Isotope Effects on the Enzymatic and Nonenzymatic Reactions of Chorismate

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• 作者：S. Kirk Wright ; Michael S. DeClue ; Ajay Mandal ; Lac Lee ; Olaf Wiest ; W. Wallace Cleland ; and Donald Hilvert
• 刊名：Journal of the American Chemical Society
• 出版年：2005
• 出版时间：September 21, 2005
• 年：2005
• 卷：127
• 期：37
• 页码：12957 - 12964
• 全文大小：176K
• 年卷期：v.127,no.37(September 21, 2005)
• ISSN：1520-5126

文摘

The important biosynthetic intermediate chorismate reacts thermally by two competitive pathways,one leading to 4-hydroxybenzoate via elimination of the enolpyruvyl side chain, and the other to prephenateby a facile Claisen rearrangement. Measurements with isotopically labeled chorismate derivatives indicatethat both are concerted sigmatropic processes, controlled by the orientation of the enolpyruvyl group. Inthe elimination reaction of [4-²H]chorismate, roughly 60% of the label was found in pyruvate after 3 h at 60C. Moreover, a 1.846 ± 0.057 ²H isotope effect for the transferred hydrogen atom and a 1.0374 ± 0.0005¹⁸O isotope effect for the ether oxygen show that the transition state for this process is highly asymmetric,with hydrogen atom transfer from C4 to C9 significantly less advanced than C-O bond cleavage. In thecompeting Claisen rearrangement, a very large ¹⁸O isotope effect at the bond-breaking position (1.0482 ±0.0005) and a smaller ¹³C isotope effect at the bond-making position (1.0118 ± 0.0004) were determined.Isotope effects of similar magnitude characterized the transformations catalyzed by evolutionarily unrelatedchorismate mutases from Escherichia coli and Bacillus subtilis. The enzymatic reactions, like their solutioncounterpart, are thus concerted [3,3]-sigmatropic processes in which C-C bond formation lags behindC-O bond cleavage. However, as substantially larger ¹⁸O and smaller ¹³C isotope effects were observedfor a mutant enzyme in which chemistry is fully rate determining, the ionic active site may favor a somewhatmore polarized transition state than that seen in solution.