nss encoded by groundnut bud necrosis virus is a bifunctional enzymenss花生芽坏死病毒编码的是一个双功能酶.pdfVIP

NSs Encoded by Groundnut Bud Necrosis Virus Is a Bifunctional Enzyme 1 2 1 1 Bhushan Lokesh , Panigrahi R. Rashmi , Bhat S. Amruta , Dharmaiah Srisathiyanarayanan , Mathur R. N. 2 1 Murthy , Handanahal S. Savithri * 1 Department of Biochemistry, Indian Institute of Science, Bangalore, Karnataka, India, 2 Molecular Biophysics Unit, Indian Institute of Science, Bangalore, Karnataka, India Abstract Groundnut bud necrosis virus (GBNV), a member of genus Tospovirus in the family Bunyaviridae, infects a large number of leguminosae and solanaceae plants in India. With a view to elucidate the function of nonstructural protein, NSs encoded by the small RNA genome (S RNA), the NSs protein of GBNV- tomato (Karnataka) [1] was over-expressed in E. coli and purified by Ni-NTA chromatography. The purified rNSs protein exhibited an RNA stimulated NTPase activity. Further, this activity was metal ion dependent and was inhibited by adenosine 59 (b, c imido) triphosphate, an ATP analog. The rNSs could also hydrolyze dATP. Interestingly, in addition to the NTPase and dATPase activities, the rNSs exhibited ATP independent 59 RNA/ DNA phosphatase activity that was completely inhibited by AMP. The 59 a phosphate could be removed from ssDNA, ssRNA, dsDNA and dsRNA thus confirming that rNSs has a novel 59 a phosphatase activity. K189A mutation in the Walker motif A (GxxxxGKT) resulted in complete loss of ATPase activity, but the 59 phosphatase activity was unaffected. On the other hand, D159A mutation in the Walker motif B (DExx) resulted in partial loss of both the activities. These results demonstrate for the first time that NSs is a bifunctional enzyme, which could particip