Format

Send to

Choose Destination
Nat Struct Biol. 1998 Sep;5(9):812-8.

Insights into transition state stabilization of the beta-1,4-glycosidase Cex by covalent intermediate accumulation in active site mutants.

Author information

1
Protein Engineering Network of Centres of Excellence, Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Canada.

Abstract

The catalytic mechanism of 'retaining' beta-glycosidases has been the subject of considerable interest and debate for many years. The visualization of a covalent glycosyl enzyme intermediate by X-ray crystallography was first accomplished with a saccharide substrate substituted with fluorine at its 2-position. The structure implicated major roles for residue His 205 and for the 2-hydroxyl position of the proximal saccharide in binding and catalysis. Here we have studied the kinetic behavior of various His 205 mutants. One of these mutants, a double mutant H205N/E127A, has been used to stabilize a covalent glycosyl-enzyme intermediate involving an unsubstituted sugar, permitting crystallographic analysis of the interactions between its 2-hydroxyl group and the enzyme.

PMID:
9731776
DOI:
10.1038/1852
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center