Chitosanase could degrade chitosan efficiently under mild conditions to prepare chitosan oligosaccharides (COSs). COS possesses versatile physiological activities and has wide application prospects in food, pharmaceutical and cosmetic fields. Herein, a new glycoside hydrolase (GH) family 46 chitosanase (CscB) was cloned from Kitasatospora setae KM-6054 and heterologously expressed in Escherichia coli. The recombinant chitosanase CscB was purified by Ni-charged magnetic beads and showed a relative molecular weight of 29.19 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). CscB showed the maximal activity (1094.21 U/mg) at pH 6.0 and 30 °C. It was revealed that CscB is a cold-adapted enzyme. CscB was determined to be an endo-type chitosanase with a polymerization degree of the final product mainly in the range of 2-4. This new cold-adapted chitosanase provides an efficient enzyme tool for clean production of COSs.