WEEK 10: DRUG TARGETS, ENZYMES AND RECEPTORS

Enzymes

Structure and function of enzymes

• Globular proteins (3⁰ and 4⁰ structures)

• Catalysts (speed up reactions)

• Lower activation energy

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)

• participate in the enzyme-catalysed reaction

Enzymes

• Are specific for what they will catalyze

• Are Reusable

• End in

–ase

-Sucrase

-Lactase

-Maltase

Methods of enzyme catalysis

• Provides a reaction surface (the active site)

• Provides a suitable environment (hydrophobic)

• Brings reactants together

• Positions reactants correctly for reaction

• Weakens bonds in the reactants

• Provides acid / base catalysis

• Provides nucleophilic groups

• Stabilises the transition state with intermolecular bonds

• Lowering the activation energy of reaction

How do enzymes reduce activation energy?

• Enzymes + substrates form enzyme-substrate complex

• Binding interactions with ‘aa’ residues hold the substrate in place

How do enzymes reduce activation energy?

• Catalytic ‘aa’ residues (R groups) facilitate the reaction

– making or breaking bonds in the substrate

• Product is formed and released

Substrate Binding: Bonding forces

• Ionic

• H-bonding

• van der Waals

Substrate binding: bonding forces

Substrate Binding: Induced fit

• Active site is flexible (not precisely complementary to substrate)

• Substrate enters active site, active site conformation changes

• Amino acids in active site move closer to form chemical bonds with substrate

• Strains bonds in substrate

• Lowers activation energy of reaction

Catalysis mechanisms: acid/base catalysis

Catalysis mechanisms: nucleophilic residues

Enzyme inhibition

Overall process of enzyme catalysis

• Binding interactions must be strong enough to hold the substrate sufficiently long for the reaction to occur

• Interactions must be weak enough to allow the product to depart

• Interactions stabilise the transition state

• Designing molecules with stronger binding interactions results in enzyme inhibitors which block the active site

Reverse inhibitors

• Inhibitor binds reversibly to the active site

• Intermolecular bonds are involved in binding

• The inhibitor undergoes no reaction

• 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 substrate, product or cofactor

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

Reversible and irreversible inhibitors

Irreversible Inhibitors - orlistat

• Orlistat is an anti-obesity drug that inhibits pancreatic lipase

• The enzyme is blocked from digesting fats in the intestine

• Fatty acids and glycerol are less absorbed as a result

• Leads to reduced biosynthesis of fat in the body

Allosteric inhibitors

• Inhibitor binds reversibly to the allosteric site

• Intermolecular bonds are formed

• Induced fit alters the shape of the enzyme

• Since active site is distorted it is not recognised by the substrate

• Increasing substrate concentration does not reverse inhibition

• Allosteric inhibitors do not have to be similar in structure to the substrate

• Inhibits the first enzyme in the biosynthesis of purines

• Blocks the biosynthesis of purines and DNA

• Used in the treatment of leukaemia

Transition-state Inhibitors

• Drugs designed to mimic the transition state of an enzyme-catalysed reaction

• Transition-state inhibitors are likely to bind more strongly than drugs mimicking the substrate or product

• Transition states are high energy, transient species and cannot be isolated or synthesised

• Design based on reaction intermediates so are closer in character to transition states than substrates or products

• These drugs mimic the stereochemistry and binding properties of the reaction intermediate, but is stable.

Suicide Substrates

• Agents which are converted to irreversible inhibitors by the enzyme-catalysed reaction

