The adsorption of a bacterial cellulase and its two isolated domains to crystalline cellulose.
CenA is a bacterial cellulase (beta-1,4-glucanase) comprised of a globular catalytic domain joined to an extended cellulose-binding domain (CBD) by a short linker peptide. The adsorption of CenA and its two isolated domains to crystalline cellulose was analyzed. CenA and CBD.PTCenA' (the CBD plus linker) adsorbed rapidly to cellulose at 30 degrees C, and no net desorption of protein was observed during the following 16.7 h. There was no detectable adsorption of the catalytic domain. Scatchard plots of adsorption data for CenA and for CBD.PTCenA were nonlinear (concave upward). The adsorption of CenA and CBD.PTCenA exceeded 7 and 8 mumol/g cellulose, respectively, but saturation was not attained at the highest total protein concentrations employed. A new model for adsorption was developed to describe the interaction of a large ligand (protein) with a lattice of overlapping potential binding sites (cellobiose residues). A relative equilibrium association constant (Kr) of 40.5 and 45.3 liter.g cellulose-1 was estimated for CenA and CBD.PTCenA, respectively, according to this model. A similar Kr value (33.3 liter.g-1) was also obtained for Cex, a Cellulomonas fimi enzyme which contains a related CBD but which hydrolyzes both beta 1,4-xylosidic and beta-1,4-glucosidic bonds. It was estimated that the CBD occupies approximately 39 cellobiose residues on the cellulose surface.
Gilkes
N R
NR
Department of Microbiology, University of British Columbia, Vancouver, Canada.
Jervis
E
E
Henrissat
B
B
Tekant
B
B
Miller
R C
RC
Warren
R A
RA
Kilburn
D G
DG
D000327:Adsorption; D001483:Base Sequence; D001665:Binding Sites; D002480:Cellulase / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D002852:Chromatography, Ion Exchange; D003460:Crystallization; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D005822:Genetic Vectors; D008969:Molecular Sequence Data
D002482:Cellulose; D002480:Cellulase