Protein expression, crystallization and preliminary X-ray crystallographic analysis of the isolated Shigella flexneri VapC toxin.

Abstract

Upon release from the stable complex formed with its antitoxin VapB, the toxin VapC (MvpT) of the Gram-negative pathogen Shigella flexneri is capable of globally down-regulating translation by specifically cleaving initiator tRNA(fMet) in the anticodon region. Recombinant Shigella flexneri VapC(D7A) harbouring an active-site mutation was overexpressed in Escherichia coli, purified to homogeneity and crystallized by the vapour-diffusion technique. A preliminary X-ray crystallographic analysis shows that the crystals diffracted to at least 1.9 Å resolution at a synchrotron X-ray source and belonged to the trigonal space group in the hexagonal setting, H3, with unit-cell parameters a = b = 120.1, c = 52.5 Å, α = β = 90, γ = 120°. The Matthews coefficient is 2.46 Å(3) Da(-1), suggesting two molecules per asymmetric unit and corresponding to a solvent content of 50.0%.