CHMI 2227E Biochemistry I

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

2. Enzymes • Enzymes: • Proteins • RNA • Catalysts: • Fully active enzyme is regenerated at the end of the reaction • Speed-up chemical reactions in cells by placing the substrate in an environment that facilitates the reaction • So: proper 3D structure (tertiary and, when applicable, quaternary) is ABSOLUTELY ESSENTIAL for the function of enzymes; CHMI 2227 - E.R. Gauthier, Ph.D.

3. Very powerful catalysts; Very specific for their substrate; Can distinguish between enantiomers Some enzymes require additional chemicals or groups to function: Metal ions Prosthetic groups: Heme Co-factors ATP Enzymes CHMI 2227 - E.R. Gauthier, Ph.D.

4. 1. oxido-reduction: oxidoreductases 2. Transfer of chemical groups: transferases 3. Hydrolysis: hydrolases 4. Removal of chemical groups: lyases 5. Isomerisation: isomerases 6.Linking two groups together: ligases Classes of enzymes • Only 6 types of chemical reactions are catalyzed by enzymes: • Enzymes are classified according to the type of reaction they catalyse: • each enzyme is given a systematic name and a IUBMB number (EC XXXX) • IUMBM = International Union of Biochemistry and Molecular Biology • The complete list of enzymes can be found at the following website: www.chem.qmul.ac.uk/iubmb/enzyme/ CHMI 2227 - E.R. Gauthier, Ph.D.

5. Example: ATP: D-glucose-6 phosphotransferase (aka hexokinase) EC 2.7.1.1 2 = transferase 7 = phosphotransferase 1 = acceptor group is OH 1 = glucose binds the phosphate group ATP + D-glucose ADP + D-glucose-6-phosphate IUBMB enzyme nomenclature CHMI 2227 - E.R. Gauthier, Ph.D.

6. 1- Oxidoreductases 2- Transferases 3- Hydrolases 4- Lyases 5- Isomerases 6- Ligases Classes of enzymes CHMI 2227 - E.R. Gauthier, Ph.D.

7. A- + B A + B- A-B + C A + B-C A-B + H2O A-H + B-OH Y X X Y Y X A=B + X-Y A-B A-B A-B A + B A-B Classes of enzymes • 1- Oxidoreductases • 2- Transferases • 3- Hydrolases • 4- Lyases • 5- Isomerases • 6- Ligases CHMI 2227 - E.R. Gauthier, Ph.D.

8. OxidoreductasesExample: Lactate dehydrogenase (EC 1.1.1.27) • Lactate = substrate • Pyruvate = product • NAD+/NADH = co-factor CHMI 2227 - E.R. Gauthier, Ph.D.

9. 2. TransferasesExample: Alanine transaminase (EC 2.6.1.2) • Alanine / a-ketoglutarate = substrates • Pyruvate / L-glutamate = products CHMI 2227 - E.R. Gauthier, Ph.D.

10. 3. HydrolasesExample: diphosphate phosphohydrolase(EC 3.6.1.1) • Pyrophosphate / H2O = substrates • Phosphate = product CHMI 2227 - E.R. Gauthier, Ph.D.

11. 4. Lyases (aka synthases)Example: Pyruvate decarboxylase(EC 4.1.1.1) • Pyruvate = substrate • Acetaldehyde and CO2 = products CHMI 2227 - E.R. Gauthier, Ph.D.

12. 5. IsomerasesExample: Alanine racemase(EC 5.1.1.1) • L-alanine = substrate • D-alanine = product CHMI 2227 - E.R. Gauthier, Ph.D.

13. 6. Ligases (aka synthetases)Example: L-glutamine synthetase(EC 6.3.1.2) • L-glutamate / NH4+ = substrates • L-glutamine = product • ATP = co-factor CHMI 2227 - E.R. Gauthier, Ph.D.

14. How do enzymes work? • Enzymatic reactions take place in multiple steps involving reaction intermediates: E + S ES EP E + P • Note that, while in theory these reactions are reversible, in practice, the low levels of the one of the reactants (S or P) usually pushes the equilibrium in one direction; • Enzymes increase the rate of chemical reactions, but do NOT alter the direction of the equilibrium. CHMI 2227 - E.R. Gauthier, Ph.D.

15. In chemical reactions, three conditions must be met for a reaction to take place: 1. the molecules must collide to react.If two molecules simply collide, however, they will not always react; therefore, the occurrence of a collision is not enough. 2. there must be enough energy (energy of activation) for the two molecules to react. If two slow molecules collide, they might bounce off one another because they do not contain enough energy to reach the energy of activation and overcome the transition state (the highest energy point). 3.the molecules must be orientated with respect to each other correctly. Enzymes and the energy barrier CHMI 2227 - E.R. Gauthier, Ph.D.

16. The transition state is not a reaction intermediate: it is a transitory molecular structure that is no longer the substrate, but not yet the product. G = Gibbs free energy the ability of the molecule to react The greater the free energy, the more unstable the molecule is. DG = Gproduct – Gsubstrate: If DG > 0 = reaction does not occur spontaneously (because S is more stable that P) If DG < 0 = reaction occurs spontaneously (because P is more stable than S) In other words: the more negative the DG, the more likely the reaction will take place; DGŧ DG Enzymes and the energy barrier CHMI 2227 - E.R. Gauthier, Ph.D.

17. Enzymes and the energy barrier • In the absence of an enzyme (or catalyst), the substrate (here the metal stick) requires a substantial amount of energy in order to reach the activation state and react; CHMI 2227 - E.R. Gauthier, Ph.D.

18. Enzymes and the energy barrier • In the presence of an enzyme, the reaction is facilitated because the enzyme provides a better environment for the reaction to occur: • Close proximity of substrate and chemical groups of the enzyme • Proper orientation of the chemical groups with respect to the substrate • The formation of the transition state is favoured; • This results in a lowering of the activation energy required for the reaction to occur. CHMI 2227 - E.R. Gauthier, Ph.D.

19. The « lock and key » myth… • The « lock and key » thing is a MYTH: • If the enzyme and substrate were perfectly complementary, like a lock and key, the interaction between E and S would be so stable that the reaction would not occur! CHMI 2227 - E.R. Gauthier, Ph.D.

20. The « lock and key » myth… • Instead, the 3D shape of the enzyme is complementary to the transition state; • by doing so, the enzymes favours the formation of the transition state, lowers the energy of activation, and accelerates the reaction…COOL! CHMI 2227 - E.R. Gauthier, Ph.D.