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Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate

Vocadlo, D J, Davies, G J, Laine, R and Withers, S G (2001) Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate. Nature. pp. 835-838. ISSN 0028-0836

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Abstract

Hen egg-white lysozyme (HEWL) was the first enzyme to have its three-dimensional structure determined by X-ray diffraction techniques(1). A catalytic mechanism, featuring a long-lived oxo-carbenium-ion intermediate, was proposed on the basis of model-building studies(2). The `Phillips' mechanism is widely held as the paradigm for the catalytic mechanism of beta -glycosidases that cleave glycosidic linkages with net retention of configuration of the anomeric centre. Studies with other retaining beta -glycosidases, however, provide strong evidence pointing to a common mechanism for these enzymes that involves a covalent glycosyl-enzyme intermediate, as previously postulated(3). Here we show, in three different cases using electrospray ionization mass spectrometry, a catalytically competent covalent glycosyl-enzyme intermediate during the catalytic cycle of HEWL. We also show the three-dimensional structure of this intermediate as determined by Xray diffraction. We formulate a general catalytic mechanism for all retaining beta -glycosidases that includes substrate distortion, formation of a covalent intermediate, and the electrophilic migration of C1 along the reaction coordinate.

Item Type: Article
Copyright, Publisher and Additional Information: © 2001 Nature Publishing Group
Keywords: ACTIVE-SITE, BETA-GLUCOSIDASE, LEAVING GROUP, HYDROLYSIS, DISTORTION, MECHANISM, INSIGHTS, MUTANTS, ACID
Academic Units: The University of York > Chemistry (York)
Depositing User: Repository Officer
Date Deposited: 13 Oct 2004
Last Modified: 17 Oct 2013 14:34
Published Version: http://dx.doi.org/10.1038/35090602
Status: Published
Refereed: Yes
Related URLs:
URI: http://eprints.whiterose.ac.uk/id/eprint/131

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