Abstract
Chandipura virus (CHAV), a member of the vesiculovirus genus, is an emerging human pathogen. As for other rhabdoviruses, CHAV entry into susceptible cells is mediated by its single envelope glycoprotein G which is both involved in receptor recognition and fusion of viral and cellular membranes. Here, we have characterized the fusion properties of CHAV-G. As for vesicular stomatitis virus (VSV, the prototype of the genus) G, fusion is triggered at low pH below 6.5. We have also analyzed the biochemical properties of a soluble form of CHAV-G ectodomain (CHAV-Gth, generated by thermolysin limited-proteolysis of recombinant VSV particles in which the G gene was replaced by that of CHAV). The overall behavior of CHAV-Gth is similar to that previously reported for VSV-Gth. Particularly, CHAV-Gth pre-fusion trimer is not stable in solution and low-pH-induced membrane association of CHAV-Gth is reversible. Furthermore, CHAV-Gth was crystallized in its low pH post-fusion conformation and its structure was determined at 3.6Å resolution. An overall comparison of this structure with the previously reported VSV-Gth post-fusion conformation, shows a high structural similarity as expected from the comparison of primary structure. Among the three domains of G, the pleckstrin homology domain (PHD) appears to be the most divergent and the largest differences are confined to the secondary structure of the major antigenic site of rhabdoviruses. Finally, local differences indicate that CHAV has evolved alternate structural solutions in hinge regions between PH and fusion domains but also distinct pH sensitive switches. Globally the comparison between the post fusion conformation of CHAV and VSV-G highlights several features essential for the protein’s function. It also reveals the remarkable plasticity of G in terms of local structures.
Highlights
Chandipura virus (CHAV; Family Rhabdoviridae, Genus Vesiculovirus) is an emerging human pathogen associated with deadly encephalitis, principally among children in the tropical areas of India
BSR cells were co-transfected with a plasmid encoding G and rabies virus (RABV) phosphoprotein fused to the green fluorescent protein (P-GFP)
Our results indicate that the fusion properties and the associated low-pH-induced conformational change are very similar for CHAV-G and VSV-G
Summary
Chandipura virus (CHAV; Family Rhabdoviridae, Genus Vesiculovirus) is an emerging human pathogen associated with deadly encephalitis, principally among children in the tropical areas of India. Most of the functional and structural information on CHAV has been inferred from studies performed on the prototype vesiculovirus, vesicular stomatitis virus serotype Indiana (VSVIND) This inference is reasonable due to the conservation of the amino acid (aa) sequences of the proteins of the two viruses [5]; for instance, aa sequences of glycoproteins G from both viruses share 40% identity and around 65% similarity [6]. At neutral pH, G exists as a population of mostly monomeric species with a minority of PRE trimers [12,15] whereas, at low pH, the G distribution is shifted toward the trimeric POST state [15,16] This is the main difference between rhabdoviral G and other viral fusion glycoproteins activated at low pH for which the PRE conformation is metastable [17,18]
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