PBS-BCH-221 (Biochemistry)
Enzymes
Enzymes
Enzymes are produced by all living organisms including
humans and present only in small amounts.
Enzymes are biological catalysts (also known as
biocatalysts) that speed up biochemical reactions in
living organisms.
Enzymes are to a large extent protein molecules,
although some enzymes are made from RNA and are
referred to as ribozymes.
Enzymes
They can also be extracted from cells and then used to
catalyse a wide range of commercially important
processes.
For example, they have important roles in the
production of sweetening agents and the modification
of antibiotics, they are used in washing powders and
various cleaning products, and they play a key role in
analytical devices and assays that have clinical,
forensic and environmental applications
Enzymes as catalysts
A catalyst does not change the chemical reaction but it
accelerates the reaction nor consumed in overall reaction.
They undergo chemical or physical change during reaction and
returns to original state at the end of reaction.
Transition state theory was proposed to explain action of
catalyst. For a chemical reaction A → B to occur, energy is
required.
When enough energy is supplied, A undergoes to transition
state which is an unstable state. So, it gets converted to
product B which is more stable.
The amount of energy needed to convert a substance from
ground state to transition state is called activation energy.
In presence of catalyst, A undergoes to transition state very fast
and requires less energy.
Enzymes as Catalysts
Enzymes reduce the reaction’s activation energy to
progress towards equilibrium quicker than the reactions that
are not catalyzed
Enzymes as Catalysts cont..
Hence, a catalyst accelerate the rate of reaction by
decreasing the energy of activation. Likewise, enzymes also
speed up reaction by lowering energy of activation. Activation
energy is very much less for a reaction in presence of
enzyme than non-enzyme catalyst
Enzymes qualify as catalysts because of four reasons
1. They are in low concentration as compared to the substrate
2. Enzymes do not affect the equilibrium of a reaction:
Equilibrium is when the rate of the forward reaction
equals the rate of the reverse reaction
3. Enzymes catalyzes equally a forward and backward
reaction
4. Enzymes remain unchanged at the end of the reaction
Characteristics of enzymes
Enzymes are protein catalysts that increase the velocity
of a chemical reaction, and are not consumed during the
reaction they catalyze.
1. Active sites: Enzyme molecules contain a special
pocket or cleft called the active site.
The active site contains amino acid side chains that
create a three dimensional surface complementary to
the substrate.
The active site binds the substrate, forming an enzyme-
substrate (ES) complex.
ES is converted to enzyme-product (EP), which
subsequently dissociates to enzyme and product.
Characteristics of enzymes cont..
2. Specificity
Enzymes are highly specific, interacting with one or a few
substrates and catalyzing only one type of chemical
reaction.
In fact, one of the properties of enzymes that makes them
so important as diagnostic and research tools is the
specificity they exhibit relative to the reactions they
catalyze.
A few enzymes exhibit absolute specificity; that is, they
will catalyze only one particular reaction.
Other enzymes will be specific for a particular type of
chemical bond or functional group. In general, there are
four distinct types of specificity:
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 and 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.
Characteristics of enzymes cont..
3- Cofactors:
Some enzymes associate with a non-protein cofactor that is
needed for enzyme activity.
Commonly encountered cofactors include metal ions such as Zn2+
or Fe2+and organic molecules, known as coenzymes, that are
often derivatives of vitamins.
For example, the coenzyme contains niacin, FAD contains
riboflavin, and coenzyme A contains pantothenic acid (vitamin B5).
Holoenzyme refers to the enzyme with its cofactor.
Apoenzyme refers to the protein portion of the holoenzyme.
In the absence of the appropriate cofactor, the apoenzyme
typically does not show biologic activity.
A prosthetic group is a tightly bound coenzyme that does not
dissociate from the enzyme.
Substrate binding
Enzymes must bind their substrates before they can catalyse
any chemical reaction.
Enzymes are usually very specific as to what substrates they
bind and then the chemical reaction catalysed.
Usually, an enzyme molecule has only one active site, and the
active site fits with one specific type of substrate.
An active site contains a binding site that binds the substrate
and orients it for catalysis.
Residues in the binding site form hydrogen bonds,
hydrophobic interactions, or temporary covalent interactions
(Van der Waals) with the substrate to make an enzyme-
substrate complex.
Substrate binding
A tighter fit between an active site and the
substrate molecule is believed to increase
efficiency of a reaction.
There are two proposed models of how enzymes
fit to their specific substrate: the lock and key
model and the induced fit model.
1. Lock and key model
2. Induced fit model
Lock and key model
Emil Fischer proposed, in
1894, that both the enzyme
and the substrate possess
specific complementary
(geometric) shapes that fit
exactly into one another
as"the lock and key".
The model explains enzyme
specificity, and assumes that
the active site is a perfect fit
for a specific substrate and
that once the substrate binds
to the enzyme no further
modification occurs.
Induced fit model
It was suggested by Daniel
Koshland in 1958).
It is a development of the lock-
and-key model and assumes
that an active site is flexible
and it changes shape until the
substrate is completely bound.
