Heavy Atom Isotope Effects Reveal a Highly Polarized Transition State for Chorismate Mutase

The conversion of chorismate (1) to prephenate (2) is catalyzed by the enzyme chorismate mutase. The enzymatic reaction is formally a Claisen rearrangement, proceeding through a chairlike transition state, but more detailed information on the structure of this high-energy species has remained elusive. Kinetic isotope effects (KIEs) are powerful tools in the determination of transition-state geometries. Secondary tritium isotope effects, for example, have shown that the uncatalyzed rearrangement of chorismate occurs via an asymmetric transition state in which the C(5)-O(5) bond is approximately 40% broken and the C(1)-C(9) bond is not detectably formed. These results are in accord with calculations based on the RHF/6-31* transition structure for this reaction and are typical for aliphatic Claisen rearrangements. Unfortunately, no isotope effects were observed for the reaction catalyzed by the bifunctional chorismate mutase-prephenate dehydrogenase from Escherichia coli. Suppression of the intrinsic isotope effect suggests a kinetically significant transition state prior to the rearrangement itself, presumably involving substrate complexation or a protein conformational change, which nevertheless precludes direct study of the chemical reaction at the enzyme active site.