Gene cloning and bioactivity analysis of chitinase gene AO-483 from Arthrobotrys oligospora XJ-A1

doi:10.3969/j.issn.1004-1524.2019.02.07

›› 2019, Vol. 31 ›› Issue (2): 222-228.DOI: 10.3969/j.issn.1004-1524.2019.02.07

• Animal Science • Previous Articles Next Articles

Gene cloning and bioactivity analysis of chitinase gene AO-483 from Arthrobotrys oligospora XJ-A1

GONG Shasha¹, MENG Qingling^{1, *}, QIAO Jun¹, ZHONG Wenqiang¹, HUANG Yunfu¹, ZHANG Guowu¹, CHEN Ying¹, CAI Xuepeng²

1.Department of Animal Science & Technology, Shihezi University, Shihezi 832003, China;;
2. Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China

Received:2018-04-20 Online:2019-02-25 Published:2019-03-06

Abstract

Abstract: In order to study the functions of chitinase, chitinase gene AO-483 of nematode-trapping fungi Arthrobotrys oligospora XJ-A1 was cloned, analyzed and expressed in prokaryotic expression system Escherichia coli. The pure prokaryotic expression recombinant fusion protein was purified by Ni column and the chitinase activity was determined by DNS method, and the biological activity of recombinant protein AO-483 was verified by dealing with larvae of Caenorhabditis elegans. The results showed that chitinase AO-483 gene encoded 309 amino acids, the homology of its gene sequence was 96.88% and the amino acid sequence was 97.73% respectively with Arthrobotrys oligospora standard strain (ATCC 24927). The chitinase AO-483 belonged to the family 18 glycoside hydrolase with a Glyco-hydro-18 structure domained at amino acid sequence of 20~297 and had two prevalent conserved catalytic domained with the sequence of GMLGG and LDGLDLDVE. The tertiary structure was a typical triosephosphate isomerase barrel structure. The protein identified by SDS-PAGE showed that the recombinant protein had a molecular weight of about 52 ku which was consistent with prediction. Western blot revealed that recombinant protein can specifically react with anti-Arthrobotrys oligospora polyclonal antibody. The activity of recombinant chitinase AO-483 was determined by the DNS method, with a value of 223.31 U·mL^-1. The recombinant AO-483 owned stronger biological activity that can degrade Caenorhabditis elegans larvae stage 1and stage 4.

Key words: Arthrobotrys oligospora, chitinase, gene cloning, biological activity

CLC Number:

GONG Shasha, MENG Qingling, QIAO Jun, ZHONG Wenqiang, HUANG Yunfu, ZHANG Guowu, CHEN Ying, CAI Xuepeng. Gene cloning and bioactivity analysis of chitinase gene AO-483 from Arthrobotrys oligospora XJ-A1[J]. , 2019, 31(2): 222-228.

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Gene cloning and bioactivity analysis of chitinase gene AO-483 from Arthrobotrys oligospora XJ-A1

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