Purification and Characterization of Cold-active β-N-Acetylglucosaminidase from Pseudoalteromonas issachenkonii KOPRI 22718

Abstract

136 marine bacteria showing chitinolytic activity were isolated from 47 sediment samples of Kara Sea, Arctic. Among them, a psychrotolerant strain (KOPRI 22718) was selected for its cold-activity at 5℃. The sequence analysis of 16S rRNA affiliates KOPRI 22718 to Pseudoalteromonas issachenkonii. An exo-acting chitinase (~100 kDa) was homogeneously purified from the culture supernatant of KOPRI 22718 through ion exchange and gel filtration chromatography. The purified chitinase (W-Chi22718) exhibited the highest activity toward pNP-GlcNAc and produced GlcNAc monomer as end-product from chitin oligosaccharides (GlcNAc2-GlcNAc6), due to its β-N-acetylglucosaminidase activity. W-Chi22718 displayed chitinase activities at 0-37 ℃ (optimal temperature of 30 ℃), and maintained its activities at pH 6.0-9.0 (optimal pH of 7.6). Interestingly, W-Chi22718 exhibited a relative activity of 13 and 35% at 0 and 10 ℃, respectively, in comparison to 100% at optimal 30 ℃, which is comparable with those of previously characterized, colad-adapted, bacterial chitinases. Among the main cations and protein denaturing reagents, W-Chi22718 activity could be enhanced by K+, Ca+, and Fe2+, but completely inhibited Cu2+ and SDS.