Irreversible Inhibition

1. Irreversible
Inhibition
Sonal Meshram

2. Irreversible
Inhibition :
Introduction
It inactivates an enzyme by bonding covalently to a
particular group at the active site. The inhibitor-enzyme
bond is so strong that the inhibition cannot be reversed by
the addition of excess substrate. The nerve gases,
especially Diisopropyl fluorophosphate (DIFP),
irreversibly inhibit biological systems by forming an
enzyme-inhibitor complex with a specific OH group of
serine situated at the active sites of certain enzymes. The
peptidases trypsin and chymotrypsin contain serine groups
at the active site and are inhibited by DIFP.

3. Irreversible inhibitors are covalently or noncovalently bound to the target
enzyme and dissociates very slowly from the enzyme. There are three types
of irreversible inhibitors:
Group specific
reagents
Affinity labels
Suicide
inhibitors

4. Group
specific
reagents
react with specific amino acid side chains like
diisopropylphosphofluoridate (DIPF) and iodoacetamide.
For example, only 1 of the 28 serine residues in chymotrypsin is
modified by DIPF. This means that this specific residue is especially
reactive; moreover, it is implied that this specific residue lies in the
active site of the enzyme chymotrypsin.
DIPF has also provided data that suggests, through its binding with
active serine residues, that there is indeed a reactive serine residue
contained within the active site of the enzyme Acetylcholinesterase.
The inactivating functionality of DIPF and similarly-shaped molecules
in acetylcholinesterase is representative of a group of compounds,
known as nerve agents.

6. Affinity labels (Reactive substrate analogs)
They are structurally similar to the substrate that can covalently bind to the
active site and are therefore more specific than group specific reagents.
An example is Tosyl-L-phenylalanine chloromethyl ketone (TPCK) which is an
analog for chymotrypsin which binds to the active site and reacts irreversibly
with the histidine residue to inhibit the enzyme.
Another example is bromoacetol phosphate, which mimics dihydroxy acetone
phosphate (DHAP) and binds covalently to the active site of triose phophate
isomerase and then modifying the enzyme so it becomes irreversibly inhibited.

8. Suicide
inhibitors
(Mechanism-
based
inhibitors)
It binds to the enzyme as a substrate and is processed by a normal
catalytic mechanism that generates a chemically reactive
intermediate that inactivates the enzyme through covalent
modification.
An example of a mechanism-based inhibitor can be seen through
the inhibitory power of the planar proline intermediate formed
during proline racemization. During this process, a trigonal
intermediate is formed, and the formation of the racemase is
inhibited because the tetrahedral intermediate necessary for
formation of the product is not formed.
The isomerization of proline through the planar transition state
underscores the essence of transition-state analogs as potent
inhibitors of enzymes.

9. Clinical
Importance of
Suicide
Inhibitors
• Aspirin:
Anti-inflammatory action Aspirin acetylates a
serine residue in the active centre of
cyclooxygenase thus inhibiting the PG synthesis
and the inflammation subsides
• 5-fluorouracil:
Used in cancer therapy,
5-fluorouracil (5-Fu) is converted to
fluorodeoxyuridylate (Fdump) by the enzymes of
the salvage pathway.
Fdump so formed inhibits the enzyme thymidylate
synthase thus inhibiting nucleotide synthesis.

10. • Difluoromethylornithine (DFMO)
Ornithine decarboxylase (ODC) catalyzes the conversion of ornithine to putrescine which is necessary for polyamine synthesis
When the ODC in Trypanosoma is inhibited ; multiplication of the parasite is arrested.
Therefore, inhibitors of ODC enzyme such as difluoromethylornithine (DFMO) has been found to be effective against
trypanosomiasis (sleeping sickness).
DFMO is initially inert, but on binding with the enzyme, forms irreversible covalent complex with the co-enzyme (pyridoxal
phosphate) and the amino acid residues of the enzyme.
In mammalian cells, the turnover rate of ODC is very high, and so the inhibition by DFMO is only transient. So, DFMO kills the
parasites with no side effects to the patient.
• Allopurinol:
The best example of suicide inhibition.
The drug is used in treatment of gout, as it inhibits the enzyme xanthine oxidase thus decreasing the uric acid formation.
allopurinol gets oxidized by the enzyme xanthine oxidase itself to form “alloxanthine” a more potent effective and stronger inhibitor
of xanthine oxidase thus potentiating the action of allopurinol

11. Thankyou