NMR ASSIGNMENTS AND RELAXATION STUDIES OF THIOBACILLUS-VERSUTUS FERROCYTOCHROME C-550 INDICATE THE PRESENCE OF A HIGHLY MOBILE 13-RESIDUES LONG C-TERMINAL TAIL

Cytochrome c-550 of Thiobacillus versutus functions as an electron tra nsfer protein in a chain of redox proteins that enables T. versutus to grow on methylamine. It is a single-heme protein of 134 residues, rel ated to mitochondrial cytochrome c. Cytochrome c-550, as well as sever al other bacterial c(2)-type cytochromes, contain a C-terminal extensi on of 13-16 amino acids of unknown function, compared to mitochondrial cytochrome c. NMR experiments were performed to obtain structural and dynamic information on the protein in solution. Far this purpose, T. versutus cytochrome c-550 was labeled with N-15 and C-13 using C-13-me thanol grown Paracoccus denitrificans as a host for heterologous expre ssion. NMR assignments were obtained for the H-1, N-15, and C-13 nucle i in the backbone and the beta-positions of the protein and the second ary structure was determined. N-15-relaxation studies were performed t o characterize the dynamic properties of the protein. The results indi cate that the main part of T. versutus ferrocytochrome c-550 exists in solution as a rigid, well-ordered molecule with a secondary structure that is very similar to that of P. denitrificans cytochrome c-550, as observed in crystals. The C-terminal extension, however, is unstructu red and highly mobile. The possible origin and function of the extensi on are discussed.