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Lookup NU author(s): Dr David Bolam, Emeritus Professor Harry Gilbert
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To investigate the mode of action of cellulose-binding domains (CBDs), the Type II CBD from Pseudomonas fluorescens subsp. cellulosa xylanase A (XYLA(CBD)) and cellulase E (CELE(CBD)) were expressed as individual entities or fused to the catalytic domain of a Clostridium thermocellum endoglucanase (EGE). The two CBDs exhibited similar K(a) values for bacterial microcrystalline cellulose (CELE(CBD), 1.62 x 106 M-1; XYLA(CBD), 1.83 x 106 M-1) and acid-swollen cellulose (CELE(CBD), 1.66 x 106 M-1; XYLA(CBD), 1.73 x 106 M-1). NMR spectra of XYLA(CBD) titrated with cello-oligosaccharides showed that the environment of three tryptophan residues was affected when the CBD bound cellohexaose, cellopentaose or cellotelraose. The K(a) values of the XYLA(CBD) for C6, C5 and C4 cello-oligosaccharides were estimated to be 3.3 x 102, 1.4 x 102 and 4.0 x 101 M-1 respectively, suggesting that the CBD can accommodate at least six glucose molecules and has a much higher affinity for insoluble cellulose than soluble oligosaccharides. Fusion of either the CELE(CBD) or XYLA(CBD) to the catalytic domain of EGE potentiated the activity of the enzyme against insoluble forms of cellulose but not against carboxymethylcellulose. The increase in cellulase activity was not observed when the CBDs were incubated with the catalytic domain of either EGE or XYLA, with insoluble cellulose and a cellulose/hemicellulose complex respectively as the substrates. Pseudomonas CBDs did not induce the extension of isolated plant cell walls nor weaken cellulose paper strips in the same way as a class of plant cell wall proteins called expansins. The XYLA(CBD) and CELE(CBD) did not release small particles from the surface of cotton. The significance of theses results in relation to the mode of action of Type II CBDs is discussed.
Author(s): Bolam D, Ciruela A, McQueen-Mason S, Simpson P, Williamson M, Rixon J, Boraston A, Hazelwood G, Gilbert H
Publication type: Article
Publication status: Published
Journal: Biochemical Journal
Year: 1998
Volume: 331
Issue: 3
Pages: 775-781
Print publication date: 01/05/1998
ISSN (print): 0264-6021
ISSN (electronic): 1470-8728
Publisher: Portland Press Ltd.
PubMed id: 9560304
