Edited by Miguel De la Rosa Keywords:Ubiquinol cytochrome bd oxidase CydAB CydX UV/vis difference spectroscopy Escherichia coli a b s t r a c t Cytochrome bd ubiquinol oxidase uses the electron transport from ubiquinol to oxygen to establish a proton gradient across the membrane. The enzyme complex consists of subunits CydA and B and contains two b-and one d-type hemes as cofactors. Recently, it was proposed that a third subunit named CydX is essential for the function of the complex. Here, we show that CydX is indeed a subunit of purified Escherichia coli cytochrome bd oxidase and that the small protein is needed either for the assembly or the stability of the active site di-heme center and, thus, is essential for oxidase activity. Structured summary of protein interactions:cydA physically interacts with cydB by affinity technology (View interaction) cydA physically interacts with cydB by molecular sieving (View interaction) cydB, cydA and cydX physically interact by molecular sieving (View interaction) cydB, cydA, and cydX physically interacts by affinity technology (1, 2)

cydB , cydA , and cydX physically interacts by affinity technology (1, 2)