Carboxymethylcellulase and Avicelase activities from a cellulolytic Clostridium strain A11.
The extracellular cellulase enzyme system of Clostridium A11 was fractionated by affinity chromatography on Avicel: 80% of the initial carboxymethylcellulase (CMCase) activity was adhered. This cellulase system was a multicomponent aggregate. Several CMCase activities were detected, but the major protein P1 had no detectable activity. Adhered and unadhered cellulases showed CMCase activity with the highest specific activity in Avicel-adhered fraction. However, only adhered fractions could degrade Avicel. Thus, efficiency of the enzymatic hydrolysis of Avicel was related to the cellulase-adhesion capacity. Carboxymethylcellulase and Avicelase activities were studied with the extracellular enzyme system and cloned cellulases. Genomic libraries from Clostridium A11 were constructed with DNA from this Clostridium, and a new gene cel1 was isolated. The gene(s) product(s) from cel1 exhibited CMCase and p-nitrophenylcellobiosidase (pNPCbase) activities. This cloned cellulase adhered to cellulose. Synergism between "adhered enzyme system" and cloned endoglucanases was observed on Avicel degradation. Conversely, no synergism was observed on CMC hydrolysis. Addition of cloned endoglucanase to cellulase complex led to increase of the Vmax without significant Km variation. Cloned endoglucanases can be added to cellulase complexes to efficiently hydrolyze cellulose.
Benoit
L
L
Laboratoire de Chimie Biologique I, Université de Nancy I, Vandoeuvre-lès-Nancy, France.
Cailliez
C
C
Gehin
A
A
Thirion
J
J
Raval
G
G
Petitdemange
H
H
D002480:Cellulase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002482:Cellulose; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D006026:Glycoside Hydrolases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D015183:Restriction Mapping; D013379:Substrate Specificity
D002482:Cellulose; D006026:Glycoside Hydrolases; D002480:Cellulase; C019826:carboxymethylcellulase