The substrate is thought to
induce a change in the shape
of the active site.
It also predicts that the
presence of certain residues
(amino acids) in the active site
will encourage the enzyme to
locate the correct substrate.
Induced fit model
Conformational changes may then occur as the substrate is
bound. After the products of the reaction move away from
the enzyme, the active site returns to its initial shape
The key difference between Induced Fit and Lock and Key
is that in induced fit theory, the binding of the substrate with
the active site of the enzyme induces the modification of the
shape of the active site into the complementary shape of
the substrate
Classification of enzymes
Enzymes can be classified according to the type of reaction
they catalyze:
1. Hydrolases;
Lipases – hydrolysis of an ester group
Nucleases – hydrolysis of a phosphate group
Proteases – hydrolysis of an amide group
2. Isomerases
Epimerases – isomerization of a stereogenic center
3. Ligases Carboxylases – Addition of CO2 Syntheses –
formation of a new bond
4. Oxidoreductases Ddehydrogenases – introducton of a double
bond by removal of hydrogen Oxidases – oxidation Reductases -
reduction
Classification of enzymes
5. Transferases
Kinases – transfer of a phosphate group
Transaminases – transfer of an amino group
Regulation of Enzyme Activities
The catalytic activities of many enzymes are regulated.
There are a number of ways through which enzyme
activities are regulated.
1. Feed back inhibition.
The enzyme that catalyzes the first step in a biosynthetic
pathway is usually inhibited by the ultimate product.
2. Enzymes are also regulated by regulatory proteins,
which can either stimulate of or inhibit their activities.
The activities of many enzymes are regulated by
calmodulin, a protein that serves as a calcium sensor in
nearly all eukaryotic cells.
Regulation of Enzyme Activities
3. Covalent modification.
Many enzymes are controlled by reversible attachment of
phosphoryl groups to specific serine and threonine residues
4. Proteolytic activation. Some enzymes are synthesized
in an inactive precursor form, which is activated at
physiological appropriate time and place.
Enzymes are stereoselective in the sense that they
catalyse only one enantiomer of a chiral substrate.
This is because enzymes are polymers of L-amino acids
hence they are enantiomerically pure chiral molecules.
Enzyme Inhibition
Enzymes can be inhibited by specific molecules and ions.
This is important because it serves as a major control mechanism
in biological stems. Also many drugs and toxic agents act by
inhibiting enzymes.
Furthermore, inhibition can be a source of insight into the
mechanism of enzyme action.
Enzyme inhibition can be either reversible or irreversible.
a. An irreversible inhibitor dissociates/detaches very slowly from
its target enzyme because it becomes very tightly bound to the
enzyme, either covalently or non-covalently.
b. Reversible inhibition is characterized by a rapid dissociation of
the enzyme-inhibitor complex.
Reversible inhibition could be competitive inhibition or
noncompetitive inhibition
Enzyme Inhibition cont..
Competitive inhibition: the enzyme can bind substrate
(forming an ES complex) or inhibitor (EI) but not both.
Many competitive inhibitors resemble the substrate and
bind to the active site of the enzyme.
The substrate is thereby prevented from binding to the
same active site.
A competitive inhibitor diminishes the rate of catalysis by
reducing the proportion of enzyme molecules bound to a
substrate.
Competitive inhibition can be overcome by increasing the
concentration of the substrate
Enzyme Inhibition cont..
Noncompetitive inhibition, which is also reversible, the
inhibitor and substrate can bind simultaneously to an
enzyme molecule.
Hence their binding sites do not overlap.
A noncompetitive inhibitor acts by decreasing the
turnover number rather that by diminishing the proportion
of enzyme molecules that are bound to the substrate.
Noncompetitive inhibition, in contrast with competitive
inhibition, cannot be overcome by increasing the
substrate concentration
Factors that affects enzyme activity
The are several factors that affects the enzyme catalyse
reaction, these are following:
Substrate concentration:
As the substrate concentration increases, the enzyme
reaction rate increases. However, at very high substrate
concentrations, the enzymes become saturated with
substrate and a higher concentration of substrate does not
increase the reaction rate.
Enzyme concentration: Assuming a sufficient concentration
of substrate is available, increasing enzyme concentration
will increase the enzyme reaction rate.
Factors that affects enzyme activity cont.
pH of the medium: Each enzyme has an optimal pH
that helps maintain its three-dimensional shape.
Changes in pH may denature enzymes by altering
the enzyme's charge.
Temperature of the medium
Effects of inhibitors on enzyme activity (Competitive
inhibition and Non-Copetitive Inhibitors)
Practice questions
1. With examples, discuss on the specificity of enzymes
2. What would happen to a biochemical reaction and life in the absence of
biological enzyme? Give a reason to your answer.
3. What kind of bonds hold enzymes-substrate together?
4. Define active site.
5. Discuss two models of enzyme inhibition
6. What is multienzyme system?
7. How is enzyme activity measured?
8. How does pH affect enzyme activity?
9. Describe allosteric regulation.
