First expression and characterization of a recombinant Cu-A-containing subunit II from an archaeal terminal oxidase complex

The branched respiratory chain of the archaeon Sulfolobus acidocaldarius co ntains a supercomplex, SoxM, consisting of a bc(1)-like subcomplex and a te rminal oxidase moiety, including a subunit II analogous polypeptide, SoxH. However, the latter component has never been identified in preparations of SoxM. We demonstrate the presence of an mRNA transcript by Northern analysi s. We succeeded in cloning and expressing the respective gene with truncate d N-terminus by deleting a 20 AS membrane anchor, which resulted in a water -soluble purple copper protein, which was further characterized. The recomb inant subunit II of the SoxM complex contains a correctly inserted binuclea r CUA cluster as revealed by UV/vis and EPR spectroscopy. The protein is hi ghly thermostable and displays a redox potential of +237 mV. In recombinant form, the metal interacts with cytochrome c as an artificial electron dono r; the physiological electron donor is still unknown, since S. acidocaldari us does not contain any c-type cytochromes. The purple copper center of Sox M shows an interesting pH dependency with a pK(a) at 6.4, suggesting proton ation of the Cu-ligating histidines. Further lowering the pH causes a rever sible transition into another cluster form with concomitant liberation of o ne copper. It may thus provide a model for the study of cluster rearrangeme nts in response to pH.