ENZYMES

 Chemical reactions in biological systems are almost

always catalyzed, otherwise they would occur too slowly
 There are two ways to accelerate a chemical reaction:
1) Heating
increases the energy of the molecules and hence,
increases the percentage of molecules with the required
energy of activation
2) Catalysts
provide the second way to accelerate a chemical
reactions; they lower the activation energy, i.e., the
energy barrier that substrates must overcome before
they can be converted into products

Biological catalytic --> Enzyme

 Special kinds of protein molecules: long chain of amino acids bound
by peptide bonds
 Amino acid  organic acid with amine (-NH2) & carboxilic acid (-
COOH) groups
 produced by living cells
 Found in all organelles (e.g., the nucleus, mitochondria, ribosomes,
etc.), and in membranes.
 absolutely essential as catalysts in biochemical reactions. i.e.,
controlling the metabolic processes, converting nutrients into
energy, breaking down food materials, etc
 Decrease activation energy
 speed up the rate of the reaction so that equilibrium is
attained rapidly
 At equilibrium, the ratio of the substrate concentration(s) to
the product concentration(s) is the same, whether or not
enzyme is present.
 do not change the equilibrium of a chemical reaction
 increase the rate of reaction without themselves undergoing
permanent chemical changes.
 enzymes are released again after a reaction ceases and can
continue in another reaction
 This processes cannot go forever  limited stabilities and,
slowly, they become inactive
 The catalytic ability of enzymes is due to its
particular protein structure  Active site
 Active site: a site where specific chemical reaction
to catalyzed reaction occurred at a small portion of
the surface of an enzyme --> site where the
substrate(s) are bound
 Some physical and chemical interactions occur at
this site to catalyze a certain chemical reaction for a
certain enzyme.
1. The active site takes up a relatively small portion of the total
volume of the enzyme.
2. The active site is a three-dimensional entity.
3. Active sites are clefts or crevices.
4. The specificity of binding depends on the precisely defined
arrangement of atoms in the active site giving a direct fit or
an induced fit.
5 Most substrates are bound to enzymes by relatively weak
forces.

The parts of the protein not contributing directly to the active

site may be required to form or stabilize the structure.
 In an enzymatic reaction, the substrate binds to the
active site of the enzyme
 The active site can lower an EA barrier by
 Orienting substrates correctly
 Straining substrate bonds
 Providing a favorable microenvironment
 Covalently bonding to the substrate
 Enzyme activity depends on interaction between a specific
protein molecule and other molecules (non protein)  affect
the enzyme structure
 The enzyme: a protein, and a combination of one or more
parts are called “cofactors.”
 A cofactor is a non-protein chemical compound that is bound
to enzymes and is required for facilitating their activity as
catalysts
 An inactive enzyme, without the cofactor is called an
apoenzyme, while the complete enzyme with cofactor is the
holoenzyme
 The role of a cofactor is to activate the enzymes protein
by changing its geometric shape, or by actually
participating in the overall reaction
E.g vitamin, metal ion
 Cofactors can be classified depending on how tightly they
bind to an enzyme: coenzymes & prosthetic group.
• Coenzymes low MW organic which loosely bound
cofactors (e.g vitamin)
• prosthetic groups  small organic which tightly
bound cofactors
• An enzyme catalyst  highly specific; reducing
the number of side reactions and by-products.
A great variety of enzymes exist, which can
catalyze a very wide range of reactions.
• The rate of an enzyme-catalyzed reaction is
usually much faster
• the rates of reaction are easily controlled by
adjustment of incubation conditions
• Only require a small amount
• mild conditions of temperature and pH
• Simple supporting units
• Sensitive or unstable molecules  require more
care
• Expensive
• in some products, enzymes must be inactivated
or removed after processing which adds to the
cost of the product
• usually coated or immobilised on carrier materials
to reduce the risk of inhalation, reduce cost
• Like other proteins, enzymes may cause allergic
responses
LOCK & KEY
•the active site has a rigid
•Only the right key fits: right size
and shape, have charges in the
correct place, have the right
hydrogen-bond donors and
acceptors, and have just the right
hydrophobic patches shape
•This is an older model, however,
and does not work for all
enzymes
•This explains enzyme
specificity
•This explains the loss of activity
when enzymes denature.
• the active site is flexible, not rigid
• Binding the correct substrate 
change enzyme structure  catalytic
groups into right position to facilitate
reaction
• the shapes of the enzyme, active site,
and substrate adjust to maximumize
the fit, which improves catalysis
• This model is more consistent with a
wider range of enzymes
• Bad substrates can not to make the
specific interactions that cause the
conformation change  enzyme stays
in its inactive conformation.
• Explaining bad substrate not good
substrate
• The bonds of the substrate are
stretched to make reaction easier
(lowers activation energy).
 Poor substrates  bind the
enzyme in the wrong
orientation
 catalytic groups and specific
interactions that accelerate
the reaction of the correct
substrate come into play in
only a very small number of
the interactions between the
enzyme and a bad substrate.
 In contrast to the induced-fit
model, this model does not
require a change in the
conformation of the enzyme
• the binding of S to E
pulls/pushes specific
chemical bonds  make
the subsequent chemical
reaction easier
• If a bond has to be broken,
the enzyme grabs onto
both sides of the bond and
pulls.
• If a bond has to be
formed, the enzyme grabs
onto both sides and
pushes.
 High activity. Increase rate by
reducing Ea
 Selectivity. Active sites3D
pocket/cleft fit only to a specific
substrates
 Regiospecificity. An enzyme can
detect differences in the spatial
arrangement of atoms in a
compound  specific ring group
 when a reaction have a potential
form multiple structural isomers 
only forms one regioisomer
(=structural isomers)
 Stereospecificity in the choice of
substrate
stereoisomer: same molecule formula,
sequence but differ in 3D orientation
 cis-trans, optical isomer L or D; due to
asimmetric of enzyme

