An assay for selective determination of exo-1,4,-beta-glucanases in a mixture of cellulolytic enzymes.
A selective procedure using synthetic substrates for determination of exo-1,4,-beta-glucanases in a mixture of exoglucanases , endoglucanases , and beta-glucosidases is formulated. The heterobiosides , p- nithrophenyl -beta-D- cellobioside ( pNPC ) or p-nitrophenyl-beta-D-lactoside ( pNPL ), were used as selective substrates for the measurement of exoglucanase activity. The exoglucanases (especially cellobiohydrolases , which split off cellobiose units from the nonreducing end of the cellulose chain) specifically act on the agluconic bond (between p-nitrophenyl and the disaccharide moiety) and not on the holosidic bond (between the two glucose units of cellobiose). The interfering effect of beta-glucosidase, which acts on both agluconic and holosidic bonds, is overcome by the addition of D-glucono-1,5-delta-lactone, a specific inhibitor of beta-glucosidases. The interference of endoglucanases , which also act on both agluconic and holosidic bonds, can be compensated for by prior standardization of the assay procedure with a purified endoglucanase from the studied mixture of cellulases.
D002475:Cellobiose / Q000378:metabolism; D002480:Cellulase / Q000032:analysis*; D002482:Cellulose / Q000378:metabolism; D043366:Cellulose 1,4-beta-Cellobiosidase; D005959:Glucosidases / Q000032:analysis*; D005960:Glucosides / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000032:analysis*; D006027:Glycosides / Q000378:metabolism; D007700:Kinetics; D008722:Methods; D001617:beta-Glucosidase / Q000032:analysis* / Q000037:antagonists & inhibitors
D005960:Glucosides; D006027:Glycosides; D002475:Cellobiose; C029976:4-aminophenyl beta-lactoside; C034170:4-nitrophenyl beta-cellobioside; D002482:Cellulose; D005959:Glucosidases; D006026:Glycoside Hydrolases; D001617:beta-Glucosidase; D002480:Cellulase; D043366:Cellulose 1,4-beta-Cellobiosidase