millions of molecules of hydrogen peroxide per second.
Hydrogen peroxide is a by-product of many normal metabolic processes.
It is a powerful oxidizing agent and is potentially damaging to cells which must be quickly converted into less dangerous substances.
Prof. R. Shanthini Sept 2011

7. Industrial use of catalase
- in the food industry for removing hydrogen peroxide from milk prior to cheese production
- in food-wrappers to prevent food from oxidizing
- in the textile industry to remove hydrogen peroxide from fabrics to make sure the material is peroxide-free
- to decompose the hydrogen peroxide which is used (in some cases) to disinfect the contact lens
Prof. R. Shanthini Sept 2011

8. Examples of Industrial Enzymes
See the hand out on the same topic
Prof. R. Shanthini Sept 2011

9. More on enzymes Enzymes are very specific.
Absolute specificity - the enzyme will catalyze only one reaction
Group specificity - the enzyme will act only on molecules that have specific functional groups, such as amino, phosphate or methyl groups
Linkage specificity - the enzyme will act on a particular type of chemical bond regardless of the rest of the molecular structure
Stereochemical specificity - the enzyme will act on a particular steric or optical isomer
Prof. R. Shanthini Sept 2011

10. Source: http://waynesword.palomar.edu/molecu1.htm
Prof. R. Shanthini Sept 2011
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11. E + S ES Source: http://waynesword.palomar.edu/molecu1.htm
Prof. R. Shanthini Sept 2011
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12. Lock & Key Theory Of Enzyme Specificity
(postulated in 1894 by Emil Fischer)
E + S ES
E + P
Prof. R. Shanthini Sept 2011
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13. Prof. R. Shanthini Sept 2011

14. Active Site Of Enzyme Blocked By Poison Molecule
Prof. R. Shanthini Sept 2011
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15. Induced Fit Model E + S ES E + P
(postulated in 1958 by Daniel Koshland )
E + S ES
E + P
Binding of the first substrate induces a conformational shift that helps binding of the second substrate with far lower energy than otherwise required. When catalysis is complete, the product is released, and the enzyme returns to its uninduced state.
Prof. R. Shanthini Sept 2011
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16. Simple Enzyme Kinetics
E + S ES
E + P k2
which is equivalent to
[E] S P
S for substrate (reactant)
E for enzyme
ES for enzyme-substrate complex
P for product
Prof. R. Shanthini Sept 2011

17. Michaelis-Menten approach to the rate equation:k1 k3
E + S ES
E + P k2
Assumptions:
Product releasing step is slower and it determines the reaction rate
2. ES forming reaction is at equilibrium
3. Conservation of mass (CE0 = CE + CES)
Initial concentration of E
Concentration of E at time t
Concentration of ES at time t
Prof. R. Shanthini Sept 2011

18. Michaelis-Menten approach to the rate equation:k1 k3
E + S ES
E + P k2
Product formation (= substrate utilization) rate:
rP = - rS = k3 CES
(1)
Since ES forming reaction is at equilibrium, we get
k1 CE CS = k2 CES
(2)
Prof. R. Shanthini Sept 2011

19. Michaelis-Menten approach to the rate equation:k1 k3
E + S ES
E + P k2
Using CE0 = CE + CES in (2) to eliminate CE, we get
k1 (CE0 – CES) CS = k2 CES
which is rearranged to give
CE0CS
CES =
(3)
k2/k1 + CS
Prof. R. Shanthini Sept 2011

20. Michaelis-Menten approach to the rate equation:k1 k3
E + S ES
E + P k2
Using (3) in (1), we get
k3CE0CS
rmaxCS
rP =
- rS = =
(4)
k2/k1 + CS
KM + CS
where rmax = k3CE0
and KM = k2 / k1
(5)
(6)
Prof. R. Shanthini Sept 2011

21. Briggs-Haldane approach to the rate equation:k1 k3
E + S ES
E + P k2
Assumptions:
1. Steady-state of the intermediate complex ES
2. Conservation of mass (CE0 = CE + CES)
Initial concentration of E
Concentration of E at time t
Concentration of ES at time t
Prof. R. Shanthini Sept 2011

22. Briggs-Haldane approach to the rate equation:k1 k3
E + S ES
E + P k2
Product formation rate:
rP = k3 CES
(7)
Substrate utilization rate:
rs = - k1 CECS + k2 CES
(8)
Since steady-state of the intermediate complex ES is assumed, we get
k1 CECS = k2 CES + k3 CES
(9)
Prof. R. Shanthini Sept 2011

23. Briggs-Haldane approach to the rate equation:k1 k3
E + S ES
E + P k2
Combining (7), (8) and (9), we get
rP = - rS = k3 CES
(10)
Using CE0 = CE + CES in (9) to eliminate CE, we get
k1 (CE0 - CES)CS = (k2 + k3)CES
which is rearranged to give
CE0CS
CES =
(11)
(k2+k3)/k1 + CS
Prof. R. Shanthini Sept 2011

24. Briggs-Haldane approach to the rate equation:k1 k3
E + S ES
E + P k2
Combining (10) and (11), we get
k3CE0CS
rmaxCS
rP =
- rS = =
(12)
KM + CS
(k2+k3)/k1 +CS
where rmax = k3CE0
and KM = (k2 + k3) / k1
(5)
(13)
When k3 << k2 (i.e. product forming step is slow),
KM = k2 / k1
(6)
Prof. R. Shanthini Sept 2011

25. Simple Enzyme Kinetics (in summary)
rmaxCS
rP =
- rS =
KM + CS
where rmax = k3CE0 and KM = f(rate constants)
rmax is proportional to the initial concentration of the enzyme
KM is a constant
Prof. R. Shanthini Sept 2011