Biotechnology Letters

, Volume 34, Issue 4, pp 689–694 | Cite as

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

  • Xiaomei Chen
  • Chao Zhai
  • Lixin Kang
  • Chunhua Li
  • Hong Yan
  • Yuling Zhou
  • Xiaolan Yu
  • Lixin Ma
Original Research Paper

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

Chitosanase High-density fermentation Pichia pastoris Thermostability 

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)

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Xiaomei Chen
    • 1
  • Chao Zhai
    • 1
  • Lixin Kang
    • 1
  • Chunhua Li
    • 1
    • 2
  • Hong Yan
    • 1
  • Yuling Zhou
    • 1
  • Xiaolan Yu
    • 1
  • Lixin Ma
    • 1
  1. 1.Hubei Key Laboratory of Industrial Biotechnology, College of Life SciencesHubei UniversityWuhanPeople’s Republic of China
  2. 2.Hubei Key Laboratory of Genetic Regulation and Integrative Biology, College of Life SciencesHuazhong Normal UniversityWuhanPeople’s Republic of China

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