Mechanism of Action of Enzymes
Enzymes accelerate reactions by providing an alternative reaction pathway with a lower activation energy (Ea). The enzyme is not consumed — it is regenerated at the end of each catalytic cycle.
Active Site
A specific 3D cleft or pocket on the enzyme surface where substrate binds. Comprises only a small portion of total amino acid residues. Two regions: Binding site (recognizes substrate) and Catalytic site (performs chemistry).
Models of Enzyme-Substrate Interaction
- Lock and Key Model (Emil Fischer, 1894): Active site has a rigid, pre-formed complementary shape to the substrate. Explains high specificity but not all enzyme behavior.
- Induced Fit Model (Koshland, 1958): Active site is flexible; undergoes conformational change upon substrate binding to achieve proper alignment of catalytic groups. Better explains enzyme behavior (e.g., Hexokinase closes around glucose, excluding water).
Chemical Mechanisms of Catalysis
- Acid-Base Catalysis: Active site amino acids (His, Asp, Glu) donate or accept protons to facilitate reaction (most common mechanism).
- Covalent Catalysis: Transient covalent bond forms between enzyme and substrate. Example: Serine proteases (trypsin, chymotrypsin) form acyl-enzyme intermediate. Nucleophilic serine at active site.
- Metal Ion Catalysis: Metal ions stabilize negative charges on transition state, mediate redox reactions, or orient substrate. Examples: Zn²⁺ in Carbonic anhydrase, Mg²⁺ in kinases.
- Proximity and Orientation Effects: Enzyme brings reactants into close proximity and correct orientation — major contributor to rate enhancement.
- Transition State Stabilization: Enzyme binds the transition state more tightly than substrate or product → lowers Ea.
Serine Proteases — Classic Example
Active site has a catalytic triad: Ser, His, Asp. Mechanism: Ser-OH acts as nucleophile → forms acyl-enzyme intermediate → water hydrolyzes the bond → enzyme regenerated. Examples: Trypsin (cleaves after Arg/Lys), Chymotrypsin (after Phe/Trp/Tyr), Elastase (after small residues).
Enzyme Action — Energetics
- Enzymes do NOT change ΔG of reaction (equilibrium position)
- They only speed up the rate to reach equilibrium
- They lower Ea for both forward AND reverse reactions
- Transition state analogs are powerful enzyme inhibitors (used as drugs)
Regulatory Sites
Allosteric enzymes have a regulatory site distinct from the active site. Binding of allosteric effectors causes conformational change that alters catalytic activity. Examples: Phosphofructokinase-1 (key regulator of glycolysis), ATCase (pyrimidine synthesis).