Biotechnology Letters

, Volume 34, Issue 4, pp 689–694

High-level expression and characterization of a highly thermostable chitosanase from Aspergillus fumigatus in Pichia pastoris

Authors

  • Xiaomei Chen
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
  • Chao Zhai
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
  • Lixin Kang
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
  • Chunhua Li
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
    • Hubei Key Laboratory of Genetic Regulation and Integrative Biology, College of Life SciencesHuazhong Normal University
  • Hong Yan
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
  • Yuling Zhou
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
  • Xiaolan Yu
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
    • Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei University
Original Research Paper

DOI: 10.1007/s10529-011-0816-0

Cite this article as:
Chen, X., Zhai, C., Kang, L. et al. Biotechnol Lett (2012) 34: 689. doi:10.1007/s10529-011-0816-0

Abstract

The sequence of an endo-chitosanase gene (CSN) from Aspergillus fumigatus was optimized based on the preferred codons of Pichia pastoris and synthesized in vitro through overlapping PCR (CSN-P). The gene was cloned into a yeast expression vector, pHBM905A, and secretorily expressed in Pichia pastoris GS115. The yield of CSN-P reached ~3 mg/ml with a high-density fermentation in a 14 l fermenter and the enzyme activity was ~25,000 U/ml. The enzyme had half-lives of 2.5 h at 80°C, 1 h at 90°C and 32 min at 100°C. It retained 70% activity after incubation with 10 M urea at room temperature for 30 min. This enzyme was used for a large-scale preparation of oligosaccharides: 3 g enzyme converted 200 kg chitosan into oligosaccharides in 24 h at 60°C.

Keywords

ChitosanaseHigh-density fermentationPichia pastorisThermostability

Supplementary material

10529_2011_816_MOESM1_ESM.doc (44 kb)
Supplementary material 1 (DOC 45 kb)
10529_2011_816_MOESM2_ESM.doc (58 kb)
Supplementary material 2 (DOC 58 kb)
10529_2011_816_MOESM3_ESM.doc (370 kb)
Supplementary material 3 (DOC 370 kb)

Copyright information

© Springer Science+Business Media B.V. 2011