The aerobic respiratory chain of the extremely acidophilic iron-oxidizing archaeon Ferroplasma acidiphilum

Respiratory supercomplexes have been extensively studied in mitochondria, but much less concerned in prokaryotes. Here we present the detailed characterization work on a supercomplex III/IV isolated from native membranes of eubacterium Aquifex aeolicus. This supercomplex III/IV survives fromdetergent solubilization and ismonodispersed in both ion-exchange and size-exclusion chromatography. It consists of cytochrome b, cytochrome c1 and Rieske protein from the cytochrome bc1 complex and subunits I, II and IIa from the cytochrome oxidase. Major subunits are clearly visible on SDS-PAGE and later on identified by MALDI-MS. Activity assay showed that this supercomplex has higher turnover numbers than the purified cytochrome oxidase, which was called “Cox2” in our former work [1]. The stoichiometry of the supercomplex has been characterized as a homo-dimeric bc1 complex and a monomeric oxidase based on heme ratios, protein amounts and metal contents. Functional analysis on the supercomplex III2IV indicates potential interaction between the cytochrome bc1 complex and the cytochrome oxidase. In order to investigate the potential interaction, crosslinking work of the supercomplex has been performed, antibodies against protein subunits have been produced, and mass spectrometry has been applied. Further functional analysis of the supercomplex would be done as soon as possible aiming to get a picture of the supercomplex functional organization.