Investigation of the Enzymatic and Nonenzymatic Cope Rearrangement of Carbaprephenate to Carbachorismate
2002-08-24T00:00:00Z (GMT) by
The dimethyl esters of carbaprephenate and 4-<i>epi</i>-carbaprephenate were prepared by modification of published procedures. In methanol these compounds are converted quantitatively to isomeric 6-hydroxytricyclo[126.96.36.199<sup>2,7</sup>]non-3-en-1,3-dimethyl esters via a two-step sequence involving an initial Cope rearrangement, followed by intramolecular Diels−Alder reaction of the dimethyl carbachorismate or 4-<i>epi</i>-carbachorismate intermediates. Carbaprephenate and its epimer were obtained by alkaline hydrolysis of the corresponding dimethyl esters. These compounds, in contrast to their ester precursors, undergo spontaneous acid-catalyzed decarboxylation in aqueous solution. Only at high pH does the Cope rearrangement compete with decarboxylation. At pH 12 and 90 °C, carbaprephenate slowly rearranges to carbachorismate, which rapidly loses water to give 3-(2-carboxyallyl)benzoic acid as the major product. A small amount of the intramolecular Diels−Alder adduct derived from carbachorismate is also observed by NMR as a minor product. Carbaprephenate is not a substrate for the enzyme chorismate mutase from <i>Bacillus subtilis</i> (BsCM), nor does carbaprephenate inhibit the normal chorismate mutase activity of this enzyme, even when present in 200-fold excess over chorismate. Its low affinity for the enzyme-active site is presumably a consequence of placing a methylene group rather than an oxygen atom proximal to the essential cationic residue Arg90. Nevertheless, BsCM variants that lack this cation (R90G and R90A) do not accelerate the Cope rearrangement of carbaprephenate either, and a catalytic antibody 1F7, which exhibits modest chorismate mutase activity, is similarly inactive. Poor substrate binding and the relatively high barrier for the Cope compared to the Claisen rearrangement presumably account for the lack of detectable catalysis. Acceleration of this sigmatropic rearrangement apparently requires more than an active site that is complementary in shape to the reactive substrate conformer.