Kyasanur Forest disease virus non-structural protein NS1 forms multimers in solution, with a distinctly identifiable tetrameric state

Kyasanur Forest Disease Virus (KFDV), a flavivirus, is predominantly present in the tropical region of southern India and is responsible for viral hemorrhagic disease in primates and non-primate animals. KFDV infection is spread by tick bites. The other medically important viruses of Flaviviridae family are dengue (DENV), Zika (ZIKV), West Nile virus (WNV) and Japanese encephalitis virus (JEV). The flaviviruses are collectively responsible for diverse disease pathologies and account for a major global health burden. A major contributing factor to disease pathogenesis of flavivirus is the secreted form of non-structural protein 1 (NS1). However, in vivo studies using lethal flavivirus challenge have demonstrated the protective role of NS1-specific antibodies and complement the hypothesis to explore possibilities of NS1-based vaccine and therapeutic candidates. Recent structural studies on DENV, ZIKV, JEV and WNV NS1 antigen have shown that the sNS1 protein exists in high-order oligomeric states. However, structural insights about the high-order oligomeric states of sNS1 of tick-borne flaviviruses and their biological significance are poorly explored. In this study, we have expressed and purified the KFDV NS1 protein in the mammalian expression system. The KFDV sNS1 protein exhibits higher oligomeric conformation in solution as determined by size exclusion chromatography (SEC), and negative stain transmission electron microscopy (NS-TEM). Single-particle analysis of KFDV NS1 reveals tetrameric arrangements that are structurally similar to previously reported NS1 structures from other flaviviruses. Our study will help to develop a future roadmap of the rational design of broad-spectrum anti-NS1 antibodies and subunit vaccines effective against tick-borne flaviviruses.