Expression of serine proteinase P186 of Arthrobotrys oligospora and analysis of its nematode-degrading activity

文献类型： 外文期刊

第一作者： Zhao, Hailong

作者： Zhao, Hailong;Qiao, Jun;Meng, Qingling;Gong, Shasha;Chen, Cheng;Liu, Tianli;Tian, Lulu;Chen, Chuangfu;Cai, Xuepeng;Luo, Jianxun

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关键词： Arthrobotrys oligospora;Pichia pastoris;Eukaryotic expression;Serine proteinase P186;Nematode-degrading activity

期刊名称：ANTONIE VAN LEEUWENHOEK INTERNATIONAL JOURNAL OF GENERAL AND MOLECULAR MICROBIOLOGY （ 影响因子：2.271； 五年影响因子：2.707 ）

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收录情况： SCI

摘要： The nematode-trapping fungi possess a unique capability of predating and invading nematodes. As a representative nematode-trapping fungus, Arthrobotrys oligospora has been widely used to study the interactions between nematode-trapping fungi and their hosts. Serine proteinase is one of the important virulence factors during process of invasion of the nematode-trapping fungi into nematodes. In this study, using reverse transcription polymerase chain reaction, we amplified the gene sequence of serine proteinase 186 from A. oligospora, cloned it into pPIC9K vector and expressed it in the yeast Pichia pastoris. The expressed recombinant serine proteinase186 (reP186) was purified via Ni-affinity chromatography. The in vitro nematode-degrading activity of reP186 was analyzed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot analysis revealed that reP186 with molecular weight of 33 kDa was successfully obtained. ReP186 was capable of degrading a series of protein substrates including casein, gelatin, bovine serum albumin, denatured collagen and nematode cortical layer. The reP186 exhibited the maximal activity at pH 8.0 and 55 A degrees C and was highly sensitive to the inhibitor, phenylmethanesulfonylfluoride. Treatment of Caenorhabditis elegans and Haemonchus contortus with reP186 for 12, 24 and 36 h, respectively, resulted in 62, 88 and 100 % of killing rates for C. elegans, and 52, 65 and 84 % of killing rates for H. contortus, respectively, indicating a relatively strong nematode-degrading bioactivity of reP186.

分类号： Q1

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