Stability of the cbb ₃ -Type Cytochrome Oxidase Requires Specific CcoQ-CcoP Interactions

Abstract

Cytochrome cbb 3 -type oxidases are members of the heme copper oxidase superfamily and are composed of four subunits. CcoN contains the heme b-Cu B binuclear center where oxygen is reduced, while CcoP and CcoO are membrane-bound c-type cytochromes thought to channel electrons from the donor cytochrome into the binuclear center. Like many other bacterial members of this superfamily, the cytochrome cbb 3 -type oxidase contains a fourth, non-cofactor-containing subunit, which is termed CcoQ. In the present study, we analyzed the role of CcoQ on the stability and activity of Rhodobacter capsulatus cbb 3 -type oxidase. Our data showed that CcoQ is a single-spanning membrane protein with a N out -C in topology. In the absence of CcoQ, cbb 3 -type oxidase activity is significantly reduced, irrespective of the growth conditions. Blue native polyacrylamide gel electrophoresis analyses revealed that the lack of CcoQ specifically impaired the stable recruitment of CcoP into the cbb 3 -type oxidase complex. This suggested a specific CcoQ-CcoP interaction, which was confirmed by chemical cross-linking. Collectively, our data demonstrated that in R. capsulatus CcoQ was required for optimal cbb 3 -type oxidase activity because it stabilized the interaction of CcoP with the CcoNO core complex, leading subsequently to the formation of the active 230-kDa cbb 3 -type oxidase complex.

A Critical Role for the cccA Gene Product, Cytochrome c 2 , in Diverting Electrons from Aerobic Respiration to Denitrification in Neisseria gonorrhoeae

“…The fourth subunit of cytochrome oxidase cbb 3 , CcoQ, has been suggested to be an assembly factor required for the formation of a fully active cytochrome oxidase complex (38,39). However, unlike most other bacteria, the gonococcal cytochrome oxidase also transfers electrons to nitrite, and the nitrite and oxygen reduction functions are essentially independent (compare data in reference 24 with those in Fig.…”

Expression of multiplecbb3cytochromecoxidase isoforms by combinations of multiple isosubunits inPseudomonas aeruginosa

“…CcoO and CcoP are transmembrane monoheme and diheme cytochromes c, respectively (5). CcoQ is known to affect the stability of the cbb 3 complex, but it is not necessarily a component of purified cbb 3 (6)(7)(8).…”

Biochemical and Biophysical Characterization of the Two Isoforms of cbb3-Type Cytochrome c Oxidase from Pseudomonas stutzeri

“…In Bradyrhizobium japonicum and R. sphaeroides, it was shown that deletion of CcoQ had no effect on assembly or catalytic activity of cbb 3 -CcO (22,56). In contrast, the activity of R. capsulatus cbb 3 -CcO was significantly reduced in the absence of CcoQ (23). Additionally, it has been demonstrated that CcoQ is required to protect the R. sphaeroides cbb 3 -CcO from degradation under aerobic conditions (24).…”

“…It has also been suggested to serve as a gas-sensing element because one reduced heme c in this subunit can bind carbon monoxide (8). CcoQ is the smallest subunit present in some cbb 3 -CcOs, which was shown to be involved in the stabilization of the cbb 3 -CcO by interaction with subunit CcoP in Rhodobacter capsulatus (23) and by protection of the cbb 3 -CcO from oxidative destabilization in Rhodobacter sphaeroides (24).…”