Identification of a novel GH13 β-galactosidase from Photobacterium gaetbulicola

Abstract:

[Background] Glycoside hydrolase family 13 (GH13), known as the largest α-amylase family, does not contain galactosidases. [Objective] To identify a protein BgalPg from the marine bacterium Photobacterium gaetbulicola. [Methods] The family of BgalPg was identified by conserved motif analysis and a phylogenetic analysis. The BgalPg gene was cloned and expressed in Escherichia coli. The enzymatic properties of purified BgalPg were determined. [Results] BgalPg possessing the conserved motifs of GH13 family is a new member of the GH13_38 subfamily, although it showed low sequence identity with characterized α-amylase family proteins. BgalPg demonstrated no catalytic activity against any substrates of the α-amylase family, while it had the activities towards pNP-β-d-galactopyranoside [pNP-β-Gal, (2.8±0.4) U/mg] and oNP-β-d-galactopyranoside [oNP-β-Gal, (1.4±0.3) U/mg]. Moreover, the enzyme hydrolyzed lactose [(0.40±0.01) U/mg], which represented a typical β-galactosidase activity. BgalPg exhibited high stability in pH 7.0–8.5 and the half-life time of 1.5 h at 60 ℃. [Conclusion] This is the first time that an enzyme of GH13 family was biochemically defined as a novel β-galactosidase.