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REGION-DIRECTED MUTAGENESIS OF RESIDUES SURROUNDING THE ACTIVE-SITE NUCLEOPHILE IN BETA-GLUCOSIDASE FROM AGROBACTERIUM-FAECALIS

TitleREGION-DIRECTED MUTAGENESIS OF RESIDUES SURROUNDING THE ACTIVE-SITE NUCLEOPHILE IN BETA-GLUCOSIDASE FROM AGROBACTERIUM-FAECALIS
Publication TypeJournal Article
Year of Publication1992
AuthorsTRIMBUR, DE, WARREN, RAJ, Withers, SG
JournalJOURNAL OF BIOLOGICAL CHEMISTRY
Volume267
Pagination10248-10251
Date PublishedMAY 25
ISSN0021-9258
Abstract

The active site nucleophile of the beta-glucosidase of Agrobacterium faecalis has recently been identified by the use of inhibitors. A combination of site-directed and in vitro enzymatic mutagenesis was carried out on the beta-glucosidase to probe the structure of the active site region. Forty-three point mutations were generated at 22 different residues in the region surrounding the active site nucleophile, Glu358. Only five positions were identified which affected enzyme activity indicating that only a few key residues are important to enzyme activity, thus the enzyme can tolerate a number of single residue changes and still function. The importance of Glu358 to enzymatic function has been confirmed and other residues important to enzyme structure or function have been identified.