Interaction of cytochrome c with cytochrome c oxidase: An NMR study on two soluble fragments derived from Paracoccus denitrificans

The functional interactions between the various components of the respiratory chain are relatively short-lived, thus allowing high turnover numbers but at the same time complicating the structural analysis of the complexes. Chemical shift mapping by NMR spectroscopy is a useful tool to investigate such transient contacts, since it can monitor changes in the electron-shielding properties of a protein as the result of temporary contacts with a reaction partner. In this study, we investigated the molecular interaction between two components of the electron-transfer chain from Paracoccus denitrificans: the engineered, water-soluble fragment of cytochrome c552and the CUAdomain from the cytochrome c oxidase. Comparison of [15N,1H]-TROSY spectra of the [15N]-labeled cytochrome c552fragment in the absence and in the presence of the CUAfragment showed chemical shift changes for the backbone amide groups of several, mostly uncharged residues located around the exposed heme edge in cytochrome c552. The detected contact areas on the cytochrome c552surface were comparable under both fully reduced and fully oxidized conditions, suggesting that the respective chemical shift changes represent biologically relevant protein-protein interactions.