The Sphingomonas paucimobilis β-glucosidase Bgl1 is encoded by the bgl1 gene, associated with an 1308bp open reading frame. The deduced protein has a potential signal peptide of 24 amino acids in the N-terminal region, and experimental evidence is consistent with the processing and export of the Bgl1 protein through the inner membrane to the periplasmic space. A His6-tagged 44.3kDa protein was over-produced in the cytosol of Escherichia coli from a recombinant plasmid, which contained the S. paucimobilis bgl1 gene lacking the region encoding the putative signal peptide. Mature β-glucosidase Bgl1 is specific for aryl-β-glucosides and has no apparent activity with oligosaccharides derived from cellulose hydrolysis and other saccharides. A structure-based alignment established structural relations between S. paucimobilis Bgl1 and other members of the glycoside hydrolase (GH) family 1 enzymes. At subsite —1, the conserved residues required for catalysis by GH1 enzymes are present in Bgl1 with only minor differences. Major differences are found at subsite +1, the aglycone binding site. This alignment seeded a sequence-based phylogenetic analysis of GH1 enzymes, revealing an absence of horizontal transfer between phyla. Bootstrap analysis supported the definition of subfamilies and revealed that Bgl1, the first characterized β-glucosidase from the genus Sphingomonas, represents a very divergent bacterial subfamily, closer to archaeal subfamilies than to others of bacterial origin.

Abbreviations used: CAZy, carbohydrate-active enZymes; DOE-JGI, Department of Energy Joint Genome Institute; GH, glycoside hydrolase; IPTG, isopropyl β-d-thiogalactoside; ORF, open reading frame; PDB, Protein Data Bank; pNPG, p-nitrophenyl β-d-glucopyranoside.

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Author notes

The nucleotide sequence data reported will appear in DDBJ, EMBL, GenBank® and GSDB Nucleotide Sequence Databases under the accession number AF305688 (or Spauc_bgl_AAG59862).