Competitive inhibitors

What is Competitive Inhibition?

❖ Enzymes experience competitive inhibition when any molecule which is not the enzyme's natural substrate
binds to the enzyme's active site, preventing the natural substrate from binding and thereby inhibiting
enzyme activity.
❖ To determine the effectiveness of the inhibitor, the ratio of a competitive inhibitor's affinity for an enzyme's
active site is compared to the natural substrate's affinity for the active site.
❖ Example of competitive inhibitor:
✓ An example of a use for a competitive inhibitor is in the treatment of influenza via the neuraminidase
inhibitor, Relenza.
✓ Competitive inhibitors such as 5-fluororacil or Sulfa drugs can be toxic.
Kinetics of Competitive enzyme inhibition
 Effect on Km: For a specified
substrate, a competitive inhibitor
rises the value of km. This means
that more substrate is needed in the
presence of a competitive inhibitor
to achieve 1⁄2Vmax.

Effect on Vmax: Increasing

substrate concentration [S] reverses
the impact of a competitive
inhibitor. The reaction velocity
reaches the Vmax observed in the
absence of inhibitors at a
(A) Plot of velocity versus substrate concentration. sufficiently elevated concentration
(B) Mechanism of competitive inhibition. of the substrate.
 Competitive inhibitors cause an apparent increase in Km, without changing Vmax

An enzyme can be inhibited competitively by substances that are similar in chemical structure to the substrate.
These compounds bind in the active site and compete with substrate for the active site of the enzyme; they cause
an apparent increase in Km, but no change in Vmax (Fig. 6.8). The inhibition is not the result of an effect on
enzyme activity, but on substrate access to the active site. The reaction scheme for competitive inhibition is:

Effect on the Lineweaver-Burk

plot: Competitive inhibition
shows a characteristic
Lineweaver-Burk plot in which
the plots of the inhibited and
uninhibited reactions intersect on
the y-axis at 1/Vmax (Vmax is
unchanged), and show different
x-axis intercepts, indicating that
the apparent Km is increased in
the presence of the competitive
(C) Lineweaver–Burk plot in the presence of a competitive inhibitor. inhibitor.
(D) Eadie–Hofstee plot in the presence of a competitive inhibitor.
K′m is the apparent Km in presence of inhibitor.
 In the presence of a competitive inhibitor, Michaelis-Menten equation-
• The Michaelis-Menten equation will be changed a little bit for this enzyme
inhibition process.

Let’s see what changes will take place o The inhibition constant (Ki) is the dissociation constant of the
in the equation. enzyme–inhibitor complex (EI) and
✓ the lower the Ki, the more efficient the inhibition of enzyme
activity.
✓ The smaller the Ki, the smaller amount of medication needed
to inhibit the activity of that enzyme.

o Regardless of the Ki, however, the rate of the enzyme-

catalyzed reaction in the presence of a competitive inhibitor
can be increased by increasing the substrate concentration,
since substrate, at higher concentration, competes more
effectively with the inhibitor.
The inhibitor constant, Ki, is an indication of how potent an inhibitor is; it is the concentration required to produce
half maximum inhibition. The value Ki is the dissociation constant describing the binding affinity between the
inhibitor and the enzyme.
 In the presence of a competitive inhibitor, Lineweaver-Burk equation

• This is the Lineweaver-Burk equation

in the presence of a competitive
inhibitor.
• In the competitive inhibition, the value
of Km changes while Vmax does not.
There is a dramatic shift of Km
towards the right side that means the
•
Km is higher than the normal Km.
• For any positive values of alpha, the
slope is steeper that means km has
changed.
✓ With the increase in
alpha value that line
would get steeper and
steeper, and the degree
of inhibition would be
stronger and stronger
because km would
increase and the
affinity for the substrate
towards enzyme would
decrease.
 Example of competitive enzyme inhibition

1. Malonate is a competitive inhibitor of succinate.

2. Donepezil competitively inhibits acetylcholinesterase
3. Methotrexate is a competitive inhibitor of DHFR
4. Competitive inhibitor, 5-Fluorouracil (5-FU) is a widely used chemotherapeutic agent
5. Sulfonamides are competitive inhibitors of Dihydropteroate Synthase
6. Ethanol is used to treat methanol toxicity
 Malonate is a competitive inhibitor of succinate.

