Molecular identification of a novel [beta]-1,3-glucanase from alkaliphilic Nocardiopsis sp. strain F96

Alkaliphilic Nocardiopsis sp. strain F96 produced three β-1,3-glucanase isozymes of different molecular masses (BglF1, BglF2 and BglF3). The N-terminal amino acid sequences of BglFs indicated that these isozymes were the products of a single gene. The β-1,3-glucanase gene (bglF) was cloned from the chromosomal DNA of strain F96. The bglF gene encoded a polypeptide of 270 amino acids including a signal sequence. The deduced amino acid sequence of mature BglF exhibited the highest homology to those of glycoside hydrolase (GH) family 16 β-1,3-glucanases, suggesting that the enzyme belonged to the GH family 16. The mature region of bglF gene was functionally expressed in Escherichia coli. The optimum pH and temperature of purified recombinant BglF were pH 9.0 and 70°C, respectively. This enzyme efficiently hydrolyzed insoluble β-1,3-glucans and showed the highest activity toward a β-1,3-1,4-glucan rather than β-1,3-glucans. These results suggested that BglF would be a novel β-1,3-glucanse. Mutational analysis revealed that Glu123 and Glu128 should be the catalytic residues of BglF.[PUBLICATION ABSTRACT]