Unit 4 - Enzymes 2 Flashcards
What are the two types of irreversible inhibition?
Affinity Labelling Agents or Reactive Substrate Analogues
- react directly with the enzyme active site
Mechanism Based Inhibitors or Suicide Inhibitors
- contain unreactive groups which are activated by the enzyme by chemical reaction and then react further with the enzyme active site to form covalent bonds to the enzyme.
What is a Mechanism Based Inhibitor also known as?
Suicide Inhibitor
What is a major cause of death in Europe and the USA?
Coronary Heart Disease
What percentage of deaths due to heart disease in USA are due to atherosclerosis?
50%
What is atherosclerosis?
Build up of fatty deposit on the inner walls of the arteries
What is most of the deposits in atherosclerosis due to?
Cholesterol
What is the rate limiting step of cholesterol synthesis?
Reduction of 3-hydroxy-3-methylglutaryl coenzyme A to mevalonic acid
- HMGCoA reductase
What was the first HIT compound that was screened when trying to find an inhibitor of HMGCoA reductase?
Mevastatin
How does compactin or mevastatin work?
Mimics the susbtrate 3-hydroxy-3-methylglutaryl coenzyme A
- HMGCoA
How do analogues of HMGCoA work?
Bind well to the enzyme active site as the enzyme stabilises these high energy intermediates as a way of increasing the rate of reaction
What are statins?
competitive inhibitors of HMGCoA reductase
How do statins bind HMGCoA?
Bind in two binding pockets in the enzyme active site
Describe the binding sites of HMGCoA
Hydroxymethylglutaryl binding pocket
Hydrophobic binding pocket
What is Ki?
Inhibition constant
What is Km?
Michaelis constant
Give some examples of currently used inhibitors of HMGCoA (statins)?
Atorvastatin
Fluvastatin
Pravastatin
Simvastatin
Why are statins so popular?
Cardiovascular disease is a major healthcare burden in affluent countries
Increasing obesity and high cholesterol levels as major risk factors
Patients and healthcare providers willing to pay for effective brand medicines
Statins are orally bioavailable
- normally a one-a-day tablet
Widespread uses
- most patients with cardiovascular disease including elderly
- diabetes patients > 40 years
- preventative for asymptomatic individuals at increased risk
How is atorvastatin metabolised?
Primary route of metabolism via CYP3A4 hydroxylation
What are the side effects of atorvastatin?
Serious side effects are rare, not however class association with muscle effects
- myopathy
- myositis
- rhabdomyolysis
Statins can also alter liver function
- hepatitis
- jaundice
Which enzyme gives rise to an increase in blood pressure?
Angiotensin converting enzyme, ACE
How do angiotensin converting enzymes cause an increase in blood pressure?
3 mechanisms
- good target for control of blood pressure
Angiotensin 2 causes arteriolar vasoconstriction increasing blood pressure
constriction of the efferent arterioles of the kidney leads to increased perfusion pressure in the glomeruli.
Stimulation by ATII of the adrenal cortex to release aldosterone, a hormone that acts on kidney tubules, causes sodium and chloride ions retention and potassium excretion. Sodium is a “water-holding” ion, so water is also retained, which leads to increased blood volume, hence an increase in blood pressure.
Where is angiotensinogen produced?
Liver
How is angiotensinogen converted to angiotensin I?
Renin
How is angiotensin I to angiotensin II?
Angiotensin converting enzyme, ACE
How does angiotensin converting enzyme increase blood pressure?
Angiotensin II is a hormone and a potent vasoconstrictor that increases blood pressure
Angiotensin II causes release of the hormone aldosterone which stimulates sodium/water retention and also increase blood pressure
ACE catalyses hydrolysis of bradykinin
- a potent vasodilator
- increasing blood pressure
What do proteases, such as ACE, usually have?
Binding pockets for each of the amino acids adjacent to the site of cleavage
N-terminal site are S1, S2, S3
C-terminal site are S1’, S2’, S3’
What was known about ACE when work started to find an inhibitor?
The enzyme was known to use Zn2+ ions
A series of tripeptides were known to be substrate
- following sequence being optimal
- aromatic amino acid-Ala-Pro
How does ACE work?
Cleaves the C-terminal 2 residues of the tripeptide substrate
Why is sulphur used in ACE inhibitors?
Binds strongly to zinc
What is the inhibitor constant, Ki, of captopril?
- 7nM
- very potent
What is the clinical use of captopril?
Effective on its own, 50%
Increased effectiveness with a diuretic, 90%
What are the side effects of captopril?
Rash and loss of taste
- possibly due to presence of sulphur
