Expression, purification and crystallization of VP4 protease from Tellina virus 1.

Abstract

Tellina virus 1 is an aquabirnavirus that was isolated from the sand-dwelling marine bivalve mollusc Tellina tenuis. The self-encoded protease viral protein 4 (VP4) processes its own polyprotein to yield the individual proteins VP2 and VP3 that are required for viral assembly. VP4 protease utilizes a serine-lysine catalytic dyad in its mechanism. A full-length VP4 construct was overexpressed in Escherichia coli and purified to homogeneity using nickel-affinity chromatography. Ion-exchange and size-exclusion chromatographic steps were utilized to isolate a monomeric fraction of the protein. The purified monomeric VP4 was subjected to limited proteolysis to yield crystallizable protein. Crystal growth was performed using the hanging-drop vapour-diffusion method and was carried out at room temperature (∼296 K). Hexagonal crystals grew in the presence of PEG 8000, ammonium sulfate and urea. These crystals diffracted to beyond 2.1 Å resolution and belonged to space group P6(4)22, with unit-cell parameters a=59.1, b=59.1, c=208.1 Å, one molecule in the asymmetric unit and a solvent content of 42%.