Enzymes and Chymotrypsin Mechanism

Enzyme Structure

Large structure with a small active site; positions groups in the active site, controls activity through inhibitor binding, and facilitates subunit interactions.

Active Site Participants

Amino acid residues, cofactors (metal ions), and coenzymes (small organic molecules).

Cofactors

Usually metal ions that cannot be substituted by other ions with the same charge due to differences in their orbitals.

Coenzymes

Small organic molecules, many of which are vitamins (e.g., NAD, FMN, Lipoic acid, biotin).

Nucleophiles

Atoms with electron density that attack partial positive charges; examples include Cysteine (C), Serine (S), Lysine (K), Histidine (H), and Water.

General Acid Catalysts

R groups that donate a proton to speed up a reaction; examples include Aspartic acid and Cysteine.

General Base Catalysts

R groups that grab a proton to speed up a reaction; same as general acid catalysts, minus Arginine.

pH Importance

Cells regulate pH using buffers; pH affects ionic and hydrogen bonds, influencing protein structure.

Chymotrypsin

Enzyme that cuts on the carboxyl side of large aromatic groups (W, Y, F).

Chymotrypsin Overall Reaction

Peptide + H2O --> Two smaller peptides (Hydrolysis)

Substrate Binding (Chymotrypsin)

Peptide (S1) enters the active site, encountering a hydrophobic pocket formed by nonpolar residues like valine, leucine, and isoleucine.

Catalytic Triad

Ser195, His57, Asp102; Serine acts as a nucleophile, Histidine acts as a general base, and Aspartate raises the pKa of histidine.

Deacylation (Chymotrypsin)

Water molecule (S2) enters the active site and attacks the carbonyl carbon of the acyl-enzyme intermediate.

Spectrophotometry Techniques

Mix in some organic solvent like ethanol, propanol, something that is miscible with water to lower the freezing point.

Enzyme Catalysis - Activation Energy

Enzymes lower the activation energy by ordering the substrate in the active site and providing R groups, coenzymes, and cofactors to facilitate the reaction.

Reaction coordinate

Progress of the reaction