Unexpected regioselectivity of Humicola insolens Cel7B glycosynthase mutants.
Four Humicola insolens Cel7B glycoside hydrolase mutants have been evaluated for the coupling of lactosyl fluoride on O-allyl N(I)-acetyl-2(II)-azido-beta-chitobioside. Double mutants Cel7B E197A H209A and Cel7B E197A H209G preferentially catalyze the formation of a beta-(1-->4) linkage between the two disaccharides, while single mutant Cel7B E197A and triple mutant Cel7B E197A H209A A211T produce predominantly the beta-(1-->3)-linked tetrasaccharide. This result constitutes the first report of the modulation of the regioselectivity through site-directed mutagenesis for an endoglycosynthase.
Blanchard
Sophie
S
CERMAV-FR-CNRS 2607, BP 53, F-38041 Grenoble, France.
Armand
Sylvie
S
Couthino
Pedro
P
Patkar
Shamkant
S
Vind
Jesper
J
Samain
Eric
E
Driguez
Hugues
H
Cottaz
Sylvain
S
D001203:Ascomycota / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D001386:Azides / Q000138:chemical synthesis / Q000737:chemistry; D002240:Carbohydrate Sequence; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D004187:Disaccharides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006027:Glycosides / Q000096:biosynthesis / Q000737:chemistry; D007785:Lactose / Q000031:analogs & derivatives / Q000737:chemistry; D016297:Mutagenesis, Site-Directed; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D013237:Stereoisomerism
D001386:Azides; D004187:Disaccharides; D006027:Glycosides; C519395:O-allyl N(1)-acetyl-2(II)-azidochitobioside; D009844:Oligosaccharides; C084793:lactosyl fluoride; D006026:Glycoside Hydrolases; D007785:Lactose