The document discusses enzymes kinetics and provides details about enzymes. It defines enzymes as proteins that act as biological catalysts and accelerate chemical reactions. It describes the lock and key theory and induced fit theory of how enzymes bind to substrates. It explains the three phases of enzyme kinetics - pre-steady state, steady state, and post-steady state. It also discusses factors that affect enzyme activity like substrate concentration, temperature, and pH. The Michaelis-Menten kinetics model and equation are explained. Finally, it provides examples of applications of studying enzyme kinetics.

1. Enzymes Kinetics
Pranjal Sharma
(20-PZO-016)

2. Enzymes
Enzymes are proteins that act as biological catalysts (biocatalysts). Catalysts
accelerate chemical reactions,often by several orders of magnitude. The molecules
upon which enzymes may act are called substrates, and the enzyme converts the
substrates into different molecules known as products.

3. Enzyme (Catalysts) lower the activation energy of the reaction.

4. Characteristics of Enzyme
● Like all catalysts, enzymes accelerate the rates of reactions while
experiencing no permanent chemical modification as a result of their
participation.
● Catalytic power and specificity
● Catalysis takes place at particular site on the enzyme called the active site.
● If and enzyme is denatured, its catalytic activity is lost, therefore Primary,
Secondary , Tertiary and Quaternary structure of enzymes are essential for
their catalytic activity.
● Enzymes like other proteins have molecular weight ranging from 12000 to
more than 1 million Dalton.

5. EXCEPTION!!!
The ribozyme discovered by Thomas Cech and others in 1986 is
a catalyst made of RNA, which acts on itself and cleaves RNA.

6. Enzyme Theories
There are two theories that describe the binding of enzymes:
1) Lock and Key Theory: The shape of the enzyme's active site is complementary
to that of its substrate, by German chemist Emil Fischer in 1899
2) Induced Fit Theory: The active site has a flexibility of shape, thus when an
appropriate substrate comes in contact with the enzyme's active site, the shape of
the active site would change to fit with the substrate,by Koshland in 1958

9. ENZYME KINETICS
● Enzymes = Biocatalysts
● Kinetics = study of motion and its causes.
● Enzyme + kinetic = The study of biochemical reaction
rates catalyzed by an enzyme.

10. Study of enzyme kinetics is useful for measuring
• concentration of an enzyme in a mixture (by its catalytic activity),
• its purity (specific activity),
• its catalytic efficiency and/or specificity for different substrates
• comparison of different forms of the same enzyme in different tissues or
organisms,
• effects of inhibitors (which can give information about catalytic mechanism,
structure of active site, potential therapeutic agents...)

11. Quick History
•Enzyme kinetics began in 1902 when Adrina Brown reported an
investigation of the rate of hydrolysis of sucrose as catalyzed by the yeast
enzyme invertase.
• Brown demonstrated – when sucrose concentration is much higher than
that of the enzyme, reaction rate becomes independent of sucrose
concentration.
• Brown proposal – overall reaction is composed of two elementary reactions
in which the substrate forms a complex with the enzyme that subsequently
decomposes to products and enzymes

12. E = Enzyme
S = Substrate
P = Product
ES = Enzyme-Substrate complex
k1= rate constant for the forward reaction
k-1 = rate constant for the breakdown of the ES to substrate
k2 = rate constant for the formation of the products

13. According to this model
• When the substrate concentration becomes high enough
to entirely convert the enzyme to the ES form, the second
step of the reaction becomes rate limiting step.
• The overall reaction rate becomes insensitive to further
increase in substrate concentration.

15. Phase Concentration of ES Rate of product
formation
Pre-steady state Rapid burst of ES
complexes form
Initially slow, waiting for
ES to form, then speeds
up
Steady state(equilibrium) ES concentration remains
constant as it is being
formed as quickly as it
breaks down
Constant rate of
formation, faster than
pre-steady state
Post-steady state Substrate depletes so
fewer ES complexes form
Slow as there are fewer
ES complexes; slows
down as substrate runs
out
Reaction Kinetics
When an enzyme is added to a substrate, the reaction that follows occurs in three
stages with distinct kinetics:

16. Kinetics Basics
m+n = order of reaction

17. Enzyme Kinetics - Factors
The catalytic properties of enzymes, and consequently their activity, are
influenced by numerous factors.
These factors include
• Physical quantities (temperature, pressure),
• The chemical properties of the solution (pH value, ionic strength),
• The concentrations of the relevant substrates, cofactors, and inhibitors.

