Enzyme Catalysis

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Enzyme

Catalysis

CHM-203
What are enzymes?

Structure and chemical nature of enzymes

Enzyme nomenclature
What are we
Mechanism of enzyme catalysis
going to
cover? Factors affecting enzyme activity

Enzyme activity – kinetics

Enzyme control
Enzymes

+ Enzymes are protein-


based biocatalysts.
+ They are efficient
+ They are potent
+ They are specific
Enzymes

Proteins Cofactors Enzymes


Cofactors

Prosthetic
Metal Ions Coenzymes Groups
Metal Ions

+ Those enzymes with metal ions


bound to them are called
metalloenzymes
+ Zinc is usually seen in
carboxypeptidase, carbonic
anhydrase, alcohol
dehydrogenase, etc.
+ Copper is usually seen in enzymes
acting on molecular oxygen.
+ Other metal ions seen are Fe, Mn,
Mo, Co
+ These ions bind to sites where
cysteine sulfhydryl groups or
imidazole groups on histidine are
seen
Prosthetic
Groups
+ Prosthetic groups are
those groups which
give special chemical
functions to the
enzyme.
+ These functions would
not be present if they
were attached as a
side-chain
Coenzymes

+ Coenzymes are molecules


that temporarily attach
themselves to the enzyme,
change throughout the
course of the catalysis and
get regenerated
+ Active carrier molecules
usually act as coenzymes for
multiple different enzymes
+ Vitamins are responsible for
the synthesis of coenzymes
Enzyme Nomenclature

+ Oxidoreductases catalyse oxidation and reduction reactions


+ Transferases transfer functional groups
+ Hydrolases catalyse hydrolysis reactions
+ Lyases catalyse breaking of bonds without using water
+ Isomerases catalyse isomerisation reactions
+ Ligases catalyse the association of two molecules
Enzymes are biocatalysts.

A lot of biochemical reactions are


Enzyme thermodynamically unstable.

Activity
For every reaction to occur, the
Mechanism activation energy barrier ought to
be crossed.

This activation barrier can be


crossed by – thermodynamic and
catalytic processes
Activation
Energy
+ For every reaction, there is a
specific activation energy (𝐸𝐴 ),
which is the minimum amount
of energy that reactants must
have before collisions between
them will be successful in giving
rise to products.
+ More specifically, reactants
need to reach an intermediate
chemical stage called the
transition state, which has a free
energy higher than that of the
initial reactants.
Activation
Energy
+ For most biologically important
reactions at normal cellular
temperatures, the activation
energy is sufficiently high that
the proportion of molecules
possessing that much energy at
any instant is extremely small.
+ However, reactants even in
thermodynamically favourable
reactions are also stable.
+ This state is called a metastable
state.
Why thermodynamic method is
unfavourable?
+ The only way a reaction involving metastable reactants will
proceed at an appreciable rate is to increase the proportion
of molecules with sufficient energy.
+ This can be achieved either by increasing the average energy
content of all molecules or by lowering the activation energy
requirement.
+ Cells are isothermal systems and increasing temperature is
not beneficial since biological processes require a stable,
constant temperature.
Catalytic
Method
+ By using a catalyst, we can reduce the
activation energy.
+ That is, interactions of reactants can be
more favoured.
+ For this, reactant molecules need to be
bound to a surface.
+ A catalyst acts by forming transient,
reversible complexes with substrate
molecules, binding them in a manner that
facilitates their interaction and stabilizes
the intermediate transition state.
+ A catalyst changes only the rate at which
equilibrium is achieved; it has no effect
on the position of the equilibrium.
Every enzyme contains a characteristic cluster of amino
acids that form the active site where the substrates
bind, and the catalytic event occurs.

Usually, the active site is an actual groove or pocket


How do with chemical and structural properties that
accommodate the intended substrate or substrates
enzymes with high specificity.

catalyse Once in the active site, the substrate molecules are


bound to the enzyme surface in just the right
orientation so that specific catalytic groups on the
reactions? enzyme can facilitate the reaction

A more refined view of the enzyme-substrate


interaction is provided by the induced-fit model.
Induced Fit
Model
+ Unlike the lock and key model,
here substrate-enzyme
interactions are more flexible
+ It involves a conformational
change in the shape of the
enzyme molecule following
substrate binding.
+ This alters the configuration of
the active site and positions the
proper reactive groups of the
enzyme optimally for the
catalytic reaction.
Enzyme
Catalysis
+ E.g., action of sucrase

+ The initial random collision of a substrate molecule—


sucrose, in this case—with the active site results in its
binding to amino acid residues that are strategically
positioned there

+ Substrate binding induces a change in the enzyme


conformation that tightens the fit between the substrate
molecule and the active site and lowers the free energy of
the transition state.

