The Hydratase Activity of Malonate Semialdehyde Decarboxylase: Mechanistic and Evolutionary Implications
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- 作者：Gerrit J. Poelarends ; Hector Serrano ; William H. Johnson ; Jr. ; David W. Hoffman ; and Christian P. Whitman
- 刊名：Journal of the American Chemical Society
- 出版年：2004
- 出版时间：December 8, 2004
- 年：2004
- 卷：126
- 期：48
- 页码：15658 - 15659
- 全文大小：43K
- 年卷期：v.126,no.48(December 8, 2004)
- ISSN：1520-5126
文摘
Malonate semialdehyde decarboxylase (MSAD) is a member of the tautomerase superfamily, a group of structurally homologous proteins that have a characteristic 
beta2.gif" BORDER=0 ALIGN="middle">-
-beta2.gif" BORDER=0 ALIGN="middle">-fold and a catalytic amino-terminal proline. In addition to its physiological decarboxylase activity, the conversion of malonate semialdehyde to acetaldehyde and carbon dioxide, the enzyme has now been found to display a promiscuous hydratase activity, converting 2-oxo-3-pentynoate to acetopyruvate, with a kcat/Km value of 6.0 × 102 M-1 s-1. Pro-1 and Arg-75 are critical for both activities, and the pKa of Pro-1 was determined to be ~9.2 by a direct 15N NMR titration. These observations implicate a decarboxylation mechanism in which Pro-1 polarizes the carbonyl oxygen of substrate by hydrogen bonding and/or an electrostatic interaction. Arg-75 may position the carboxylate group into a favorable orientation for decarboxylation. Both the hydratase activity and the pKa value of Pro-1 are shared with trans-3-chloroacrylic acid dehalogenase, another tautomerase superfamily member that precedes MSAD in a bacterial degradation pathway for trans-1,3-dichloropropene. Hence, MSAD and CaaD could have evolved by divergent evolution from a common ancestral protein, retaining the necessary catalytic components for the conjugate addition of water.
beta2.gif" BORDER=0 ALIGN="middle">--beta2.gif" BORDER=0 ALIGN="middle">-fold and a catalytic amino-terminal proline. In addition to its physiological decarboxylase activity, the conversion of malonate semialdehyde to acetaldehyde and carbon dioxide, the enzyme has now been found to display a promiscuous hydratase activity, converting 2-oxo-3-pentynoate to acetopyruvate, with a kcat/Km value of 6.0 × 102 M-1 s-1. Pro-1 and Arg-75 are critical for both activities, and the pKa of Pro-1 was determined to be ~9.2 by a direct 15N NMR titration. These observations implicate a decarboxylation mechanism in which Pro-1 polarizes the carbonyl oxygen of substrate by hydrogen bonding and/or an electrostatic interaction. Arg-75 may position the carboxylate group into a favorable orientation for decarboxylation. Both the hydratase activity and the pKa value of Pro-1 are shared with trans-3-chloroacrylic acid dehalogenase, another tautomerase superfamily member that precedes MSAD in a bacterial degradation pathway for trans-1,3-dichloropropene. Hence, MSAD and CaaD could have evolved by divergent evolution from a common ancestral protein, retaining the necessary catalytic components for the conjugate addition of water.