Molecular cloning and heterologous expression of an acid stable xylanase gene from Alternaria sp. HB186

  • Liangwei Mao
  • Po Meng
  • Cheng Zhou
  • Lixin Ma
  • Guimin ZhangEmail author
  • Yanhe Ma


A new xylanase gene, named xyn186, was cloned by the genome-walking PCR method from the Alternaria sp. HB186. The sequence of xyn186 contains a 748 bp open reading frame separated by one intron with the size of 52 bp. The cDNA was obtained by DpnI-mediated intron deletion. The cDNA was cloned into pHBM905A and transformed into Pichia pastoris GS115 to screen xylanase-secreting transformants on RBB-xylan plates. The molecular mass of the enzyme was estimated to be 23 kDa on SDS-PAGE. The optimal pH and temperature of the purified enzyme is 6 and 50°C, respectively. The K m and V max valued for birchwood xylan are 1.404 mg ml−1 and 0.2748 mmol min−1 mg−1, respectively. The inhibitory effects of various metal ions were investigated, Cu2+ and Hg2+ ions inhibited most of the enzyme activity. The gene copy number of xyn186 in the genome of P. pastoris was estimated as two by the Real-time PCR. To date, xyn186 gene is the first xylanase gene cloned from the genus Alternaria.


Alternaria sp. HB186 Heterologous expression Pichia pastoris Acid stable xylanase Gene copy number Real-time PCR 



This study was supported by the National Nature Science Foundation (30600014), Hubei Province Nature Science Foundation Key Project, Knowledge Innovative Program of the Chinese Academy of Sciences (KSCX2-EW-G-8) and the Ministry of Sciences and Technology of China (973 programs 2012CB721000).


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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Liangwei Mao
    • 1
  • Po Meng
    • 1
  • Cheng Zhou
    • 2
  • Lixin Ma
    • 1
  • Guimin Zhang
    • 1
    Email author
  • Yanhe Ma
    • 2
  1. 1.College of Life SciencesHubei UniversityWuhanChina
  2. 2.State Key Laboratory of Microbial ResourcesInstitute of Microbiology, Chinese Academy of SciencesBeijingChina

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