Title | ENZYMATIC CLEAVAGE OF GLYCOSIDES - HOW DOES IT HAPPEN |
Publication Type | Journal Article |
Year of Publication | 1995 |
Authors | Withers, SG |
Journal | PURE AND APPLIED CHEMISTRY |
Volume | 67 |
Pagination | 1673-1682 |
Date Published | OCT |
ISSN | 0033-4545 |
Abstract | Many glycosidases operate through a double-displacement mechanism in which a glycosyl-enzyme intermediate is formed and hydrolysed via oxocarbenium ion-like transition states with acid/base catalysis. The two key active site residues involved in this mechanism, the active site nucleophile and the acid/base catalyst have been identified by novel means. The nucleophile is identified using a mechanism-based inactivator which functions by formation of a stabilised glycosyl-enzyme intermediate. Identification of the labelled peptide from proteolytic hydrolysates is achieved using a new tandem mass spectrometric method. The acid/base catalyst is identified by detailed kinetic analysis of candidate amino acids chosen on the basis of sequence similarities. |
DOI | 10.1351/pac199567101673 |
