White Rose University Consortium logo
University of Leeds logo University of Sheffield logo York University logo

Modifying the stereochemistry of an enzyme-catalyzed reaction by directed evolution

Williams, G.J, Domann, S., Nelson, A. and Berry, A. (2003) Modifying the stereochemistry of an enzyme-catalyzed reaction by directed evolution. Proceedings of the National Academy of Sciences, 100 (6). pp. 3143-3148. ISSN 0027-8424

Full text available as:
[img]
Preview
Text
berrya1.pdf
Available under License : See the attached licence file.

Download (410Kb)

Abstract

Aldolases have potential as tools for the synthesis of stereochemically complex carbohydrates. Here, we show that directed evolution can be used to alter the stereochemical course of the reaction catalyzed by tagatose-1,6-bisphosphate aldolase. After three rounds of DNA shuffling and screening, the evolved aldolase showed an 80-fold improvement in k-cat/K-m toward the non-natural substrate fructose 1,6-bisphosphate, resulting in a 100-fold change in stereospecificity. (31)P NMR spectroscopy was used to show that, in the synthetic direction, the evolved aldolase catalyzes the formation of carbon—carbon bonds with unnatural diastereoselectivity, where the >99:<1 preference for the formation of tagatose 1,6-bisphosphate was switched to a 4:1 preference for the diastereoisomer, fructose 1,6-bisphosphate. This demonstration is of considerable significance to synthetic chemists requiring efficient syntheses of complex stereoisomeric products, such as carbohydrate mimetics.

Item Type: Article
Copyright, Publisher and Additional Information: © Copyright 2003 National Academy of Sciences, U.S.A.
Academic Units: The University of Leeds > Faculty of Biological Sciences (Leeds) > Institute of Molecular and Cellular Biology (Leeds)
Depositing User: Sherpa Assistant
Date Deposited: 13 Mar 2006
Last Modified: 08 Feb 2013 17:01
Published Version: http://www.pnas.org/cgi/content/full/100/6/3143
Status: Published
Refereed: Yes
Identification Number: 10.1073/pnas.0635924100
URI: http://eprints.whiterose.ac.uk/id/eprint/557

Actions (login required)

View Item View Item