Identification and enzymatic characterization of NS2B-NS3 protease of Alkhurma virus, a class-4 flavivirus

We report here on the identification of the ALKV NS2B and NS3 domains and the expression and purification of a catalytically active viral protease as a hexahistidine recombinant protein. Its enzymatic properties were characterized in vitro using a para-nitroanilide substrate. This constitutes the first characterization of the proteinase from a class-4 flavivirus. Our results indicate that the association of NS3 with a short segment of NS2B is necessary and sufficient for protease activity. The developed system could help to identify or design inhibitors potentially active as antiviral drugs against ALKV and other pathogenic flaviviruses.