Novel Alkali-Stable, Cellulase-Free Xylanase from Deep-Sea Kocuria sp. Mn22

A novel xylanase gene, Kxyn, was cloned from Kocuria sp. Mn22, a bacteria isolated from the deep sea of the east Pacific. Kxyn consists of 1,170 bp and encodes a protein of 390 amino acids that shows the highest identity (63%) with a xylanase from Thermobifida fusca YX. The mature protein with a molecular mass of approximately 40 kDa was expressed in Escherichia coli BL21 (DE3). The recombinant Kxyn displayed its maximum activity at 55°C and at pH 8.5. The K m , V max , and k cat values of Kxyn for birchwood xylan were 5.4 mg/ml, 272 μmol/min·mg, and 185.1/s, respectively. Kxyn hydrolyzed birchwood xylan to produce xylobiose and xylotriose as the predominant products. The activity of Kxyn was not affected by Ca 2+ , Mg 2+ , Na + , K + , β-mercaptoethanol, DTT, or SDS, but was strongly inhibited by Hg 2+ , Cu 2+ , Zn 2+ , and Pb 2+ . It was stable over a wide pH range, retaining more than 80% activity after overnight incubation at pH 7.5―12. Kxyn is a cellulase-free xylanase. Therefore, these properties make it a candidate for various industrial applications.