18. Substrate concentration
For a typical enzyme, as substrate concentration is increases the rate of reaction
i.e. initial velocity(Vi) will also increases, untill it reaches the max value (Vmax)
after that further increase in substrate concentration the rate of reaction will not
increase, Because enzyme is saturated with substrate.

19. Effect of Temperature
● Raising the temperature increases the rate of enzyme catalyzed reactions.
● The temperature range over which an enzyme remain stable is varies from
organism to organism, for example Humans exhibit stability around 37.5°C.
● The rate of most the of the biological reactions doubles for every 10° rise in
temperature.
However ,the enzyme catalyze reaction cannot
increases indefinitely with the rise in temperature
because most of the enzyme tend to denature and
loose their catalytic activity after certain limit.

20. Effect of pH or H+ ion concentration
● All the enzyme catalyze reactions exhibit significant dependence on H+ ion
concentration.
● Most intracellular enzyme exhibit optimal activity at pH 5 and 9.
● The relationship of enzyme activity to H+ ion concentration reflects the
balance between enzyme denaturation at high or low pH.

21. Michealis-Menten Analysis
• Michaelis–Menten kinetics is one of the simplest and best-known models of enzyme
kinetics.
• The model serves to explain how an enzyme can cause kinetic rate enhancement of a
reaction and why the rate of a reaction depends on the concentration of enzyme present.
• To begin our discussion of enzyme kinetics, let's define the number of moles of product
(P) formed per time as V.
• The variable, V, is also referred to as the rate of catalysis of an enzyme.
• For different enzymes, V varies with the concentration of the substrate, S.
• At low S, V is linearly proportional to S, but when S is high relative to the amount of total
enzyme, V is independent of S.

22. The Michaelis - Menten Equation

23. It shows that the maximal velocity (Vmax) is approached asymptotically.
(Km) is the substrate concentration yielding a velocity of Vmax/2

24. Significance of Km
Km is the concentration of substrate at which half the active sites are filled. It
provides a measure of the substrate concentration required for significant
catalysis to occur.

25. Significance of Vmax
The maximal rate, Vmax, reveals the turnover number of an enzyme
● It is the number of substrate molecules converted into product by an enzyme
molecule in a unit time when the enzyme is fully saturated with substrate.
● The maximal rate, Vmax, reveals the turnover number of an enzyme if the
concentration of active sites [E]T is known,

26. Applications of Enzyme Kinetics Study
One of the applications of enzyme kinetics is the determination of dissociation
constants for antigen-antibody interactions in solution.
According to researchers double reciprocal plots of Elisa signals versus antigen
concentration helps in studying antigen antibody binding and hence aids drug
designing.
1. Kinetics study also helps in studying drug metabolism
2. Since the enzyme’s activity is affected by concentration, ph etc studying the
kinetics might help in providing suitable conditions for drug to be efficient.

28. IT’S Time For
QUIZ

29. 1. Name an enzyme which is not proteinaceous in nature?
A. Cellulases
B. Xylanases
C. Ribozyme
D. Peptidiase

30. 2. Which statement about enzymes is true
(a) enzymes accelerate reactions by lowering the activation energy
(b) enzymes are proteins whose three-dimensional form is key to their
function
(c) enzymes do not alter the overall change in free energy for a reaction
(d) all of these

31. 3.Name an enzyme that digests fat?
A. Lipase
B. Sucrase
C. Maltase
D. Fructose

32. 4.In humans salivary amylase enzyme breaks down starch. The
optimum pH for this reaction is:
A. 6
B. 6.2
C. 6.4
D. 6.8

33. 5. Who coined the word enzyme?
A. Wilhelm Kuhne
B. Alfred Russel
C. Robert Koch
D. Rosalind Franklin

34. ANSWERS
1. C
2. D
3. A
4. D
5. A

35. References
● https://teachmephysiology.com/biochemistry/molecules-and-signalling/enzym
e-kinetics/
● https://www.biologydiscussion.com/enzymes/the-kinetics-of-enzyme-action-wi
th-diagram/3297
● https://www.biologydiscussion.com/enzymes/kinetics-of-enzymatic-reactions-
biochemistry/71432
● https://www.slideshare.net/KamalKishor31/enzyme-kinetics-57408548
● https://www.slideshare.net/mohdsakharkar/enzyme-final
● https://www.slideshare.net/sakshisaxena9256/enzyme-kinetics-42456967
● https://www.slideshare.net/RIZWANABBAS3/enzyme-kinetics-81061982
● https://www.slideshare.net/manjunathasanka/enzyme-kinetics-74490821
● https://www.slideshare.net/sachithGamage/enzyme-kinetics-72294243

36. Thank you for
listening to my
presentation.
❤❤