The Role of the Supernumerary Subunit of Rhodobactersphaeroides Cytochrome bc1 Complex

The smallest molecular weight subunit (subunit IV), which contains no redox prosthetic group,is the only supernumerary subunit in the four-subunit Rhodobacter sphaeroides bc^sub 1^ complex.This subunit is involved in Q binding and the structural integrity of the complex. When thecytochrome bc^sub 1^ complex is photoaffinity labeled with [^sup 3^H]azido-Q derivative, radioactivity isfound in subunits IV and I (cytochrome b), indicating that these two subunits are responsiblefor Q binding in the complex. When the subunit IV gene (fbcQ) is deleted from the R.sphaeroides chromosome, the resulting strain (RSΔIV) requires a period of adaptation beforethe start of photosynthetic growth. The cytochrome bc^sub 1^ complex in adapted RSΔIVchromatophores is labile to detergent treatment (60-75% inactivation), and shows a four-fold increasein the K^sub m^ for Q^sub 2^H^sub 2^. The first two changes indicate a structural role of subunit IV; the thirdchange supports its Q-binding function. Tryptophan-79 is important for structural andQ-binding functions of subunit IV. Subunit IV is overexpressed in Escherichia coli as a GSTfusion protein using the constructed expression vector, pGEX/IV. Purified recombinant subunitIV is functionally active as it can restore the bc^sub 1^ complex activity from the three-subunit corecomplex to the same level as that of wild-type or complement complex. Three regions in thesubunit IV sequence, residues 86-109, 77-85, and 41-55, are essential for interaction withthe core complex because deleting one of these regions yields a subunit completely or partiallyunable to restore cytochrome bc^sub 1^ from the core complex.[PUBLICATION ABSTRACT]