An enzyme E catalyzes the bimolecular reaction A + B → P in aqueous solution at 298 K. Kinetic studies show that the uncatalyzed reaction is extremely slow, but when E is present the rate constant increases by more than 10^6. Spectroscopic data indicate the following:
• A and B bind simultaneously to a cleft on the surface of E, forming the ternary complex E·A·B.
• A carboxylate side chain of the enzyme donates a proton to B in the rate-determining step and is reprotonated before product release.
Answer the following questions about the catalytic strategy used by E.
(a) Identify each type of catalysis (binding/orientation and/or covalent) that is operative in the mechanism of E. Justify your classification using the information provided.
(b) Explain, at the molecular level, how simultaneous binding of A and B to the enzyme cleft lowers the activation energy relative to the uncatalyzed reaction.
(c) The proton transfer involving the carboxylate side chain generates a new, short-lived intermediate. Explain why this step is classified as covalent (acid–base) catalysis and describe how the catalyst is regenerated by the end of the cycle.