ENZYMES

1. ENZYMES

2. Definition Enzymes are protein molecules that catalyze the conversion of one or more substrates into one or more different products enhancing the rate of reaction without affecting equilibrium.

3. Many enzymes contain small non protein molecules and metal ions that participate directly in substrate binding or catalysis are termed “ Prosthetic groups , Cofactors , and Coenzymes”

4. Prosthetic groups:- Are tight , stable incorporation in the protein structure of enzyme by covalent or noncovalent bonding. Examples: FMN, FAD, Biotin, Co, Cu ,Mg ,Mn

5. Cofactors: Serve a function as prosthetic group but binds transient ,dissociable manner either to enzyme or to the substrate. Example: ATP

6. Coenzymes: • Serve as a recyclable shuttles that transport substrates from site of generation to the site of utilization. • Example : Coenzyme A

7. Enzyme classification and nomenclature • According to the International union Of Biochemistry an enzyme name has two parts: -First part is the name of the substrates for the enzyme. -Second part is the type of reaction catalyzed by the enzyme. This part ends with the suffix “ase”. Example: Lactate dehydrogenase

8. Six Classes 1. Oxidoreductases 2. Transferases 3. Hydrolases 4. Lyases 5. Isomerases 6. Ligases

9. 1. Oxidoreductase catalyze oxidation reduction reactions , also called oxidases, dehydrogenases, or reductases.

10. 2. Transferases • Transferases – catalyze group transfer reactions, excluding oxidoreductases (which transfer hydrogen or oxygen ) these are of the general form: A-X + B ↔ BX + A

11. 3. Hydrolases • Hydrolases – catalyze hydrolytic reactions. Includes lipases, esterases, peptidases/proteases. A-X + H2O ↔ X-OH + HA

12. 4. Lyases • Catalyze non-hydrolytic removal of functional groups from substrates, often creating a double bond in the product; or the reverse reaction, • Example : decarboxylases and aldolases

13. 5. Isomerases • Catalyzes isomerization reactions, including racemization and cis-tran isomerizations.

14. 6. Ligases • catalyzes the synthesis of various (mostly C-X) bonds, coupled with the breakdown of energy-containing substrates, usually ATP

15. 1. Structure and function of enzymes • Globular proteins acting as the body’s catalysts • Speed up time for reaction to reach equilibrium • Lower the activation energy of a reaction Example: LDH = Lactate dehydrogenase (enzyme) NADH2 = Nicotinamide adenosine dinucleotide (reducing agent & cofactor) Pyruvic acid = Substrate

16. Energy Energy Transition state New transition state Act. energy Act. energy Starting material Starting material ∆G ∆G Product Product WITH ENZYME WITHOUT ENZYME 1. Structure and function of enzymes Lowering the activation energy of reaction • Enzymes lower the activation energy of a reaction but DG remains the same

17. 1. Structure and function of enzymes Methods of enzyme catalysis • Provide a reaction surface (the active site) • Provide a suitable environment (hydrophobic) • Bring reactants together • Position reactants correctly for reaction • Weaken bonds in the reactants • Provide acid / base catalysis • Provide nucleophiles

18. Active site Active site ENZYME 2. The active site • Hydrophobic hollow or cleft on the enzyme surface • Accepts reactants (substrates and cofactors) • Contains amino acids which • - bind reactants (substrates and cofactors) • - catalyse the reaction

19. Substrate S 3. Substrate binding 3.1 Induced fit Induced fit • Active site is nearly the correct shape for the substrate • Binding alters the shape of the enzyme (induced fit) • Binding will strain bonds in the substrate • Binding involves intermolecular bonds between functional groups in the substrate and functional groups in the active site

20. S vdw interaction H-bond Active site ionic bond H Phe O Ser CO2 Asp Enzyme 3. Substrate binding 3.2 Bonding forces • Ionic • H-bonding • van der Waals • Example:

21. O H-Bond H Ionic bond H3N Possible interactions vdw-interactions H-Bond van der Waals Ionic 3. Substrate binding 3.2 Bonding forces • Ionic • H-bonding • van der Waals Example: Binding of pyruvic acid in LDH

22. Phe Phe S S H O H O Ser Ser CO2 Induced fit CO2 Asp Asp 3. Substrate binding 3.2 Bonding forces • Induced fit - Active site alters shape to maximise intermolecular bonding Intermolecular bond lengths optimised Susceptible bonds in substrate strained Susceptible bonds in substrate more easily broken Intermolecular bonds not optimum length for maximum bonding

23. O O O 3. Substrate binding Example: Binding of pyruvic acid in LDH O H H3N

24. pi bond weakened 3. Substrate binding Example: Binding of pyruvic acid in LDH O H H3N

25. L-Serine L-Cysteine 4. Catalysis mechanisms 4.1 Acid/base catalysis • Histidine Non-ionised Acts as a basic catalyst (proton 'sink') Ionised Acts as an acid catalyst (proton source) 4.2 Nucleophilic residues

26. 4. Catalysis mechanisms Serine acting as a nucleophile

27. E E P S S E + S P E E E ES EP E + P 5. Overall process of enzyme catalysis • Binding interactions must be; • - strong enough to hold the substrate sufficiently long for the reaction to occur • - weak enough to allow the product to depart • Implies a fine balance • Drug design - designing molecules with stronger binding interactions results in enzyme inhibitors which block the active site

28. S I I E E E 6. Competitive (reversible) inhibitors • Inhibitor binds reversibly to the active site • Intermolecular bonds are involved in binding • No reaction takes place on the inhibitor • Inhibition depends on the strength of inhibitor binding and inhibitor concentration • Substrate is blocked from the active site • Increasing substrate concentration reverses inhibition • Inhibitor likely to be similar in structure to the substrate

29. X X Covalent Bond OH OH O Irreversible inhibition 7. Non competitive (irreversible) inhibitors • Inhibitor binds irreversibly to the active site • Covalent bond formed between the drug and the enzyme • Substrate is blocked from the active site • Increasing substrate concentration does not reverse inhibition • Inhibitor likely to be similar in structure to the substrate

30. Active site unrecognisable Active site Induced fit (open) Enzyme ENZYME Allosteric site ACTIVE SITE Allosteric inhibitor (open) Enzyme ENZYME 8. Non competitive (reversible) allosteric inhibitors • Inhibitor binds reversibly to the allosteric site • Intermolecular bonds are formed • Induced fit alters the shape of the enzyme • Active site is distorted and is not recognised by the substrate • Increasing substrate concentration does not reverse inhibition • Inhibitor is not similar in structure to the substrate

31. Biosynthetic pathway S P P’’’ P’ P’’ Feedback control (open) Enzyme ENZYME 8. Non competitive (reversible) allosteric inhibitors Inhibition • Enzymes with allosteric sites often at start of biosynthetic pathways • Enzyme is controlled by the final product of the pathway • Final product binds to the allosteric site and switches off enzyme • Inhibitor may have a similar structure to the final product