Gene Cloning, Expression, and Characterization of a Second β-N-Acetylglucosaminidase from the Chitinolytic Bacterium Aeromonas hydrophila Strain SUWA-9

A gene coding for a second β-N-acetylglucosaminidase (nagB) was isolated from the chitinolytic bacterium Aeromonas hydrophila strain SUWA-9. The nagB open reading frame encoded a polypeptide of 618 amino acid residues with a molecular mass of 68.8 kDa. It did not contain a sequence characteristic of a signal peptide at the N-terminus. The deduced amino acid sequence showed high similarity to those of bacterial β-N-acetylhexosaminidases classified into family 20 of glycosyl hydrolases. The nagB gene was successfully expressed in Escherichia coli, and the recombinant protein hydrolyzed N-acetylchitooligomers from dimer to hexamer and produced monomer as a final product. Reverse transcription-mediated PCR (RT-PCR) analysis revealed that nagB was transcribed when SUWA-9 cells were grown in the presence of colloidal chitin. In the upstream of the nagB gene, three genes, coding for putative N-acetylglucosamine kinase, β-glucosidase, and ATP-binding protein of ABC-type transporter, were identified, and these gen...