SOLUTION STRUCTURE OF DESULFOVIBRIO-VULGARIS (HILDENBOROUGH) FERROCYTOCHROME C(3) - STRUCTURAL BASIS FOR FUNCTIONAL COOPERATIVITY

Desulfovibrio vulgaris cytochrome c(3), is a 14 kDa tetrahaem cytochro me that plays a central role in energy transduction. The three-dimensi onal structure of the ferrocytochrome at pH 8.5 was solved through two -dimensional H-1-NMR. The structures were calculated using a large amo unt of experimental information, which includes upper and lower distan ce limits as well as dihedral angle restraints. The analysis. allows f or fast-flipping aromatic residues and flexibility in the haem plane. The structure was determined using 2289 upper and 2390 lower distance limits, 63 restricted ranges for the phi torsion angle, 88 stereospeci fic assignments out of the 118 stereopairs with non-degenerate chemica l shifts (74.6%), and 115 out of the 184 nuclear Overhauser effects to fast-flipping aromatic residues (62.5%), which were pseudo-stereospec ifically assigned to one or the other side of the ring. The calculated NMR structures are very well defined, with an average root-mean-squar e deviation value relative to the mean coordinates of 0.35 Angstrom fo r the backbone atoms and 0.70 Angstrom for all heavy-atoms. Comparison of the NMR structures of the ferrocytochrome at PH 8.5 with the avail able X-ray structure of the ferricytochrome at pH 5.5 reveals that the general fold of the molecule is very similar, but that there are some distinct differences. Calculation of ring current shifts for the resi dues with significantly different conformations confirms that the NMR structures represent better its solution structure in the reduced form . Some of the localised differences, such as a reorientation of Thr24, are thought to be state-dependent changes that involve alterations in hydrogen bond networks. An important rearrange ment in the vicinity o f the propionate groups of haem I and involving the covalent linkage o f haem IT suggests that this is the critical region for the functional cooperativities of this protein. (C) 1998 Academic Press.