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Purification and identification of an antibacterial protein from the symbiotic bacteria associated with novel entomopathogenic nematode, Rhabditis (Oscheius) sp.

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Abstract

Entomopathogenic nematodes (EPN) belonging to the families steinernematidae and heterorhabditidae and their symbiotic bacteria Xenorhabdus and Photorhabdus are well-known as biological control agents and are found to produce a wide range of bioactive secondary metabolites. Studies carried out at the Central Tuber Crops Research Institute (CTCRI) on entomopathogenic nematodes resulted in the identification of novel EPN belonging to the family Rhabditidae. This study reports the purification of a high molecular weight antibacterial protein from culture filtrates of a bacterium (Bacillus cereus) symbiotically associated with a novel entomopathogenic nematode Rhabditis (Oscheius) species, maintained at CTCRI laboratory. Fermentation conditions were standardized and optimum antibacterial activity was observed in tryptic soy broth after 48 h incubation at 30 °C. The aqueous extracts yielded antibacterial proteins which were purified by ammonium sulfate precipitation followed by ion exchange chromatography and size exclusion chromatography. Native gel electrophoresis indicated an active protein of molecular mass 220KDa which resolved into a major band of 90 kDa and a minor band of about 40 kDa on SDS-PAGE. The 90 kDa protein showed antibacterial activity and was further analysed by MALDI TOF—MS/MS. The protein was identified as a TQXA (Threonine–glutamine dipeptide) domain containing protein from Bacillus cereus. The protein was found to be active against Bacillus subtilis MTCC2756, Staphylococus aureus MTCC902 and Escherichia coli MTCC 2622 and was thermally stable.

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Acknowledgments

The authors are grateful to the Director, Central Tuber Crops Research Institute (CTCRI) for providing facilities and to Life Science Research Board (LSRB) for financial support. K. M. Anju also availed University Junior Research fellowship (JRF) from University of Kerala.

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Authors and Affiliations

  1. Division of Crop Protection, Central Tuber Crops Research Institute, Thiruvananthapuram, 695 017, Kerala, India

    K. M. Anju, M. M. Archana & C. Mohandas

  2. Division of Crop Utilization, Central Tuber Crops Research Institute, Thiruvananthapuram, India

    Bala Nambisan

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  1. K. M. Anju
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  2. M. M. Archana
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  3. C. Mohandas
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  4. Bala Nambisan
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Correspondence to K. M. Anju.

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Anju, K.M., Archana, M.M., Mohandas, C. et al. Purification and identification of an antibacterial protein from the symbiotic bacteria associated with novel entomopathogenic nematode, Rhabditis (Oscheius) sp.. World J Microbiol Biotechnol 31, 621–632 (2015). https://doi.org/10.1007/s11274-015-1816-3

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  • DOI: https://doi.org/10.1007/s11274-015-1816-3

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