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Applied Microbiology and Biotechnology

, Volume 75, Issue 2, pp 319–328 | Cite as

Cloning and identification of novel cellulase genes from uncultured microorganisms in rabbit cecum and characterization of the expressed cellulases

  • Yi Feng
  • Cheng-Jie Duan
  • Hao Pang
  • Xin-Chun Mo
  • Chun-Feng Wu
  • Yuan Yu
  • Ya-Lin Hu
  • Jie Wei
  • Ji-Liang Tang
  • Jia-Xun FengEmail author
Biotechnologically Relevant Enzymes and Proteins

Abstract

A metagenomic cosmid library was prepared in Escherichia coli from DNA extracted from the contents of rabbit cecum and screened for cellulase activities. Eleven independent clones expressing cellulase activities (four endo-β-1,4-glucanases and seven β-glucosidases) were isolated. Subcloning and sequencing analysis of these clones identified 11 cellulase genes; the encoded products of which shared less than 50% identities and 70% similarities to cellulases in the databases. All four endo-β-1,4-glucanases and all seven β-glucosidases, respectively, belonged to glycosyl hydrolase family 5 (GHF 5) and family 3 (GHF 3) and formed two separate branches in the phylogenetic tree. Ten of the 11 cloned cellulases exhibited highest activities at pH 5.5 ∼ 7.0 and 40 ∼ 55°C, a condition similar to that in the rabbit cecum. All the four endo-β-1,4-glucanases could hydrolyze a wide range of β-1,4-, β-1,4/β-1,3- or β-1,3/β-1,6-linked polysaccharides. One endo-β-1, 4-glucanase gene, umcel5G, was overexpressed in E. coli, and the purified recombinant enzyme was characterized in detail. The enzymes cloned in this work represented at least some of the cellulases operating efficiently in the rabbit cecum. This work provides the first snapshot on the cellulases produced by bacteria in rabbit cecum.

Keywords

Cellulase Cloning Metagenome Rabbit cecum Expression Enzyme Properties 

Notes

Acknowledgment

This work was supported by the National Natural Science Foundation of China (30560003), International Science and Technology Collaborating Key Project Program of China (2002AA217121) and Program for New Century Excellent Talents in University of China (NCET-05-0752). The authors are grateful to Kunio Ohmiya and Shuichi Karita for their helpful discussions.

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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Yi Feng
    • 1
  • Cheng-Jie Duan
    • 1
  • Hao Pang
    • 1
  • Xin-Chun Mo
    • 1
  • Chun-Feng Wu
    • 1
  • Yuan Yu
    • 1
  • Ya-Lin Hu
    • 1
  • Jie Wei
    • 1
  • Ji-Liang Tang
    • 1
  • Jia-Xun Feng
    • 1
    • 2
    Email author
  1. 1.Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, The Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, and College of Life Science and TechnologyGuangxi UniversityNanningPeople’s Republic of China
  2. 2.College of Life Science and TechnologyGuangxi UniversityNanningPeople’s Republic of China

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