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UBC Theses and Dissertations

Characterization of cbg : a cloned gene encoding an extracellular [beta]-glucosidase from Cellulomonas fimi Bates, Nancy Carol


A group of Escherichia coli clones harbouring recombinant pBR322 plasmid, containing Cellulomonas fimi DNA inserts, that reacted with antiserum to C.fimi culture supernatant, was screened for their ability to hydrolyze carboxymethyl cellulose (CMC) and 4-methylumbeliferyll-β-D-cellobioside (MUC). A clone, pEC62, hydrolyzed MUC but did not hydrolyze CMC. The recombinant enzyme encoded by pEC62 was shown to be a β-glucosidase (cellobiase). C.fimii itself was shown to encode an extracellular β-glucosidase in C.fimi. This is the first report of an extracellular β-glucosidase from a bacterium. Deletion analysis localized the cloned gene (cbg)to the tet promoter proximal region of the 7.0 kilobase insert of pEC62. Further analysis and sequence data showed a highly active derivative of pEC62 contained a translational gene fusion between lacZ of pUC13 and cbg. From this data, a location for the cbg start site was proposed.

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