Chapter 6.

Enzymes and biocatalysis

6.1. The nature of chemical reactions
 Metabolism is the sum of the chemical reactions in an organism.
 Catabolism is the energy-releasing processes.
– Breaking chemical bonds between molecules
– hydrolysis (use water)
– Digestion
 Anabolism is the energy-using processes. biosynthetic,
 building of complex molecules from simpler ones, involve
dehydration synthesis reactions (reactions that release water)
 dehydration synthesis
 synthesis

1
A. Endergonic reactions
 Chemical reaction that requires a net in put of energy.
e.g. Photosynthesis

6CO2 + 6H2O C6H12O6 + 6O2 photons

SUN Light
B. Exergonic reactions Energy
 Chemical reactions that releases energy.
e.g. Cellular respiration.
C6H12O6 + 6O2  6CO2 + 6H2O +

(glucose)
ATP
Energy

2
6.2. Catalyst
 Catalyst speeds up a chemical reaction without being permanently
altered themselves
6.3 Enzyme
 Enzymes are biological catalysts produced in cells
 Every chemical reactions in a cell catalyzed by enzymes
 Globular proteins with characteristic three-dimensional shapes.
 Cells need enzymes because reactions too slow to maintain life
without enzymes

3
6.4. Enzymes and their properties
Properties of enzymes are:
 Excellent catalysts
 Speeding up reactions 108 to 1020 fold from normal
 A catalase decompose 40 million molecules of H2O2 per
second
 Specific – one enzyme for one substrate
 Unchanged at the end of the reaction - (not used up by the
reaction)
 It works rapidly and efficiently therefore is required in small
quantity.
 Require optimum conditions at which they work best
 Regulated: some enzymes can sense metabolic signals.

4
6.5. Activation energy (AE)
 AE is the energy required to break the existing bonds and begin
the reaction.
 Amount of energy needed to destabilize/weaken the bonds of a
molecule.
 During this part of the reaction the molecules are said to be in a
transition state.
– large biomolecules are stable
– must absorb energy to break bonds
How enzymes reduce AE?
-By weakening bonds and lowering the AE necessary to initiate a
chemical reaction.

5
Reaction pathway

6
7
6.6. Enzyme specificity
 Most enzymes catalyze only one particular reaction
 Folded conformation of enzymes creates an area known as the
active site.
 Active site is the portion of the enzyme to which substrate binds.
 The area on the enzyme where the substrate or substrates attach to
is called the active site.
 Most enzymes are highly specific, Why?
 Because the shape of the active site is closely related to the
shape of the substrate.
 The substrates of enzymes are the reactants that are activated by
the enzymes
 Enzymes are specific to their substrates
 The specificity is determined by the active site

8
6.7 Mechanisms of enzymatic action/work
How enzymes work???
Three mechanisms by which enzymes accelerate reactions:
(a) Maintaining precise substrate orientation.
(b) Changing substrate reactivity.
(c) Exerting physical stress on bonds in the substrate to be broken.
 In enzyme catalysed reaction
 Substrate first binds to the active site of enzyme to form the
enzyme-substrate complex.
 The substrate is transformed into products.
 The products are then released and

9
Finally the enzyme is recovered unchanged and can be re-used
again until the reaction finished.
The entire enzymatic reaction may take only a few
milliseconds

10
• Transition state has features of both substrate and product and
falls apart to yield product, which dissociates from the
enzyme.
Mechanisms of enzyme action

11
6.8. Enzyme mode of action
 Enzyme mode of action has two models:
 Lock and key model
 Induced fit model
A. Lock and key model
 Enzyme is assumed as the lock & substrate as the key.
 substrate fit exactly to the active site (AS)
 The enzyme is generally much larger than the substrate(s).
 the active site has a rigid shape, not flexible
- only substrates with the matching shape can fit
This is an older model, however, and does not work for all enzymes.

12
13
B. Induced fit enzyme model
 This model assumes that the enzyme active site is more a flexible
pocket.
 Active site changes its shape (conformation) to accept substrate.
 This change works to distort and strain substrate (S) forcing it
into transition state.
 The active site assumes shape complementary to S only when S is
bound.
 Active site changes its shape called induced fit model.
 the active site is flexible, not rigid
- the shapes of the enzyme, active site, and substrate adjust to
maximumize the fit, which improves catalysis
- there is a greater range of substrate specificity
 This model is more consistent with a wider range of enzymes

14
Induced fit model of enzyme
15
Are all enzymes made up of only proteins?
6.9. Cofactors
 Some enzymes are indeed consist only pure proteins (e.g. pepsin).
 However, most enzymes are consisting of a non protein portion
(cofactor) in addition to a protein portion (apoenzyme).
 An enzyme containing apoenzyme and cofactor is known as
holoenzymes.
 Holoenzymes perform catalytic activity.
 But neither the apoenzyme nor the cofactor alone has catalytic
activity

16
Cofactor: is a non-protein chemical compound that is bound (either
tightly or loosely) to an enzyme and is required for catalysis.
– Types of Cofactors:
• Coenzymes.
• Prosthetic groups.
• Coenzyme:
• The non-protein component, loosely bound to apoenzyme by
non-covalent bond.
• Examples: Complex organic molecules such as NAD+, NADP+,
FAD, vitamin or hormone
• Prosthetic group
• The non-protein component, tightly bound to the apoenzyme
by covalent bonds.
• Examples: Inorganic elements such as iron, copper, magnesium,
zinc, or calcium.

