FOOD SCIENCE ›› 2019, Vol. 40 ›› Issue (14): 99-106.doi: 10.7506/spkx1002-6630-20180809-086

• Bioengineering • Previous Articles     Next Articles

High-Level Expression of Bacillus subtilis Chitosanase in Pichia pastoris and Its Hydrolytic Properties

MA Shuai, YANG Shaoqing, LIU Yihao, YAN Qiaojuan, JIANG Zhengqiang   

  1. 1. Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China; 2. College of Engineering, China Agricultural University, Beijing 100083, China
  • Online:2019-07-25 Published:2019-07-23

Abstract: In this study, the gene encoding chitosanase (BsCsn46) was cloned from Bacillus subtilis and expressed at high level in Pichia pastoris GS115. The enzymatic and hydrolytic properties of BsCsn46 were further investigated. The extracellular chitosanase activity and protein concentration was up to 50 370 U/mL and 15.7 mg/mL after the high cell-density fermentation of the recombinant P. pastoris in a 5 L fermentor, respectively. BsCsn46 was purified by ion-exchange chromatography using Q-Sepharose Fast Flow. The purified enzyme had a high specific activity of 4 065.7 U/mg towards chitosan and showed maximal activity at pH 6.0 and 55 ℃, and it was stable at temperature below 45 ℃. The hydrolysis of 3 g/100 mL chitosan by BsCsn46 yielded predominantly chitooligosaccharides with degree of polymerization (DP) 2–4. The degree of hydrolysis reached 92.8% and the yield of chitooligosaccharides was up to 90.9%. The high enzyme production level and the remarkable hydrolysis efficiency make BsCsn46 a promising candidate for application in the industrial preparation of chitosanase and chitooligosaccharides.

Key words: Bacillus subtilis, chitosanase, high-level expression, enzymatic properties, chitooligosaccharides

CLC Number: