Cloning, sequencing and analysis of expression of a Butyrivibrio fibrisolvens gene encoding a β-glucosidase

Summary: 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 91800. In Escherichiu 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 94000. 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.