Abstract
Cytochrome c maturation (Ccm) is a post-translational process that occurs after translocation of apocytochromes c to the positive (p) side of energy-transducing membranes. Ccm is responsible for the formation of covalent bonds between the thiol groups of two cysteines residues at the heme-binding sites of the apocytochromes and the vinyl groups of heme b (protoporphyrin IX-Fe). Among the proteins (CcmABCDEFGHI and CcdA) required for this process, CcmABCD are involved in loading heme b to apoCcmE. The holoCcmE thus formed provides heme b to the apocytochromes. Catalysis of the thioether bonds between the apocytochromes c and heme b is mediated by the heme ligation core complex, which in Rhodobacter capsulatus contains at least the CcmF, CcmH, and CcmI components. In this work we show that the heme chaperone apoCcmE binds to the apocytochrome c and the apocytochrome c chaperone CcmI to yield stable binary and ternary complexes in the absence of heme in vitro. We found that during these protein-protein interactions, apoCcmE favors the presence of a disulfide bond at the apocytochrome c heme-binding site. We also establish using detergent-dispersed membranes that apoCcmE interacts directly with CcmI and CcmH of the heme ligation core complex CcmFHI. Implications of these findings are discussed with respect to heme transfer from CcmE to the apocytochromes c during heme ligation assisted by the core complex CcmFHI.