1 / 18

CHMI 2227E Biochemistry I

CHMI 2227E Biochemistry I. Enzymes: Inhibition. Enzyme inhibition. Enzyme inhibitors inactivate the enzyme; Two main types of inhibition exist: Reversible enzyme inhibition : enzyme activity can be recovered by removing the inhibitor (e.g. dialysis, gel filtration);

licia
Télécharger la présentation

CHMI 2227E Biochemistry I

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript


  1. CHMI 2227EBiochemistry I Enzymes: Inhibition CHMI 2227 - E.R. Gauthier, Ph.D.

  2. Enzyme inhibition • Enzyme inhibitors inactivate the enzyme; • Two main types of inhibition exist: • Reversible enzyme inhibition: enzyme activity can be recovered by removing the inhibitor (e.g. dialysis, gel filtration); • Irreversible enzyme inhibition: inhibitor binds covalently to enzyme, which is then irreversibly inactivated. • The inhibition (i.e. inactivation) of an enzyme can tell us a lot about the way it works; • Enzyme inhibitors are frequently used to define biological phenomena; • Enzyme inhibitors are also sought by the big pharma to block enzymes involved in diseases; CHMI 2227 - E.R. Gauthier, Ph.D.

  3. VEGF (Vascular Endothelial Growth Factor): Produced in embryos and tumours; Acts via a cell surface receptor to trigger the growth of blood vessels; Why inhibit VEGF-R: Blocking the action of VEGF (an enzyme) will block the growth of blood vessels and starve tumours to death! VEGF-R VEGF Endothelial cell growth/migration ZD6474 Endothelial cell Enzyme inhibitionExample 1 - VEGF Receptor inhibitors: CHMI 2227 - E.R. Gauthier, Ph.D. British Journal of Cancer (2005) 92(Suppl 1), S6 – S1

  4. Sildenafil: cGMP-Phosphodiesterase inhibitor; Initially tested as an anti-hypertension drug; Vascular smooth muscle cell Endothelial cell Acetylcholine GMP PDE Muscle relaxation Arginine Blood vessel Dilation Sildenafil Nitric Oxide Synthase (NOS) cGMP GTP Guanylate cyclase NO NO Enzyme inhibitionExample 2 – Sildenafil: Int. J. Impot. Res. (2004) 16, S11–S14 CHMI 2227 - E.R. Gauthier, Ph.D.

  5. Enzyme inhibitionExample 3 – Acetaminophen (tylenol): CHMI 2227 - E.R. Gauthier, Ph.D.

  6. P Trypsin inhibitor Reversible Enzyme inhibition1- Competitive inhibition • Most frequently encountered inhibitors; • I is very similar to S (i.e. it is a structural analog) • I and S compete for the same binding site on the enzyme: the active site; • Vmax stays the same: • At high enough [S], S will outcompete I • Km is increased (Kmapp): • Because I can bind E, the amount of S required to reach ½ Vmax will be increased. CHMI 2227 - E.R. Gauthier, Ph.D.

  7. Reversible Enzyme inhibition1- Competitive inhibition • The value of Kmapp can be used to obtain Km and Ki (the dissociation constant for the inhibitor): • Kmapp = Km (1 + [I]/Ki) • Ki = [E][I]/[EI] • Ki is a measure of the affinity of I for E: the smaller Ki, the more potent the inhibition. CHMI 2227 - E.R. Gauthier, Ph.D.

  8. Reversible Enzyme inhibition2- Uncompetitive inhibition • I only bind to ES, not the free enzyme; • Example: glycophosphate (Round-up herbicide) • Vmax is decreased: • Some of the E is converted into an inactive ESI complex. • Km is decreased: • I reduces the amount of E that can participate in the reaction; • ESI shifts the E + S ES to the right, leading to an apparent decrease in Km. CHMI 2227 - E.R. Gauthier, Ph.D.

  9. Reversible Enzyme inhibition2- Uncompetitive inhibition • Vmaxapp= Vmax / (1 + [I]/Ki) • Kmapp= Km / (1 + [I]/Ki) CHMI 2227 - E.R. Gauthier, Ph.D.

  10. Reversible Enzyme inhibition3- Noncompetitive inhibition • I and S bind to different sites on E; • Binding of I on E doesn’t affect the binding of S on E (and vice versa); • So: Km is unchanged, but Vmax is decreased (I reduces the [E] that can generate P); • E.g. deoxycyclin (an antibiotic), which inhibits collagenase (a proteolytic enzyme involved in periodontal diseases). CHMI 2227 - E.R. Gauthier, Ph.D.

  11. Reversible Enzyme inhibition3- Noncompetitive inhibition • Vmaxapp= Vmax / (1 + [I]/Ki) CHMI 2227 - E.R. Gauthier, Ph.D.

  12. Irreversible enzyme inhibition • Irreversible inhibitors bind covalently to the enzyme and permanently inhibit it. • Very useful to identify the amino acids involved in catalysis • Three types: • Group-specific • Active site-directed reagents (aka Affinity labels) • Suicide inhibitors CHMI 2227 - E.R. Gauthier, Ph.D.

  13. React with amino acid side chains; Lead to inhibition by interfering with the catalysis (e.g. by reacting with side-chains important for the catalysis); E.g. diisopropyl fluorophosphate (DFP); Nerve gas Inhibits acetylcholine esterase (and many other proteases with Ser at the active site) Irreversible enzyme inhibition1. Group-specific inhibitors CHMI 2227 - E.R. Gauthier, Ph.D.

  14. Irreversible enzyme inhibition2. Affinity labels • Inhibitor is structurally similar to S; • Reacts with active site residues; • I reacts with E to form a covalent bond that cannot be hydrolysed; CHMI 2227 - E.R. Gauthier, Ph.D.

  15. Modified substrates; Initially processed by E as if it were the normal S; However, an reaction intermediate covalently and irreversibly binds the E, leading to its inhibition; Example 1: monoamine oxidase (MAO) inhibitors (MAO – breaks down certain neurotransmitters, e.g. serotonine, adrenaline)  high MAO activity = depression; Irreversible enzyme inhibition3. Suicide inhibitors CHMI 2227 - E.R. Gauthier, Ph.D.

  16. Sugars Tetrapeptide pentaGly bridges Pen Structure of the bacterial cell wall Irreversible enzyme inhibition3. Suicide inhibitors - penicillin • Interfere with the synthesis of the bacterial cell wall • Makes bacteria much less resistant to stress; • Cell wall: • Peptidoglycan • Penicillin blocks the formation of the link between the tetrapeptide and the pentaGly bridge; CHMI 2227 - E.R. Gauthier, Ph.D.

  17. Glycopeptide transpeptidase Glycopeptide transpeptidase pentaGly bridge Penicillin Tetrapeptide Glycopeptide transpeptidase Irreversible enzyme inhibition3. Suicide inhibitors - penicillin CHMI 2227 - E.R. Gauthier, Ph.D.

  18. Irreversible enzyme inhibition3. Suicide inhibitors - penicillin CHMI 2227 - E.R. Gauthier, Ph.D.

More Related