+ This facilitates the conversion of substrate into products—


glucose and fructose, in this case.

+ The products are then released from the active site,


enabling the enzyme molecule to return to its original
conformation, with the active site now available for
another molecule of substrate

+ This entire sequence of events takes place in a sufficiently


short time to allow hundreds or even thousands of such
reactions to occur per second at the active site of a single
enzyme molecule
Factors affecting enzyme action

Substrate
Temperature pH
concentration

Presence of
inhibitors
Temperature

+ At low temperatures, the rate of


an enzyme-catalyzed reaction
increases with temperature.
+ At some point, however, further
increases in temperature result
in the denaturation of the
enzyme molecule.
+ The temperature range over
which an enzyme denatures
varies from enzyme to enzyme
and especially from organism to
organism
pH

+ Most enzymes are


active only in a 3-4 pH
unit range.
+ This pH dependence is
usually due to the
presence of one or
more charged amino
acids at the active site
and/or on the substrate
itself.
Substrate
Concentration
+ Most enzymes display
Michaelis-Menten
kinetics.
Understanding Michaelis-Menten
Kinetics
+ At very low substrate concentration, rate increases linearly with respect to
concentration
+ At very high substrate concentration, the rate is close to the maximum
velocity.
+ 𝐾𝑚 is the substrate concentration at a rate half of that of the maximum
velocity
+ 𝐾𝑚 also gives the affinity towards a substrate. Low 𝐾𝑚 implies high affinity
Enzyme inhibitors

+ Enzymes show diminishing catalytic properties in the


presence of a wide range of molecules called enzyme
inhibitors
+ They are usually heavy metal ions (Hg, Pb, Cu)
+ They may be enzyme-specific or non-specific as well
Enzyme Inhibition

Non-
Competitive Irreversible
competitive
Competitive
Inhibition
+ This occurs when the inhibitor
mimics the substrate molecule
and binds to the active site of
the enzyme in a reversible
manner.
+ This reduces enzyme activity
because many of the active sites
of the enzyme molecules are
blocked by bound inhibitor
molecules and thus cannot bind
substrate molecules at the
active site
Non-competitive
inhibition
+ A noncompetitive inhibitor, on
the other hand, binds to the
enzyme surface at a location
other than the active site.
+ It does not block substrate
binding directly but inhibits
enzyme activity indirectly by
causing a change in protein
conformation that can either
inhibit substrate binding to the
active site or greatly reduce the
catalytic activity at the active
site.
Irreversible
Inhibition
+ An irreversible inhibitor
binds to the enzyme
covalently, causing
permanent loss of
catalytic activity.
+ Some irreversible
inhibitors of enzymes
can be used as
therapeutic agents
Enzyme Control

Feedback inhibition

Allosteric regulation

Reversible covalent modification

Irreversible covalent modification


+ Feedback inhibition is one of the most common
Feedback mechanisms used by cells to ensure that the activities of
reaction sequences are adjusted to cellular needs.

Inhibition + The enzyme is inactivated when enough concentration of


the final product synthesized ahead in the metabolic
pathway
Allosteric
Regulation
+ It is a kind of feedback inhibition present
in enzymes that can exist in two different
forms.
+ In one of the two forms, the enzyme has a
high affinity for its substrate(s), leading to
high activity. In the other form, it has little
or no affinity for its substrate, giving little
or no catalytic activity. Enzymes with this
property are called allosteric enzymes.
+ Whether the active or inactive form of an
allosteric enzyme is favored depends on
the cellular concentration of the
appropriate regulatory substance, called
an allosteric effector.
+ An effector may be either an allosteric
inhibitor or an allosteric activator,
depending on the effect it has.
Allosteric Regulation
Reversible Covalent
Modification
+ Phosphorylation and
dephosphorylation are
examples of reversible
covalent modification
Irreversible Covalent
Modification

+ Proteolytic cleavage is an
example of irreversible covalent
modification.
+ This is usually seen in digestive
tract enzymes.
+ They are usually released as
inactive precursors called
zymogens since in their active
state, they are harmful where
they are released
+ When they have to be activated,
certain polypeptide chains are
cleaved to activate the enzyme.

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