Correlative Variations in Enzyme-Derived and Substrate-Derived Structures of Catalytic Transition States. Implications for the Catalytic Strategy of Acyl-Transfer Enzymes

Article abstract of DOI:10.1021/ja00346a056

Acetylchymotrypsin, acetyl elastase and (carbobenzyloxy)glycyl elastase all undergo hydrolysis with the same overall solvent isotope effect, which arises from a single protonic site n/k₁ = 2.45(1-n + n/2.45)>. <(Carbobenzyloxy)glycyl>chymotrypsin, however, shows a larger effect arising from at least two sites n/k₁ = 3.34(1-n + n/1.85)²>.Formylchymotrypsin and acetylchymotrypsin undergo deacylation with α-deuterium and β-deuteium secondary isotope effects, respectively, that suggest fractional tetrahedral character at the transition state of about 0.44 (vs. 0.58 - 0.66 for similar nonenzymic reactions) when compared to equilibrium isotope effects for complete addition.The effect for acetyl elastase suggests much less tetrahedral character (0.27).Addition of an N-acyl function leads to a more inverse isotope effect, per deuterium, and thus to an apparent increase in tetrahedral character: to 0.84 for <(carbobenzyloxy)glycyl>chymotrypsin; to 0.43 for (carbobenzyloxy)glycyl elastase.It is concluded that enzyme-substrate interactions at the transition state can alter both enzyme structure, as shown by the solvent isotope effects, and substrate structure as shown by the substrate isotope effects.Such alterations, in the combination of enzyme with natural substrate, probably adjust both structures for optimal catalytic interaction.