Strictosidine Synthase: Mechanism of a Pictet-Spengler Catalyzing Enzyme

Authors

Justin J. Maresh, Massachusetts Institute of Technology
Lesley Ann Giddings, Massachusetts Institute of TechnologyFollow
Anne Friedrich, Massachusetts Institute of Technology
Elke A. Loris, Zhejiang University
Santosh Panjikar, European Molecular Biology Laboratory Hamburg
Bernhardt L. Trout, Massachusetts Institute of Technology
Joachim Stöckigt, Zhejiang University
Baron Peters, University of California, Santa Barbara
Sarah E. O'Connor, Massachusetts Institute of Technology

Document Type

Article

Publication Date

1-16-2008

Publication Title

Journal of the American Chemical Society

Abstract

The Pictet-Spengler reaction, which yields either a β-carboline or a tetrahydroquinoline product from an aromatic amine and an aldehyde, is widely utilized in plant alkaloid biosynthesis. Here we deconvolute the role that the biosynthetic enzyme strictosidine synthase plays in catalyzing the stereoselective synthesis of a β-carboline product. Notably, the rate-controlling step of the enzyme mechanism, as identified by the appearance of a primary kinetic isotope effect (KIE), is the rearomatization of a positively charged intermediate. The KIE of a nonenzymatic Pictet-Spengler reaction indicates that rearomatization is also rate-controlling in solution, suggesting that the enzyme does not significantly change the mechanism of the reaction. Additionally, the pH dependence of the solution and enzymatic reactions provides evidence for a sequence of acid-base catalysis steps that catalyze the Pictet-Spengler reaction. An additional acid-catalyzed step, most likely protonation of a carbinolamine intermediate, is also significantly rate controlling. We propose that this step is efficiently catalyzed by the enzyme. Structural analysis of a bisubstrate inhibitor bound to the enzyme suggests that the active site is exquisitely tuned to correctly orient 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 the mechanism. Importantly, these calculations suggest that a spiroindolenine intermediate, often invoked in the Pictet-Spengler mechanism, does not occur. A detailed mechanism for enzymatic catalysis of the β-carboline product is proposed from these data.

Volume

130

Issue

2

First Page

710

Last Page

723

DOI

10.1021/ja077190z

ISSN

00027863

Comments

Peer reviewed accepted manuscript.

Recommended Citation

Maresh, Justin J.; Giddings, Lesley Ann; Friedrich, Anne; Loris, Elke A.; Panjikar, Santosh; Trout, Bernhardt L.; Stöckigt, Joachim; Peters, Baron; and O'Connor, Sarah E., "Strictosidine Synthase: Mechanism of a Pictet-Spengler Catalyzing Enzyme" (2008). Chemistry: Faculty Publications, Smith College, Northampton, MA.
https://scholarworks.smith.edu/chm_facpubs/59