Enzymes II

Question 1

1) What is the difference between competitive and non-competitive enzyme inhibitors?

Answer 1

• Competitive: block the enzyme active site so substrate cannot bind (e.g. malonate inhibition of succinate dehydrogenase)
• Non-competitive: interferes with the catalytic mechanism in some other way
  > reversible: inhibition of Mg2+, requiring an enzyme by addition of chelator
  > irreversible: organophosphorous inhibition of cholinesterase
  [some enzyme can display multiple properties]

Question 2

2) Define the term allosteric binding

Answer 2

Binding to another site of the enzyme (not the active site) in order to inhibit it
- produces inactive enzyme or enzyme-substrate complexes

Question 3

3) State one way in which competitive inhibitors can be overcome

Answer 3

Increasing substrate concentration

Question 4

4) What is the effect of competitive and non-competitive enzyme inhibitors on enzyme kinetic parameters (Vmax and Km)?

Answer 4

Competitive: Increase Km and same Vmax
- increased gradient of slope on LWB plot
Non-competitive: Same Km, lower Vmax
- decreased gradient of slope of LWB plot

Question 5

5) What is ACE (angiotensin converting enzyme) involved in and what are risks associated with high levels of ACE activity?

Answer 5

• involved in angiotensin production
• converts angiotensin I to angiotensin II which leads to peripheral vasoconstriction, increasing blood pressure
• if levels of A-II are high, this increases the risk of heart failure

Question 6

6) How do ACE inhibitors treat heart failure?

Answer 6

• inhibit formation of angiotensin II (by inhibiting ACE) and so reduces peripheral vasoconstriction so blood pressure decreases and there is a lower risk of heart failure

Question 7

7) Describe the action of Acetylcholine esterase inhibitor when reversible

Answer 7

• AChE-I binds to active enzyme
• EI (enzyme-inhibitor) complex formed
• groups on enzyme are transferred (deactivation)
• there is a slow hydrolysis of the inactive enzyme, as it reverts back to its active form

Question 8

8) Describe the action of Acetylcholine esterase inhibitor when irreversible

Answer 8

• AChE-I binds to active enzyme
• EI (enzyme-inhibitor) complex formed
• Enzyme is phosphorylated, no spontaneous hydrolysis occurs -> irreversible deactivation of enzyme

Question 9

9) Describe how the addition of a cure can reactivate this enzyme

Answer 9

• ‘Cure’ molecule interacts with enzyme, is more attractive than the inhibitor
• Allows enzyme to be reactivated as a phosphate group is transferred back and active enzyme is reformed

Question 10

10) State 3 ways of metabolic regulation of enzyme activity

Answer 10

• Allosteric binding sites (+ or - effectors)
• covalent modification by other enzymes -> phosphorylation by kinases or dephosphorylation by phosphorylases
• induction or repression of enzyme synthesis

Question 11

11) What curve do allosteric enzymes present?

Answer 11

• sigmoidal curve
• this shows low enzyme activity at low substrate concentration and an immediate, rapid increase in enzyme activity to Vmax, as [S] increases
• since this does not fit the michaelis menton equation, K0.5 is used instead of Km

Question 12

12) What is the difference between a positive and negative allosteric modulator and how does the MM plot change?

Answer 12

+ve: activator - lowers [S] required for Vmax
curve is above normal sigmoid
allows substrate to bind (changes enzyme shape to increase efficacy)
-ve: inhibitor - higher [S] needed for Vmax
curve is below normal sigmoid
prevents substrate from binding

Question 13

13) Define the two types of allosteric enzyme

Answer 13

1) homotrophic allosteric : multi-subunit enzymes, same binding site on each subunit functions as active and regulatory site
- the substrate/other molecule can be the effector
- binding of one substrate alters enzyme conformation and enhances binding of subsequent substrates
2) heterotrophic allosteric : the substrate is not the effector - a different molecule will bind to the regulatory site, the substrate can only bind at ONE site

Question 14

14) Give 2 examples of positive allosteric effectors and negative allosteric effectors

Answer 14

• Positive: Phosphoenolpyruvate (PEP) and F1,6BP on pyruvate kinase
• Negative: ATP and citrate on phosphofructokinase (while AMP is positive)

Question 15

15) Define covalent modification

Answer 15

Typically the addition or removal of phosphate from serine/threonin/tyrosine/histadine residues
- phosphorylation increases the activity of glycogen phosphorylase (degrades glycogen) and decreases the activity of glycogen synthase

Question 16

16) What are the potential effects of protein phosphorylation (3) and give 4 examples

Answer 16

• Increase in activity, decrease in activity, catalysis of a different reaction
• adenylylation - tyrosine residues
• uridylylation - tyrosine residues
• ADP-ribosylation - arginine, glutamine residues
• Methylation - glutamate residues, can alter gene expression

Question 17

17) Define proteolytic activation

Answer 17

• Some enzymes are synthesised as larger, inactive precursor forms that are called proenzymes or zymogens
• activation involves the irreversible hydrolysis of one or more peptide bonds to produce an active form