Skip Navigation


PEDS Advance Access originally published online on March 14, 2005
Protein Engineering Design and Selection 2005 18(1):33-40; doi:10.1093/protein/gzi003
This Article
Right arrow Full Text Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
18/1/33    most recent
gzi003v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (16)
Right arrowRequest Permissions
Google Scholar
Right arrow Articles by Müllegger, J.
Right arrow Articles by Withers, S. G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Müllegger, J.
Right arrow Articles by Withers, S. G.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2005. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions{at}oupjournals.org

Engineering of a thioglycoligase: randomized mutagenesis of the acid–base residue leads to the identification of improved catalysts

Johannes Müllegger1, Michael Jahn1, Hong-Ming Chen1, R.Antony J. Warren2 and Stephen G. Withers1,3

Protein Engineering Network of Centres of Excellence, 1Department of Chemistry and 2Department of Microbiology, University of British Columbia, Vancouver, BC V6T 1Z1, Canada

3 To whom correspondence should be addressed. E-mail: withers{at}chem.ubc.ca

Thioglycoligases are recently introduced variants of retaining glycosidases in which the acid–base catalyst has been mutated, rendering them capable of thioglycoside synthesis. The original acid–base mutant of Agrobacterium sp. ß-glucosidase (E170A) was previously shown to be an effective thioglycoligase carrying out glycosyltransfer from 2,4-dinitrophenyl glycosides to several different thio sugar acceptors. Here we report the generation of a screen for improved thioglycoligases, randomized mutagenesis of the acid–base catalyst E170 and identification of variants superior to E170A. Furthermore we have established a coupled assay allowing kinetic analysis of isolated variants and found that Abg E170Q is 5-fold faster than Abg E170A when 2,4-dinitrophenyl glucoside is used as donor and 100-fold faster when glucosyl azide is used. To demonstrate its utility, different acceptor and donor sugar combinations were employed to produce thio-linked di- or trisaccharides in high yields, showing the considerable versatility of the system for the synthesis of carbohydrate mimetics.

Received November 18, 2004; revised January 19, 2005; accepted January 20, 2005.

Edited by Dan Tawfik


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Protein Eng Des SelHome page
J. Paramesvaran, E. G. Hibbert, A. J. Russell, and P. A. Dalby
Distributions of enzyme residues yielding mutants with improved substrate specificities from two different directed evolution strategies
Protein Eng. Des. Sel., July 1, 2009; 22(7): 401 - 411.
[Abstract] [Full Text] [PDF]


Disclaimer: Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.