 Controllability by the amount of

substrate & by other factors
(temperature, pH, etc.) active site
 Environmentally friendly  lower by-
product produced. Natural substrates
 natural products
 Simplest enzymatic reaction:

 E+S ES P
 Real enzyme mechanism  more complexes; never proceed
through just 1 ES complex.
 Involved many steps pathway
 The slowest one determines the rate of overall reaction
 reaction rate is affected by reaction conditions (S,P,E)
  important, to understand the effectiveness &
characteristics of enzyme reaction; for process design
The reaction will start at a high rate
and slow down over time for
several reasons:
 The substrate will be used up and
the reaction rate will slow down as
each enzyme molecule spends
more time diffusing through the
solution before it collides with a
new substrate molecule.
 As the reaction proceeds, product
will accumulate which may tend to
inhibit the enzyme.
 The enzyme molecules may
gradually lose activity owing to
random denaturation.
1 Temperature
2 pH
3 Concentration of substrate
4 Concentration of enzyme
5 Concentration of cofactors (activators and coenzymes)
 An activator is a substance, other than the catalyst or one of
the substrates, that increases the rate of a catalyzed
reaction.
6 Concentration of inhibitors
 An inhibitor is a substance that diminishes the rate of a
chemical reaction  the process is called inhibition
 As with other chemical
reactions, the rates of
enzyme-catalyzed
reactionsare increasedby
increases in temperature.
 However, because
enzymes are proteins,
thermal denaturation of
the protein with
increasing temperature
will decrease the effective
concentration of the
enzyme.
 The upper limit for most
human enzymes is 40 to
50˚C (normal body
temperature 37˚C).
 The effect of
temperature
Optimum

