Chitosanase (EC 3.2.1.132) is an important chitosan-degrading enzyme involved in industrial applications. In this study, a chitosanase gene (BbCSN-1) from Beauveria bassiana, an insect fungal pathogen, was cloned and expressed in Pichia pastoris. The amount of BbCSN-1 in the fermentation broth of P. pastoris gradually increased after induction with methanol from one to 6 d, reaching 398 μg/ml on the 6th day. The molecular characteristics of BbCSN-1 were measured with colloidal chitosan as a substrate. The purified BbCSN-1 exhibited optimum activity at pH 5 and 30 °C and was stable at pH 2-8 and below 40 °C. The Km value of BbCSN-1 was approximately 0.8 mg/ml at 30 °C (pH 6.0). The activity of BbCSN-1 was significantly enhanced by Mn2+ but inhibited by Co2+ and Cu2+. These results indicated that BbCSN-1 from B. bassiana could be easily expressed in P. pastoris, which provided a basis for further study on its application.

中文翻译：

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球孢白僵菌壳聚糖酶（BbCSN-1）在毕赤酵母中的表达和重组蛋白的酶促分析。

壳聚糖酶（EC 3.2.1.132）是一种重要的壳聚糖降解酶，涉及工业应用。在这项研究中，克隆了一种来自球孢白僵菌（Beauveria bassiana）的壳聚糖酶基因（BbCSN-1），它是一种昆虫真菌病原体，并在巴斯德毕赤酵母中表达。甲醇诱导发酵后，毕赤酵母发酵液中BbCSN-1的含量从1天到6 d逐渐增加，第6天达到398μg/ ml。以胶体壳聚糖为底物测量了BbCSN-1的分子特性。纯化的BbCSN-1在pH 5和30°C下显示最佳活性，在pH 2-8和40°C以下稳定。BbCSN-1的Km值在30°C（pH 6.0）下约为0.8 mg / ml。Mn2 +显着增强了BbCSN-1的活性，但Co2 +和Cu2 +抑制了BbCSN-1的活性。这些结果表明来自B的BbCSN-1。