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Demonstration of helicase activity in the nonstructural protein, NSs, of the negative-sense RNA virus, Groundnut bud necrosis virus

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Abstract

The nonstructural protein NSs, encoded by the S RNA of groundnut bud necrosis virus (GBNV) (genus Tospovirus, family Bunyaviridae) has earlier been shown to possess nucleic-acid-stimulated NTPase and 5′ α phosphatase activity. ATP hydrolysis is an essential function of a true helicase. Therefore, NSs was tested for DNA helicase activity. The results demonstrated that GBNV NSs possesses bidirectional DNA helicase activity. An alanine mutation in the Walker A motif (K189A rNSs) decreased DNA helicase activity substantially, whereas a mutation in the Walker B motif resulted in a marginal decrease in this activity. The parallel loss of the helicase and ATPase activity in the K189A mutant confirms that NSs acts as a non-canonical DNA helicase. Furthermore, both the wild-type and K189A NSs could function as RNA silencing suppressors, demonstrating that the suppressor activity of NSs is independent of its helicase or ATPase activity. This is the first report of a true helicase from a negative-sense RNA virus.

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Acknowledgments

We thank Sir David Baulcombe (Cambridge University, London) for the gift of N. benthamiana 16c seeds, and Dr. Kazuyuki Mise (Kyoto University, Japan) for the pBIC vectors expressing GFP, dsGFP and P19. We also thank the Department of Science and Technology for the J. C. Bose fellowship to Prof. H. S. Savithri, Department of Biotechnology, and Indian Institute of Science for financial support.

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Author notes

  1. Lokesh Bhushan

    Present address: Cincinnati Children’s Hospital Medical Center, Cincinnati, USA

  2. Vipin Singh Rana

    Present address: Department of Zoology, University of Delhi, New Delhi, 110 007, India

  3. Sunil Kumar Mukherjee

    Present address: Department of Genetics, University of Delhi, New Delhi, 110 0012, India

Authors and Affiliations

  1. Department of Biochemistry, New Biological Sciences, Indian Institute of Science, Bangalore, 560012, Karnataka, India

    Lokesh Bhushan, Ambily Abraham & Handanahal Subbarao Savithri

  2. International Centre for Genetic Engineering and Biotechnology, New Delhi, 110067, India

    Nirupam Roy Choudhury, Vipin Singh Rana & Sunil Kumar Mukherjee

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  1. Lokesh Bhushan
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Correspondence to Handanahal Subbarao Savithri.

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L. Bhushan and A. Abraham contributed equally.

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Bhushan, L., Abraham, A., Choudhury, N.R. et al. Demonstration of helicase activity in the nonstructural protein, NSs, of the negative-sense RNA virus, Groundnut bud necrosis virus. Arch Virol 160, 959–967 (2015). https://doi.org/10.1007/s00705-014-2331-9

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