Abstract
From the culture filtrate of the fungus Dactylellina varietas (syn. Dactylella varietas), an extracellular protease (designed Dv1) was purified by cation exchange and hydrophobic interaction chromatography. The purified protease showed a molecular mass of approximately 30 kDa and displayed an optimal activity at pH 8 and 60.5°C (more than 20 min). This protease could degrade a broad range of substrates including casein, gelatin, BSA (bovine serum albumin), and nematode cuticle. However, its proteolytic activity was highly sensitive to the serine protease inhibitor Phenylmethylphonylfuoride (1 mM), indicating that it belongs to the serine-type peptidase group. This protease could immobilize the free-living nematodes Panagrellus redivivus and Caenorhabditis elegans and hydrolyze the purified cuticle of P. redivivus, suggesting it may play a role in infection against nematodes. The encoding gene of Dv1 and its promoter sequence were cloned using degenerate primers and the DNA walking technology. Its open-reading frame contains 1,224 base pairs and without any intron. The deduced amino-acid sequence shared low identity to serine proteases from other nematode-trapping fungi. Our report identified a novel pathogenic protease from the nematode-trapping fungus D. varietas, and the three-dimensional structure of this protease was predicted using the Swiss-Prot method.
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Acknowledgement
We thank Drs. Chenggang Zhou, Xiaowei Huang and Ms Wei Zhou for their help and advice in our studies and in preparing this manuscript. This work was funded by projects from the National Natural Science Foundation of China (approved nos. 30630003 and 30660107), by the Department of Science and Technology of Yunnan Province (approval Nos. 2004C0001Z and 2005NG05) and by Yunnan University (grant no. 2005Q008B).
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Jinkui Yang and Lianming Liang contributed equally to this work.
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Yang, J., Liang, L., Zhang, Y. et al. Purification and cloning of a novel serine protease from the nematode-trapping fungus Dactylellina varietas and its potential roles in infection against nematodes. Appl Microbiol Biotechnol 75, 557–565 (2007). https://doi.org/10.1007/s00253-007-0839-6
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DOI: https://doi.org/10.1007/s00253-007-0839-6