Stereochemical course of hydrolysis and hydration reactions catalysed by cellobiohydrolases I and II from Trichoderma reesei.
Cellobiohydrolase I from Trichoderma reesei catalyzes the hydrolysis of methyl beta-D-cellotrioside (Km = 48 microM, kcat = 0.7 min-1) with release of the beta-cellobiose (retention of configuration). The same enzyme catalyzes the trans-hydration of cellobial (Km = 116 microM, kcat = 1.16 min-1) and lactal (Km = 135 microM, kcat = 1.35 min-1), presumably with glycosyl oxo-carbonium ion mediation. Protonation of the double bond is from the direction opposite that assumed for methyl beta-cellotrioside, but products formed from these prochiral substrates are again of beta configuration. Cellobiohydrolase II from the same microorganism hydrolyzes methyl beta-D-cellotetraoside (Km = 4 microM, kcat = 112 min-1) with inversion of configuration to produce alpha-cellobiose. The other reaction product, methyl beta-cellobioside, is in turn partly hydrolysed by cellobiohydrolase II to form methyl beta-D-glucoside and D-glucose, presumably the alpha-anomer. Reaction with cellobial is too slow to permit unequivocal determination of product configuration, but clear evidence is obtained that protonation occurs from the si-direction, again opposite that assumed for protonating glycosidic substrates. These results add substantially to the growing evidence that individual glycosidases create the anomeric configuration of their reaction products by means that are independent of substrate configuration.
Claeyssens
M
M
Laboratorium voor Biochemie, State University Ghent, Belgium.
Tomme
P
P
Brewer
C F
CF
Hehre
E J
EJ
D043366:Cellulose 1,4-beta-Cellobiosidase; D006026:Glycoside Hydrolases / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D003904:Mitosporic Fungi / Q000201:enzymology*; D013379:Substrate Specificity; D014242:Trichoderma / Q000201:enzymology*
D007527:Isoenzymes; D006026:Glycoside Hydrolases; D043366:Cellulose 1,4-beta-Cellobiosidase