Crystallization and preliminary X-ray analysis of the small subunit (R2F) of native ribonucleotide reductase from Corynebacterium ammoniagenes

The crystallization of the metallo-cofactor (R2F) of native ribonucleotide reductase isolated from the Mn-requiring Gram-positive bacterium C. ammoniagenes is described. The crystals diffracted to 1.36 Å resolution. Ribonucleotide reduction, the unique step in DNA-precursor biosynthesis, involves radical-dependent redox chemistry and diverse metallo-cofactors. The metallo-cofactor (R2F) encoded by the nrdF (nucleotide reduction) gene in Corynebacterium ammoniagenes ATCC 6872 was isolated after homologous expression and a new crystal form of ribonucleotide reductase R2F was obtained. R2F was crystallized at 277 K using the vapour-diffusion method with PEG as the precipitating agent. A data set was collected to 1.36 Å resolution from a single crystal at 100 K using synchrotron radiation. The crystal belonged to space group C2, with unit-cell parameters a = 96.21, b = 87.68, c = 83.25 Å, β = 99.29°. The crystal contained two molecules per asymmetric unit, with a Matthews coefficient (V_M) of 2.69 ų Da⁻¹; the solvent content was estimated to be 54.3%. X-ray fluorescence spectroscopy and MAD diffraction data indicated the presence of manganese in the molecule and the absence of iron