Increasing
number of Denaturation
collisions (Q10)
Enzyme
activity

0 10 20 30 40 50
© 2017 Paul Billiet ODWS Temperature / °C
 Changes in pH will change
the ionization of the enzyme
that is, amphoteric that
can react both as an acid
as well as a base
 If the substrate possesses
ionizable groups, its ionic
state will change with
changes in pH
 Thus, as the ionic nature of
either the substrate or the
enzyme is changed from
that form that combines
with the other
 The pH optimum may be either narrow or broad.
 As the pH increases or decreases above or below the
optimum pH, the activity of the enzyme (the velocity
of the enzyme-catalyzed reaction) will decrease
because:
a. There will be decrease in enzyme substrate binding
due to repulsion or to loss of charge on either the
substrate or the enzyme.
b. the catalytic functional groups will be in the wrong
ionization state.
c. The enzyme may become denatured.
Illustration how a bell-shaped pH profile might occur:
 binding S-E is due to different charge
 E has a group at the active site which must be positively
charged in order for substrate to bind (pK is, e.g., 8.0)
 S has to be negatively charged in order for it to beattracted
to the enzyme active site (pK is, e.g., 4.0)
 At low pH : S is protonated (SH), will not bind to E that is
protonated ---> low rate
 At higher pH less than 6: proton removed from S (S-), closer
to conditin for active site to bind--> increase E activity
 At pH 6: 100% S as (S-) and max E activity
 pH > 6: S become more (S+), E get more (E+)
Plotting r0 at different  [S] very low  a
[S] & [E]: straight line graph
 r is proportional to [S]  when [S] is low, [S] 
(1st-order reaction)  the limiting factor; an
increase [S] produces a
when the [S] is in the proportional increase in
low range the rate
 graph will curve at higher [S] & will level off at very
high [S]
 The rate  does not depend on the [S] when the [S]
is high
 Reaction rate changes gradually from first order to
zero order as [S] is increased.
 [S] increases, [E]  the limiting factor
 a further increase in substrate  produces a less
than-proportional increase in reaction rate.
 the enzyme becomes "saturated" and the reaction
rate reaches a constant value doesn't increase
significantly as yet more substrate is added.
• [S] low  they are all bound to
the enzyme molecules
• there are more substrate
molecules some of them are
free in solution, not bound to an
enzyme molecule.
• [S] high  all the enzyme
molecules have a bound
substrate.
• The number of enzyme
molecules with bound substrate
is an indication of the reaction
rate  those enzymes "act" and
convert the substrate to
product.
• The purple highlight indicates
the region of the graph that
corresponds to illustration in the
left.
 Some enzymes need to be activated before to be
bound to the substrate (case of essential
activation).
decreases the velocity of the enzyme-catalyzed
reaction
3 types of inhibitor:
competitive,
uncompetitive and
non-competitive
• the inhibitor competes with the
substrate for the active site of the
enzyme, but only one can be bound
at the same time
• Thus, competitive inhibitors are
usually structurally similar to the
substrate.
• The inhibitor binds to the enzyme
only.
 In the case of an uncompetitive inhibition, the
inhibitor is not in competition with the substrate
for the active site of the enzyme.
 It binds only the substrate-enzyme complex.
 The substrate facilitates the binding of the
inhibitor to the enzyme.
 In the case of a non-competitive inhibition (also
said mixed inhibition), both types of inhibition
are present: the inhibitor can bind either the
free enzyme or the enzyme substrate complex.
ENZYME SPECIFICITY
 Enzymes have varying degrees of specificity for
substrates
 Enzymes may recognize and catalyze:
- a single substrate
- a group of similar substrates
- a particular type of bond
1. Oxidoreductases
• Catalyze oxidation-reduction in which hydrogen or oxygen
atoms or electrons are transferred between molecules
• E.g: oxidases, peroxidases, dehydrogenases
2. Transferases
 catalyze the transfer of an atom or group of atoms (e.g., acyl-,
alkyl-, and glycosyl-), from one molecular and/or functional
groups to another
 E.g: Phosphotransferases (Kinases), Transmethylases,
Transpeptidases
3 Hydrolases
 These involve hydrolytic reactions and their reversal
(degradation of H2O to OH− and H+ products).
 Catalyse the hydrolysis of various bonds Add water across a
bond.
 E.g: Protein hydrolyzing enzymes (Peptidases), Carbohydrases
(Amylase, Maltase, Lactase), Lipid hydrolyzing enzymes
(Lipase).
4. Lyases
 involve elimination reactions in which a group of atoms is
removed from the substrate
 Cleave various bonds by means other than hydrolysis and
oxidation.
 Add water, ammonia or carbon dioxide across double bonds,
or remove these elements to produce double bonds.
 Examples: Fumarase, Carbonic anhydrase.

5. Isomerases
 Catalyse isomerization changes within a single molecule.
 Carry out many kinds of isomerization: L to D isomerizations;
Mutase reactions (Shifts of chemical groups).
 Examples: Isomerase, Mutase.

6. Ligases
 catalyze the condensation of two molecules together with the
cleavage of ATP or another pyrophosphate bond
 Catalyse reactions in which two chemical groups are joined
(or ligated) with the use of energy from ATP.
 Examples: Acetyl~CoA Carboxylase,Glutamine synthetase
 Each enzyme has classification number consisting of four
digits:
 EC:   (2.7.1.1) HEXOKINASE
 EC: (2.7.1.1) these components indicate the following
groups of enzymes:
 2. IS CLASS (TRANSFERASE)
 7. IS SUBCLASS (TRANSFER OF PHOSPHATE)
 1.IS SUB-SUB CLASS (ALCOHOL IS PHOSPHATE
ACCEPTOR)
 1. SPECIFIC NAME
ATP,D-HEXOSE-6-PHOSPHOTRANSFERASE (Hexokinase)