Production and purification of a protease, a chitosanase, and chitin oligosaccharides by Bacillus cereus TKU022 fermentation.

A protease- and chitosanase-producing strain was isolated and identified as Bacillus cereus TKU022. The protease and chitosanase were both produced using 1.5% (w/v) shrimp head powder (SHP) as the sole carbon/nitrogen source, and these enzymes were purified from the culture supernatant. The molecular masses of the TKU022 protease and chitosanase determined using SDS-PAGE were approximately 45 and 44kDa, respectively. The high stability of the TKU022 protease toward surfactants, an optimal pH of 10 and an optimal temperature of 50-60°C suggest that this high-alkaline protease has potential applications for various industrial processes. Concomitant with the production of the TKU022 chitosanase, N-acetyl chitooligosaccharides were also observed in the culture supernatant, including (GlcNAc)(2), (GlcNAc)(4), (GlcNAc)(5), and (GlcNAc)(6) at concentrations of 201.5, 12.4, 0.5, and 0.3μg/mL, respectively, as determined using an HPLC analysis. The chitin oligosaccharides products were also characterized using a MALDI-TOF mass spectrometer. A combination of the HPLC and MALDI-TOF MS results showed that the chitin oligosaccharides of the TKU022 culture supernatant comprise oligomers with degree of polymerization (DP) from 2 to 6. Using this method, the production of a protease, a chitosanase, and chitin oligosaccharides may be useful for various industrial and biological applications.