Identification, expression, and characterization of a marine-derived chitinase Ce0303 from Chitiniphilus eburneus YS-30 with exo- and endo-hydrolytic properties.

N-acetyl-oligosaccharides exhibit antioxidant and antibacterial activities. However, the low catalytic efficiency of chitinase on crystalline chitin hinders the eco-friendly production of N-acetyl-oligosaccharides. A marine-derived chitinase-producing strain Chitiniphilus eburneus YS-30 was screened in this study. The genome of C. eburneus YS-30 spans 4,522,240 bp, with a G + C content of 63.96 % and 4244 coding genes. Among the chitinases secreted by C. eburneus YS-30, Ce0303 showed the highest content at 19.10 %, with a molecular weight of 73.5 kDa. Recombinant Ce0303 exhibited optimal activity at 50 °C and pH 5.0, maintaining stability across pH 4.0-10.0. Ce0303 demonstrated strict substrate specificity, with a specific activity toward colloidal chitin of 6.41 U mg, K of 2.34 mg mL, and k of 3.27 s. The specific activity of Ce0303 toward α-chitin was 18.87 % of its activity on colloidal chitin. Ce0303 displayed both exo- and endo-hydrolytic properties, primarily producing (GlcNAc) from colloidal chitin. The structure of Ce0303 includes one catalytic domain and two chitin-binding domains. Docking results revealed that the GlcNAc at -1 subsite formed two hydrogen bonds with conserved Trp380. The hydrolytic properties of Ce0303 will provide technical support for the comprehensive utilization of crustacean raw materials.