✓ Succinate is converted into fumarate by succinate dehydrogenase.

✓ Malonate which has a similar structure like succinate can competitively inhibit succinate dehydrogenase and as
a result no product is formed. that's how malonate act like a competitive inhibitor of succinate.

How we can reverse the effect of

competitive inhibition

We can increase the substrate

concentration and if we increase the
substrate concentration the probability
that the substrate would encounter the
enzymes active site is far more than
the probability of interaction between
inhibitor and substrate and that's how
the effect of these competitive inhibitor
can be reversed and that's how the we
can get rid of the inhibition.
Succinate dehydrogenase is an enzyme that is present in the inner mitochondrial membrane. It catalyzes the
oxidation of succinate to fumarate. It is a conjugated protein as it has flavoprotein which has FMN or FAD
molecule as a prosthetic group. Malonate acts as an inhibitor that competes with the succinate and prevents its
attachment to the enzyme.

Figure: Competitive Inhibition.

(a) Succinate binds to the enzyme

succinate dehydrogenase. A
dehydrogenation reaction
occurs, and the product—
fumarate—is released from
the enzyme.

(b) Malonate also binds to the

active site of succinate
dehydrogenase. In this case,
however, no subsequent
reaction occurs while malonate
remains bound to the enzyme.
✓ According to the ACh hypothesis, a reduction in brain ACh levels is thought to be the cause of AD. AChE is an
enzyme that breaks down ACh and makes it inactive.
✓ Donepezil is given as a treatment for Alzheimer's disease. Donepezil competitively inhibits acetylcholinesterase
from degrading acetylcholine, causing acetylcholine levels to increase in the Alzheimer's patient's brain.
✓ Donepezil impedes AChE and prevents the breakdown of ACh. This elevates the concentration of free ACh in
the synaptic cleft, thereby activating cholinergic nerves and stopping the progression of cognitive failure
mechanisms in AD patients.

Neurotransmission of cholinergic synapse. In the synaptic cleft

Donepezil impedes AChE resulting in an increase of Ach
of cholinergic neuron, acetylcholine (ACh) is decomposed into
level in the synaptic cleft
choline and acetic acid by acetylcholinesterase (AChE) https://basicmedicalkey.com/the-practical-pharmacology-of-donepezil/
Methotrexate is a competitive inhibitor of DHFR ✓ When methotrexate is given,
it acts on DHFR (by mimicking
DHFR) which is responsible for
the formation of FH4

✓ If FH4 is not formed, then

N5N10-Methylene-FH4 is not
formed

✓ N5N10-Methylene-FH4 is
needed for the production of
dTMP (dUMP -> dTMP) so
without it no dTMP is formed

✓ dTMP is a DNA building block

and is needed for DNA
synthesis - therefore when
there is no dTMP produced,
there is inhibition of DNA
synthesis and inhibition of cell
division
✓ Methotrexate has a very
similar structure to the
substrate of DHFR (FH2) so
binds with a higher affinity
for DHFR
✓ Competitive inhibitor, 5-Fluorouracil (5-FU), a
widely used chemotherapeutic agent, is an
antimetabolic and antineoplastic agent.

✓ This uracil analog, which mimics both uracil and 5-FU is a nucleobase analog of uracil in which the
thymine, interferes with nucleoside metabolism by hydrogen at position 5 is replaced by fluorine. It is a
incorporating into RNA and DNA triggering pyrimidine antagonist that functions by inhibiting DNA
cytotoxicity and cell death. synthesis.

