Enzyme catalysis: lessons from stereochemistry

Stereoselectively labeled samples of E—2an Z—[9—2H,3H]— chorismate have been synthesized from Fand z-[3H, Hlphosphoen6lpyruvate by the enzymes of the shikimate pathway. The position of the tritium label at the 9—position of chorismate has been determined, from which it is evident that in the reaction catalyzed by 3-phosphoshikimate 1—carboxyvinyltransferase, the addition step has the opposite stereo— chei9l course from the elimination step. These samples of [9— H, H]chorismate have been used to determine the stereochemical course of the reaction catalyzed by chorismate mutace. It is evident that the mutase proceeds via a transition state of chair—like geometry. The stereochemical course of enzyme—catalyzed reactions has been a fruitful source of information about the mechanisms of enzymic catalysis, and has both provided important generalizations and posed challeigig questions on the nature of these1gross. For example, the use of the chiral [ H, H, Himethyl group and the chiral [ 0, 0, 0]phospho group has demonstrated that whenever methyl or phospho transfers are mediated by enzymes, the reactions proceed via in—line associative paths that invert the configuration at carbon (or phosphorus) at each transfer. Such findings allow us more tightly to define the nature of the transition state for the catalyzed process, and usefully constrain our mechanistic postulates. In other areas, stereochemical investigations have generated interesting puzzles. For instance, why should enzymic aldol condensations seemingly always proceed with retention of configuration, yet the closely related Claisen condensations apparently always go with inversion? The reasons behind such evident stereochemical imperatives will surely, when we understand them, illuminate the larger question of why enzymes are such dramatically good catalysts. In this paper, we focus on one stereochemical problem, that relates to the mechanism of chorismate mutase. This enzyme catalyzes the reaction shown in Fig. 1, which is 4___ COO chorismate mutase — OH chorismafe prepheriate 1 2 Fig. 1. The 3,3-sigmatropic rearrangement of chorismate (1) to prephenate (2), catalyzed by the enzyme chorismate mutase. — formally a Claisen rearrangement. Indeed, the mutase is the only characterized example of an enzyme that catalyzes what is —— superficially at least -— a pericyclic reaction. While there are a number of transformations in alkaloid biosynthesis that involve the Claisen—like rearrangement of dimethylallyl ethers, the enzymes that mediate these reactions have never been isolated. The stereochemical problem posed by chorismate mutase concerns the geometry of the transition state, and there are two possible formulations: 'chair—like' and 'boat—like' (Fig. 2). While unbiased non—enzymic systems generally prefer a chair geometry, this preference is rather marginal in energetic terms, and the energy difference is certainly small (2—4 kcal/mol) when compared with the large 1005 00C