FURTHER CHARACTERIZATION OF THE 2 TETRAHEME CYTOCHROMES C(3) FROM DESULFOVIBRIO-AFRICANUS - NUCLEOTIDE-SEQUENCES, EPR SPECTROSCOPY AND BIOLOGICAL-ACTIVITY

The genes encoding the basic and acidic tetraheme cytochromes c(3) fro m Desulfovibrio africanus have been sequenced. The corresponding amino acid sequences of the basic and acidic cytochromes c(3) indicate that the mature proteins consist of a single polypeptide chain of 117 and 103 residues, respectively. Their molecular masses, 15102 and 13742 Da , respectively, determined by mass spectrometry, are in perfect agreem ent with those calculated from their amino acid sequences. Both D. afr icanus cytochromes c(3) are synthesized as precursor proteins with sig nal peptides of 23 and 24 residues for the basic and acidic cytochrome s, respectively. These cytochromes c(3) exhibit the main structural fe atures of the cytochrome c(3) family and contain the 16 strictly conse rved cysteine + histidine residues directly involved in the heme bindi ng sites. The D. africanus acidic cytochrome c(3) differs from all the other homologous cytochromes by its low content of basic residues and its distribution of charged residues in the amino acid sequence. The presence of four hemes per molecule was confirmed by EPR spectroscopy in both cytochromes c(3). The g-value analysis suggests that in both c ytochromes, the angle between imidazole planes of the axial histidine ligands is close to 90 degrees for one heme and much lower for the thr ee others. Moreover, an unusually high exchange interaction (similar t o 10(-2) cm(-1)) was evidenced between the highest potential heme (-90 mV) and one of the low potential hemes in the basic cytochrome c(3). The reactivity of D. africanus cytochromes c(3) with heterologous [NiF e] and [Fe] hydrogenases was investigated. Only the basic one interact s with the two types of hydrogenase to achieve efficient electron tran sfer, whereas the acidic cytochrome c(3) exchanges electrons specifica lly with the basic cytochrome c(3). The difference in the specificity of the two D. africanus cytochromes c(3) has been correlated with thei r highly different content of basic and acidic residues. (C) 1997 Else vier Science B.V.