Strictosidine Synthase: Mechanism of a Pictet-Spengler Catalyzing Enzyme
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- 作者：Justin J. Maresh ; Lesley-Ann Giddings ; Anne Friedrich ; Elke A. Loris ; Santosh Panjikar ; Bernhardt L. Trout ; Joachim Stö ; ckigt ; Baron Peters ; Sarah E. O'Connor
- 刊名：Journal of the American Chemical Society
- 出版年：2008
- 出版时间：January 16, 2008
- 年：2008
- 卷：130
- 期：2
- 页码：710 - 723
- 全文大小：389K
- 年卷期：v.130,no.2(January 16, 2008)
- ISSN：1520-5126
文摘
The Pictet-Spengler reaction, which yields either a MG SRC="/images/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-carboline or a tetrahydroquinoline productfrom an aromatic amine and an aldehyde, is widely utilized in plant alkaloid biosynthesis. Here wedeconvolute the role that the biosynthetic enzyme strictosidine synthase plays in catalyzing the stereoselective synthesis of a 
mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-carboline product. Notably, the rate-controlling step of the enzyme mechanism, asidentified by the appearance of a primary kinetic isotope effect (KIE), is the rearomatization of a positivelycharged intermediate. The KIE of a nonenzymatic Pictet-Spengler reaction indicates that rearomatizationis also rate-controlling in solution, suggesting that the enzyme does not significantly change the mechanismof the reaction. Additionally, the pH dependence of the solution and enzymatic reactions provides evidencefor a sequence of acid-base catalysis steps that catalyze the Pictet-Spengler reaction. An additionalacid-catalyzed step, most likely protonation of a carbinolamine intermediate, is also significantly ratecontrolling. We propose that this step is efficiently catalyzed by the enzyme. Structural analysis of abisubstrate inhibitor bound to the enzyme suggests that the active site is exquisitely tuned to correctlyorient the iminium intermediate for productive cyclization to form the diastereoselective product. Furthermore,ab initio calculations suggest the structures of possible productive transition states involved in themechanism. Importantly, these calculations suggest that a spiroindolenine intermediate, often invoked inthe Pictet-Spengler mechanism, does not occur. A detailed mechanism for enzymatic catalysis of themages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-carboline product is proposed from these data.