Datensatz

Zusammenfassung

Cytochrome c oxidases (CcOs), members of the heme-copper containing oxidase (HCO) superfamily, are the terminal enzymes of aerobic respiratory chains. The cbb₃-type cytochrome c oxidases (cbb₃-CcO) form the C-family and have only the central catalytic subunit in common with the A- and B-family HCOs. In Pseudomonas stutzeri, two cbb₃ operons are organized in a tandem repeat. The atomic structure of the first cbb₃ isoform (Cbb₃-1) was determined at 3.2 Å resolution in 2010 (S. Buschmann, E. Warkentin, H. Xie, J. D. Langer, U. Ermler, and H. Michel, Science 329:327–330, 2010, http://dx.doi.org/10.1126/science.1187303). Unexpectedly, the electron density map of Cbb₃-1 revealed the presence of an additional transmembrane helix (TMH) which could not be assigned to any known protein. We now identified this TMH as the previously uncharacterized protein PstZoBell_05036, using a customized matrix-assisted laser desorption ionization (MALDI)–tandem mass spectrometry setup. The amino acid sequence matches the electron density of the unassigned TMH. Consequently, the protein was renamed CcoM. In order to identify the function of this new subunit in the cbb₃ complex, we generated and analyzed a CcoM knockout strain. The results of the biochemical and biophysical characterization indicate that CcoM may be involved in CcO complex assembly or stabilization. In addition, we found that CcoM plays a role in anaerobic respiration, as the ΔCcoM strain displayed altered growth rates under anaerobic denitrifying conditions