Ab initio solution and refinement of two high-potential iron protein structures at atomic resolution

The crystal structure of the reduced high-potential iron protein (HiPIP) fr om Chromatium vinosum has been redetermined in a new orthorhombic crystal m odification, and the structure-of its H42Q mutant has been determined in or thorhombic (H42Q-1) and cubic (H42Q-2) modifications. The first two were so lved by ab initio direct methods using data collected to atomic resolution (1.20 and 0.93 Angstrom, respectively). The recombinant wild type (rc-WT) w ith two HiPIP molecules in the asymmetric unit has 1264 protein atoms and 3 35 solvent sites, and is the second largest structure reported so far that has been solved by pure direct methods. The solutions were obtained in a fu lly automated way and included more than 80% of the protein atoms. Restrain ed anisotropic refinement for re-WT and H42Q-1 converged to R-1 = Sigma \\F -0\ - \ F-c\/Sigma \ F-0\ of 12.0 and 13.6%, respectively [data with I > 2 sigma(I)], and 12.8 and 15.5% tall data). H42Q-2 contains two molecules in the asymmetric unit and diffracted only to 2.6 Angstrom. In both molecules of rc-WT and in the single unique molecule of H42Q-1 the [Fe4S4](2+) cluste r dimensions are very similar and show a characteristic tetragonal distorti on with four short Fe-S bonds along four approximately parallel cube edges, and eight long Fe-S bonds. The unique protein molecules in H42Q-2 and re-W T are also very similar in other respects, except for the hydrogen bonding around the mutated residue that is at the surface of the protein, supportin g the hypothesis that the difference in redox potentials at lower pH values is caused primarily by differences in the charge distribution near the sur face of the protein rather than by structural differences in the cluster re gion.