CLONING, SEQUENCING AND ANALYSIS OF EXPRESSION OF A BUTYRIVIBRIO-FIBRISOLVENS GENE ENCODING A BETA-GLUCOSIDASE

The cloning, expression and nucleotide sequence of a 3.74 kb DNA segment on pLS215 containing a β-glucosidase gene (bglA) from Butyrivibrio fibrisolvens H17c was investigated. The B. fibrisolvens bglA open reading frame (ORF) of 2490 bp encoded a β-glucosidase of 830 amino acid residues with a calculated M(r) of 91.800. In Escherichia coli C600(pLS215) cells the β-glucosidase was localized in the cytoplasm and these cells produced an additional protein with an apparent M(r) of approximately 94.000. The bglA gene was expressed from its own regulatory region in E. coli and a single mRNA initiation point was identified upstream of the bglA ORF and adjacent to a promoter consensus sequence. The primary structure of the β-glucosidase showed > 40% similarity with a domain of 237 amino acids present in the β-glucosidases of Kluyveromyces fragilis and Clostridium thermocellum. The B. fibrisolvens β-glucosidase hydrolysed cellobiose to a limited extent, cellotriose to cellobiose and glucose, and cellotetraose and cellopentaose to predominantly glucose.