Nipah virus (NiV) and related viruses form a distinct henipavirus genus within the Paramyxoviridae family. NiV continues to spillover into the humans causing deadly outbreaks with increasing human-bat interaction. NiV encodes the large protein (L) and phosphoprotein (P) to form the viral RNA polymerase machinery. Their sequences show limited homologies to those of non-henipavirus paramyxoviruses. We report two cryo-electron microscopy (cryo-EM) structures of the Nipah virus (NiV) polymerase L-P complex, expressed and purified in either its full-length or truncated form. The structures resolve the RNA-dependent RNA polymerase (RdRp) and polyribonucleotidyl transferase (PRNTase) domains of the L protein, as well as a tetrameric P protein bundle bound to the L-RdRp domain. L-protein C-terminal regions are unresolved, indicating flexibility. Two PRNTase domain zinc-binding sites, conserved in most Mononegavirales, are confirmed essential for NiV polymerase activity. The structures further reveal anchoring of the P protein bundle and P protein X domain (XD) linkers on L, via an interaction pattern distinct among Paramyxoviridae. These interactions facilitate binding of a P protein XD linker in the nucleotide entry channel and distinct positioning of other XD linkers. We show that the disruption of the L-P interactions reduces NiV polymerase activity. The reported structures should facilitate rational antiviral-drug discovery and provide a guide for the functional study of NiV polymerase.
Nipah Virus/chemistry* ; Cryoelectron Microscopy ; Viral Proteins/genetics* ; RNA-Dependent RNA Polymerase/genetics* ; Phosphoproteins/genetics* ; Humans ; Models, Molecular ; Protein Binding

In this study, Recombinant baculoviruses rBac-NF and rBac-NG were generated for expressing F and G proteins Nipah virus (NiV) . The expression of recommbinant G (rNG) and F (rNF) protein in rBac-NF and rBac-NG infected cells were confirmed by western-blot. Both rNG and rNF showed sensitive and specific antigenic reaction to rabbit serum anti-Nipah virus in indirect immunofluorescence detection and indirect ELISA. Immunization with rBac-NF and rBac-NG infected insect cells elicited G ad F protein specific antibody responses in mice. Furthermore, the G ad F specific antibodies could neutralize the infectivity of the VSVdeltaG* F/G, the NiV F and G envelope glycoproteins psudotyped recombinant Vesicular Stomatitis Virus expressing green fluorescence protein. The results demonstrated F and G protein expressed by the recombinant baculoviruses could be safe economic diagnostic antigens for the surveillance and monitoring of NiV and promising subunit vaccines for the prevention of NiV.
Animals ; Antigens, Viral ; immunology ; Baculoviridae ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; Nipah Virus ; chemistry ; genetics ; Rabbits ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Recombination, Genetic ; Viral Envelope Proteins ; biosynthesis ; genetics ; immunology