Nine-haem cytochrome c from Desulfovibrio desulfuricans ATCC 27774: primary sequence determination, crystallographic refinement at 1.8 and modelling studies of its interaction with the tetrahaem cytochrome c(3)

A monomeric nine-haem cytochrome c (9Hcc) with 292 amino acid residues was isolated from cells of the sulfate- and nitrate-reducing bacterium Desulfov ibrio desulfuricans ATCC 27774 grown under both nitrate- and sulfate-respir ing conditions. The nucleotide sequence encoding the 292 residues was deter mined, allowing the correction of about 10% of the previous primary structu re, determined from 1.8 Angstrom electron density maps. The refinement at 1 .8 Angstrom resolution of the structural model was completed, giving an R-v alue of 16.5%. The nine haem groups are arranged into two tetrahaem cluster s, located at both ends of the molecule, with Fe-Fe distances and local pro tein fold very similar to tetrahaem cytochromes c(3) and the extra haem is located asymmetrically between the two regions. The new primary sequence de termination confirmed the 39% sequence homology found between this cytochro me and the C-terminal region (residues 229-514) of the high-molecular-weigh t cytochrome c (Hmc) from D. vulgaris Hildenborough, providing strong evide nce of structural similarity between 9Hcc and the C-terminal region of Hmc. The interaction between 9Hcc and the tetrahaem cytochrome c3 from the same organism was studied by modelling methods, and the results suggest that a specific interaction is possible between haem 4 of tetrahaem cytochrome c3 and haem 1 or haem 2 of 9Hcc, in agreement with previous kinetic experiment s which showed the catalytic effect of the tetrahaem cytochrome c3 upon the reduction of 9Hcc by the [NiFe] hydrogenase from D. desulfuricans ATCC 277 74. These studies suggest a role for 9Hcc as part of the assembly of redox proteins involved in recycling the molecular hydrogen released by the cell as a result of substrate oxidation.