17
What are the factors that affect enzyme activity?
6.10. Factors affecting enzyme activity
A. Temperature
 Temperature increment is fatal for cell
 Enzymes, like all proteins, change shape when exposed to heat
or cold.
 Temp. strongly influences enzyme activity.
 Optimum temp. for maximum enzyme function is usually about
35-40oC.
 At low temp. reactions occur slowly or not at all
 Above 45oC most enzymes are denatured (lost their shape and
function).

18
19
B. pH
 Most enzymes have an optimal pH range (work best).
 Extreme pH changes will denature the enzyme.
 Optimum pH in most living things is close to 7 (neutral).
e.g. The optimal pH for human enzymes is b/n 6 and 8.
 High/low pH levels usually slow enzyme activity.
• A few enzymes (e.g. gastric protease, pepsin) work best at a pH
2.
• Extreme pH levels will produce denaturation
• The structure of the enzyme is changed
• The active site is distorted and the substrate molecules will no
longer fit in it

20
Enzyme
activity Trypsin

Pepsin

1 3 5 7 9 11
pH

21
C. Enzyme concentration
 The more the enzymes concentration the more the reaction
faster.
The initial rate of an enzyme-catalyzed reaction is always
proportionate to the concentration of enzyme.

[S]>>[E] V∝[E]

22
D Substrate concentration
 Enzymatic activity increases as substrate concentration increases
until the enzymes are saturated.
 The rate of reaction will increase to a point and then level off
when there is a fixed amount of enzyme and excess substrate.
 The increase in velocity is proportional to the substrate
concentration.
• Faster reaction but it reaches a saturation point when all the
enzyme molecules are occupied
• Alter the concentration of the enzyme then Vmax will change too

23
 Substrate concentration: Enzymic reactions

Vmax

Reaction
velocity

Substrate concentration

24
 Reaction
Velocity (v0)

Substrate Concentration/arbitrary Units

25
E. Enzyme inhibitors
 Enzyme inhibitors are molecules that are able to bind to an
enzyme and reduce its activity.
 Inhibitors affect the enzyme activity by compete with the
substrate for the active site of the enzyme
 Enzyme inhibitors can be divided in to two types: reversible and
irreversible.
1.Reversible inhibition
 Reversible inhibitors bind only loosely to an enzyme, and thus
are readily displaced.
 Reduces product formation temporarily.
 Enzyme activity is restored when the inhibitor dissociates from
the enzyme-inhibitor complex.
 Reversible inhibitors can be classified competitive and non
competitive.
26
A. Competitive inhibition
 inhibitors structurally resemble the substrate and compete with
substrate to bind at the normal active site.
 Inhibitor and the substrate compete to bind the same active site.
 The effectiveness of a competitive inhibitor depends on its
relative affinity for the enzyme.
B. Noncompetitive inhibition
 Substrate and inhibitor do not compete for the same binding
site.
 So the inhibitor acts at a site other than the enzyme’s active site
(allosteric site or other space).
 These inhibitors modify the shape of the active site once bound
elsewhere on enzyme and making it nonfunctional

27
Competitive inhibitors
28
2. Irreversible Inhibition
 Irreversible inhibitors bind very tightly to an enzyme by
forming a covalent bond to one of its amino acid residues
(active sites).
 These inhibitors prevent/inhibit formation of the E-S
complex and the products.
 occurs when an inhibitor forms a covalent bond with a
specific functional group of an enzyme, thereby inactivating
it.
–Irreversible inhibitors include:
-Arsenic (poisonous elements)
-Snake venom
-Nerve gas

29
Naming of enzymes
 Most enzymes have names that end in the suffix -ase.
 The first part of the name often indicates the substrate.
e.g. Urease, maltase, cellulase, lactase.
 Other enzymes are named for the substrate and the reaction
catalyzed.
e.g. Lactate dehydrogenase, Pyruvate decarboxylase.
 Some names are historical. e.g. Catalase, pepsin, trypsin.
-No direct relationship to substrate or reaction type.
 Enzyme classification is based on a system originally established
by the Commission on Enzymes of the International Union of
Biochemistry (CEIUB) (1979).

30
There are six main classes, grouped according to the type of
reaction catalysed:
1. Oxidoreductases: catalyse oxidation/reduction reactions in which
oxygen and hydrogen are gained or lost.
e.g. Cytochrome oxidase, lactate dehydrogenase.
2. Transferases: catalyse transfer of a group from one molecule to
another. e.g. Acetate kinase, alanine deaminase.
3. Hydrolases: catalyse hydrolysis, the cleavage of bonds by addition
of a water molecule. e.g. Lipase, sucrase.
4. Lyases: catalyse splitting bonds, other than via hydrolysis or
oxidation. e.g. Oxalate decarboxylase, isocitrate lyase.
5. Isomerases: catalyse structural rearrangements of molecules.
e.g. Glucose-phosphate isomerase, alanine racemase.
6. Ligases: catalyse the formation of new bonds using energy
(breakdown of ATP). e.g. Acetyl-CoA synthetase, DNA ligase.
31
REGULATION OF ENZYME ACTIVITY
1. Allosteric binding sites: Allosteric enzymes are regulated by
molecules called effectors (modifiers) that binds nonconvalently
at a site other than the active site.
2. By Covalent Modification: Many enzymes are regulated by
covalent modification, most frequently by the addition or
removal of ‘phosphate’ group to serine, threonine or tyrosine
residue of the enzyme by kinases. (enzyme)
3. Induction and repression of enzyme synthesis: Cells can also
regulate the amount of enzymes present by altering the rate of
enzyme synthesis.
32