• React with the target enzyme once formed

Aspirin – suicide substrate of Cyclooxygenase 1 (Cox1) enzyme

Enzyme targets for useful medications

1. Antibacterial agents

   • Dihydropteroate synthetase, transpeptidase

2. Antiviral agents

   • HIV reverse transcriptase, HIV protease, viral DNA polymerase

3. Anti-inflammatory agents

   • Cyclooxygenase

4.  Cholesterol lowering agents

   • HMG-CoA reductase

5.  Antidepressants

   • Monoamine oxidase

6.  Anticancer agents

   • Tyrosine kinase, dihydrofolate reductase, thymidylate synthase, aromatase etc

7. Antihypertensive agents

   • Renin, angiotensin converting enzyme

8.  Treatment of male erectile dysfunction

   • Phosphodiesterase

9. Anti-gout agents

   • Xanthine oxidase

10. Anti-ulcer agents

    • Proton pump

11. Alzheimers disease

    •  Cholinesterases

12. Diuretics

    • Carbonic anhydrase

Agonists vs Antagonists

• Agonists are drugs designed to mimic the natural messenger

 • Antagonists are drugs designed to block the natural messenger

Receptors: specific & membrane bound

Natural messenger binding

• Receptors contain a binding site, which is nearly the correct shape for the messenger

• Binding alters the shape of the receptor (induced fit)

• Altered receptor shape leads to further effects - signal transduction

• Chemical messenger does not enter the cell, is not permanently bound and departs unchanged.

Design of Agonists

• Agonists bind reversibly to the binding site and produce the same induced fit as the natural messenger - receptor is activated

• Similar intermolecular bonds formed as with natural messenger

• Agonists are often similar in structure to the natural messenger

Requirements:

• The agonist must have the correct binding groups

• The binding groups must be correctly positioned to interact with complementary binding regions (regions in binding site)

• The drug must have the correct shape to fit the binding site

• Identify important binding interactions in natural messenger

• Agonists are designed to have functional groups capable of same interactions

• Usually require same number of interactions

Correct position of binding groups

• Binding groups must be positioned to enable interaction of complementary binding regions at the same time

• Example has three binding groups, but only two can bind simultaneously → Example will have poor activity

• One enantiomer of a chiral drug normally binds more effectively than the other

• Different enantiomers likely to have different biological properties

• Agonist must have correct size and shape to fit binding site • Groups preventing access are called steric shields or steric blocks

Antagonists

• Antagonists are drugs designed to block the natural messenger

• Antagonists tend to have stronger and/or more binding interactions, resulting in a different induced fit such that the receptor is not activated.

Reversible antagonists

• Antagonist binds reversibly to the binding site

• Intermolecular bonds involved in binding

• Different induced fit means receptor is not activated

• The antagonist does not undergo any reaction

• Level of antagonism depends on strength of antagonist binding and concentration

• Messenger is blocked from the binding site

• Increasing the messenger concentration reverses antagonism

• Antagonists bind to the binding site but fail to produce the correct induced fit - receptor is not activated • Normal messenger is blocked from binding

Antagonists can form binding interactions with binding regions in the binding site not used by the natural messenger

Antagonists can form binding interactions with binding regions in the binding site not used by the natural messenger

Induced fit resulting from binding of the normal messenger

Different induced fit resulting from extra binding interaction

• Antagonist binds irreversibly to the binding site

• Different induced fit means that the receptor is not activated

• Covalent bond is formed between the drug and the receptor

• Messenger is blocked from the binding site

• Increasing messenger concentration does not reverse antagonism

• Often used to label receptors

Allosteric antagonists

• Antagonist binds reversibly to an allosteric binding site

• Intermolecular bonds formed between antagonist and binding site

• Induced fit alters the shape of the receptor

• Binding site is distorted and is not recognised by the messenger

• Increasing messenger concentration does not reverse antagonism

Antagonists by the Umbrella Effect

• Antagonist binds reversibly to a neighbouring binding site

• Intermolecular bonds formed between antagonist and binding site

• Antagonist overlaps the messenger binding site

• Messenger is blocked from the binding site

• Agent binds but does not produce the ideal induced fit for maximum effect

• Agent binds to binding site in two different modes, one where the agent acts as an agonist and one where it acts as an antagonist

• Agent binds as an agonist to one receptor subtype, but as an antagonist to another receptor subtype

Inverse Agonists

Properties shared with antagonists

• Bind to receptor binding sites with a different induced fit from the normal messenger

• Receptor is not activated

• Normal messenger is blocked from binding to the binding site

 Properties not shared with antagonists

• Block any inherent activity related to the receptor (e.g. GABA receptor)

• Inherent activity = level of activity present in the absence of a chemical messenger

• Receptors are in an equilibrium between constitutionally active and inactive forms

How drugs affect receptor equilibria