The gene fragment encoding the cellulose-binding domain (CBD) of an exoglucanase (Cex) from Cellulomonas fimi was subcloned and expressed in Escherichia coli. Transcription from the lac promoter coupled with translation from a consensus prokaryotic ribosome binding site led to the production of large quantities of CBDCex (up to 25% total soluble cell protein). The polypeptide leaked into the culture supernatant (up to 50 mg · L−1), facilitating one-step purification by affinity chromatography on cellulose. The 11-kDa polypeptide reacted with Cex antiserum. Absence of free thiols indicated that the two Cys residues of CBDCex form a disulfide bridge. It had the same N-terminal amino acid sequence as CBDCex prepared from Cex by proteolysis, plus two additional N-terminal amino acid residues (Ala and Ser) encoded by the Nhel site introduced during plasmid construction. CBDCex bound to a variety of β-1, 4-glycans with different affinities and saturation levels. Adsorption to bacterial microcrystalline cellulose was dependent on the temperature, but not on the pH. © 1993 John Wiley & Sons, Inc.
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