• 5-FU is a DNA/RNA synthesis inhibitor, which interrupts nucleotide synthetic by inhibiting thymidylate synthase
(TS) in tumor cells. It irreversibly inhibits TS leading to a dNTP imbalance resulting in DNA damage and cell
death.
• Fluorouracil induces apoptosis and can be used in the treatment of HIV.
https://www.invivogen.com/5-fluorouracil
 Mode of action of 5-FU:
✓ As with all pyrimidine antagonists, 5-FU is a pro-drug and
the intracellular conversion of 5-FU into 5-fluoro-uridine-
monophosphate (FUMP) and 5-fluoro-deoxy-uridine-
monophosphate (FdUMP) is essential for its action.
✓ The active metabolite 5-FdUMP is a inhibitor of
thymidylate synthase (TS) and hence of DNA synthesis
through deoxythymidine triphosphate (dTTP) deprivation
which in turn leads to apoptosis.
✓ As 5-FU is similar in shape but does not undergo the same
chemistry as uracil, it inhibits RNA replication enzymes,
thereby blocking RNA synthesis and stopping the growth
of cancerous cells.
✓ Hence, the antitumor activity results from the inhibition of TS by FdUMP, as well as from the incorporation of 5-FU
metabolites into RNA and DNA.
✓ Sulfonamides (also called sulfa drugs) are a group
of broad spectrum and action antibiotics derived
from sulfanilamide.
✓ The first class of antibiotics discovered, they stop the
growth and replication of bacteria by preventing
bacterial folic acid synthesis.
✓ Sulfa drugs inhibit bacteria from creating folic acid
production by inhibiting dihydropteroate synthase.
This causes bacteria to perish due to a lack of folic
Sulphanilamide and other sulfa drugs are structural
acid. analogs of para-aminobenzoic acid (PABA). So, sulfa
✓ Sulfonamides can be prescribed to patients with drugs act as competitive inhibitor and occupy the
active site of some bacterial enzyme catalyzing this
cystic fibrosis (CF), but their use has diminished with reaction. Thus, many sulfa drugs act as antibiotics.
newer and more effective antibiotics.
 Sulfonamides prevent bacterial infections in humans

✓ Many microorganisms like bacteria synthesize the vitamin

folic acid from para-aminobenzoic acid.
✓ When this reaction is affected, it blocks the folic acid
biosynthesis which is essential for the growth of
microorganisms, ultimately results in the death of the
microorganisms.
✓ This group of antibiotics are chemical analogues of p-
aminobenzoic acid (PABA) that interfere with folic acid
synthesis.
✓ They work by binding and blocking a specific enzyme,
dihydropteroate synthase (DHPS), that is essential for the
synthesis of folate, a vitamin bacteria — like all living
organisms — need to survive.
• Folic acids are necessary for the
bacterial protein synthesis and
synthesis of amino acids. Folic acid synthesis inhibitors.
• Trimethoprim and Sulfonamides
are the antimicrobial classes that
interfere with the synthesis of
folic acid at different levels and
are bacteriostatic.
• Trimethoprim and Sulfonamides
are used as combination therapy
in treating different infections
like Traveller’s Diarrhoea, UTI,
etc due to their synergistic
mechanisms.
• Sulfonamides are the widely
used antimicrobials.
• Trimethoprim prevents bacterial
DNA synthesis that inhibits -Bacteria have to synthesize folate using
bacterial dihydrofolate dihydropteroate synthase (only bacterial) and
reductase (DHFR). dihydrofolate reductase (DR) (bacteria and humans)
https://thebiologynotes.com/folic-acid-synthesis-inhibitors/
 Sulfonamides
✓ Sulfonamides are structural analogues of PABA (Para-AminoBenzoic Acid)
✓ Sulfonamides are competitive inhibitors of Dihydropteroate Synthase (DS).
✓ DS incorporates PABA into the Dihydropteroate acid.
✓ Dihydropteroic acid is an immediate precursor of Folic acid
✓ No dihydropteroaic acid = No Dihydrofolate acid -> No folic acid
synthesis

✓ Bacteria needs PABA for the biosynthesis of Folic acid

✓ Sulfa drugs closely resembles to PABA
✓ Sulfa drugs are used for the treatment of many bacterial infections .

