From beta-glucanase to beta-glucansynthase: glycosyl transfer to alpha-glycosyl fluorides catalyzed by a mutant endoglucanase lacking its catalytic nucleophile

Removal of the catalytic nucleophile Glu134 of the retaining 1,3-1,4-beta-g lucanase from Bacillus licheniformis by mutation to alanine yields an enzym e with no glycosidase activity, The mutant is able to catalyze the regio- a nd stereospecific glycosylation of alpha-laminaribiosyl fluoride with diffe rent glucoside accepters through a single-step inverting mechanism. The mai n advantage of the mutant as glycosylation catalyst with respect to the kin etically controlled transglycosylation using the wild-type enzyme is that t he reaction products cannot be hydrolyzed by the mutant enzyme, and glycosy lation yields rise to 90%. (C) 1998 Federation of European Biochemical Soci eties.