MOLECULAR-STRUCTURE OF THE OXIDIZED HIGH-POTENTIAL IRON-SULFUR PROTEIN ISOLATED FROM ECTOTHIORHODOSPIRA-VACUOLATA

The high-potential iron-sulfur protein (iso-form II) isolated from Ect othiorhodospira vacuolata has been crystallized and its three-dimensio nal structure determined by molecular replacement procedures and refin ed to 1.8-Angstrom resolution with a crystallographic R factor of 16.3 %. Crystals employed in the investigation belonged to the space group C222(1) with unit cell dimensions of alpha = 58.4 Angstrom, b 64.7 Ang strom and 39.3 Angstrom and one molecule per asymmetric unit. Like tho se HiPIPs structurally characterized thus far, the E. vacuolata molecu le contains mostly reverse turns that wrap around the iron-sulfur clus ter with cysteine residues 34, 37, 51, and 65 ligating the metal cente r to the polypeptide chain. There are 57 ordered solvent molecules, mo st of which lie at the surface of the protein. Two of these water mole cules play important structural roles by stabilizing the loops located between Asp 42 and Lys 57. The metal center binding pocket is decided ly hydrophobic with the closest solvent molecule being 6.9 Angstrom fr om S2 of the [4Fe-4S] cluster. The E. vacuolata HiPIP molecules pack i n the crystalline lattice as dimers with their iron-sulfur centers app roximately 17.5 Angstrom apart. On the basis of biochemical properties , it was anticipated that the E. vacuolata HiPIP would be structurally more similar to the HiPIP isolated from Ectothiorhodospira halophila than to the protein obtained from Chromatium vinosum. In fact, the E. vacuolata molecule is as structurally close to the C. vinosum HiPIP as it is to the E. halophila protein due to the presence of various inse rtions and deletions that disrupt local folding. The E. vacuolata HiPI P structure thus calls into question whether molecular modeling experi ments, based on primary structure homology alone, are valid when numer ous insertions and deletions are present.