The DsrMKJOP transmembrane complex has a most important function in dissimilatory sulfur metabolism and consists of cytoplasmic, periplasmic and membrane integral proteins carrying FeS centers and b- and c-type cytochromes as cofactors. In the present study the DsrMKJOP transmembrane complex was investigated and characterized by biochemical, biophysical and functional analyses. Each of the proteins was produced in E. coli as recombinant proteins and purified. DsrJ is an unusual triheme cytochrome and EPR spectroscopy provided evidence for a possible, but only partial, His/Cys heme ligation in one of the hemes. DsrM was identified as a diheme cytochrome b and the two hemes were found to be in low spin state. Although no hemes were predicted for DsrP, it was also clearly identified as a b-type cytochrome. DsrO and DsrK were both experimentally proven to be Fe-S-containing proteins and DsrK was shown to be membrane-associated. Coelution assays provide support for the proposed interaction of DsrK with DsrC, which might be its substrate. The results obtained in this study are combined in a model for the function of the DsrMKJOP complex in A. vinosum.