Groundnut bud necrosis virus encoded movement protein NSm binds to GTP and ATP

Groundnut bud necrosis virus (GBNV) is a tripartite negative sense RNA virus that belongs to tospoviridae family. The M RNA encodes non-structural protein-m (NSm), a movement protein in tospoviruses. In this communication, we demonstrate that, GBNV NSm interacts with ATP and GTP. UV crosslinking with [γ-32P] ATP indicates that GBNV NSm forms two distinct complexes with ATP one of them is Mg²⁺ dependent and the other is Mg²⁺ independent. It also binds to ATP- and GTP-coupled agarose resin and shows competition with free ATP and GTP but not with UTP and CTP. The NSm-NTP interaction was further validated by intrinsic fluorescence quenching studies. NTPs and dNTPs both could quench the intrinsic fluorescence of NSm. However, maximum quenching of fluorescence occurred in the presence of GTP, followed by ATP, suggesting that it is the preferred ligand. The extent of fluorescence quenching with different concentrations of GTP was used to calculate the binding constant, and it was found to be 3 μM, lower than that reported for other proteins that can bind NTP. This is the first report of the GTP and ATP binding property of NSm from any Tospoviruses. Further, NSm could also hydrolyze GTP. Preliminary sequence analysis suggests the presence of two putative atypical Walker A motif from amino acid sequences 51-58 and 267-274, indicating that this sequence might be involved in NTP binding. This motif is conserved in most of the tospoviruses. NSm from GBNV an Asian clade, localize to ER network and remodels it to vesicles which has been proposed to be involved in movement through plasmodesmata (PD). Therefore, GTP-NSm interaction might be involved in signaling cell to cell trafficking.