✓ Sulfonamides substitute PABA that results in the blocking of

enzymes that are essential for the biosynthesis of metabolic
reactions for the RNA formation.
[Tetrahydrofolic acid is used in the synthesis of
✓ Inhibition of growth and replication occurs that cannot use
nucleic acid bases (DNA, RNA), some amino acids
dietary folate which shows bacteriostatic activity. and other metabolites]
-Bacteria have to synthesize folate using
dihydropteroate synthase (only bacterial) and
https://thebiologynotes.com/folic-acid-synthesis-inhibitors/
dihydrofolate reductase (DR) (bacteria and humans)
Trimethoprim
✓ Trimethoprim is bactericidal.
✓ It inhibits an enzyme dihydrofolate reductase (DR).
✓ Dihydrofolate reductase is the main enzyme that catalyzes
the formation of THF (Tetrahydrofolic acid).
✓ This drug binds with dihydrofolate reductase interfering
biosynthesis of nucleic acids and proteins resulting in
bacterial lysis.

✓ bacteria have to synthesize folate using dihydropteroate

synthase (only bacterial) and dihydrofolate reductase
(bacteria and humans)
✓ sulfonamides are competitive inhibitors of DS
✓ trimethoprim inhibits DR
[Tetrahydrofolic acid is used in the synthesis of
➢ nucleic acid bases (DNA, RNA),
➢ some amino acids and
➢ other metabolites]
https://thebiologynotes.com/folic-acid-synthesis-inhibitors/
 Ethanol is used to treat methanol toxicity because ethanol is a competitive inhibitor
of alcohol dehydrogenase

✓ Methanol is included in many home

chemicals, fluids, varnishes, stains and dyes.

✓ Toxicity results from its metabolism by

alcohol dehydrogenase (ADH) to formic acid,
which accumulates and results in metabolic
acidosis and organ injuries (Figure 1).

✓ Small ingested amounts as little as 10 mL of

pure methanol may be sufficient to cause
life-threatening toxicity and permanent
blindness.
Methanol toxicity is important to recognize because of its conversion to acid metabolites that can
cause a metabolic acidosis, blindness, and neurologic damage.

Fomepizole or ethanol serves as

alcohol dehydrogenase
inhibitors to stop the conversion
of methanol to its toxic
metabolite, formate.
Captopril is an angiotensin converting enzyme (ACE) inhibitors. ACE normally will convert angiotensin into
angiotensin II, a compound that works to increase blood pressure through mechanisms such as vasoconstriction, as
well as the inactivation of bradykinin (potent vasodilator).

https://www.stepwards.com/?page_id=1443
 Treatment with an inhibitor of angiotensin-converting enzyme (ACE)

A 50-year-old man was admitted to hospital suffering from general fatigue, a stiff shoulder, and headache. The
patient was 1.8 m tall and weighed 84 kg. His blood pressure was 196/98 mmHg (normal below 140/90 mmHg;
optimal below 120/80 mmHg) and his pulse was 74. He was diagnosed as hypertensive. The patient was given
captopril, an angiotensin-converting enzyme (ACE) inhibitor. After 5 days' treatment, his blood pressure returned to
near-normal levels.

Comment.

Renin in the kidney converts angiotensinogen into angiotensin I, which is then proteolytically cleaved to angiotensin II
by ACE. Angiotensin II increases renal fluid and electrolyte retention, contributing to hypertension. Inhibition of ACE
activity is therefore an important target for hypertension treatment. Captopril inhibits ACE competitively, decreasing
blood pressure.

• ideal blood pressure is considered to be between 90/60mmHg and 120/80mmHg.

• high blood pressure is considered to be 140/90mmHg or higher.
• low blood pressure is considered to be 90/60mmHg or lower.

The medical term for high blood pressure is hypertension. In adults, blood pressure is considered
to be normal under a systolic value of 140 mmHg and under a diastolic value of 90 mmHg.