NMR OF CHROMATIUM-VINOSUM FERREDOXIN - EVIDENCE FOR STRUCTURAL INEQUIVALENCE AND IMPEDED ELECTRON-TRANSFER BETWEEN THE 2 [4FE-4S] CLUSTERS

The 2[4Fe-4S] ferredoxin from Chromatium vinosum has been investigated by H-1 and C-13 nuclear magnetic resonance. H-1 NMR sequence-specific assignments have been obtained for a large majority of the residues. They indicate that the protein folds along a pattern similar to that p reviously evidenced for shorter 2[4Fe-4S] ferredoxins. However, C. vin osum ferredoxin differs from other ferredoxins by the occurrence of a turn in an eight amino acid region separating two successive cysteines , Cys-40 and Cys-49, liganding one cluster. Also, the unique C-termina l end of C. vinosum ferredoxin contains a 10 amino acid alpha-helix wh ich interacts with one side of the above turn. The only cysteine of th e sequence not involved in the ligation of the [4Fe-4S] clusters is Cy s-57. Specific NMR experiments helped characterizing the signals arisi ng from the ligands of these clusters: most of them display properties reminiscent of those of homologous ferredoxins, except for the signal s associated with Cys-40. Despite the general similarity between C, vi nosum ferredoxin and other 2[4Fe-4S] ferredoxins, the electron paramag netic resonance and NMR spectra of the former reduced protein are sign ificantly different from those previously observed for S = 1/2 [4Fe-4S ](+) clusters. In addition, the intramolecular electron transfer rate in C. vinosum is far slower than in other similar cases. This is the f irst report of impeded electron exchange between two [4Fe-4S] clusters expected to be less than 12 Angstrom apart.