Crystallization, preliminary X-ray analysis of a native and selenomethionine D-hydantoinase from Thermus sp.

ABSTRACT

A D-hydantoinase from Thermus sp. was expressed in Escherichia coli, purified to homogeneity and crystallized both as native and Se-Met labelled protein. The crystals belong to the orthorhombic space group C222(1), with unit-cell parameters a = 125.9, b = 215.8, c = 207.5 A. A three-wavelength MAD data set was collected to 2.5 A resolution and a native data set was collected to 1.7 A resolution. Crystal packing and self-rotation calculations led to the assumption of six protomers per asymmetric unit, corresponding to a V(M) value of 2.28 A(3) Da(-1) and a solvent content of 46%. As each protomer contains nine Se-Met residues, 54 selenium sites per asymmetric unit were present and could be unambigously located in the course of the MAD experiment. This selenium substructure is one of the largest selenium substructures that have been solved to date. The resulting phases obtained at a high-resolution limit of 3.0 A could be extended to 1.7 A and refined by application of density-modification techniques, especially non-crystallographic symmetry.