Structural and functional analysis of aa3-type and cbb3-type cytochrome c oxidases of Paracoccus denitrificans reveals significant differences in proton-pump design.
In Paracoccus denitrificans the aa3-type cytochrome c oxidase and the bb3-type quinol oxidase have previously been characterized in detail, both biochemically and genetically. Here we report on the isolation of a genomic locus that harbours the gene cluster ccoNOOP, and demonstrate that it encodes an alternative cbb3-type cytochrome c oxidase. This oxidase has previously been shown to be specifically induced at low oxygen tensions, suggesting that its expression is controlled by an oxygen-sensing mechanism. This view is corroborated by the observation that the ccoNOOP gene cluster is preceded by a gene that encodes an FNR homologue and that its promoter region contains an FNR-binding motif. Biochemical and physiological analyses of a set of oxidase mutants revealed that, at least under the conditions tested, cytochromes aa3, bb3 and cbb3 make up the complete set of terminal oxidases in P. denitrificans. Proton-translocation measurements of these oxidase mutants indicate that all three oxidase types have the capacity to pump protons. Previously, however, we have reported decreased H+/e- coupling efficiencies of the cbb3-type oxidase under certain conditions. Sequence alignment suggests that many residues that have been proposed to constitute the chemical and pumped proton channels in cytochrome aa3 (and probably also in cytochrome bb3) are not conserved in cytochrome cbb3. It is concluded that the design of the proton pump in cytochrome cbb3 differs significantly from that in the other oxidase types.
de Gier
J W
JW
Department of Molecular and Cellular Biology, BioCentrum Amsterdam, Vrije Universiteit, Netherlands.
Schepper
M
M
Reijnders
W N
WN
van Dyck
S J
SJ
Slotboom
D J
DJ
Warne
A
A
Saraste
M
M
Krab
K
K
Finel
M
M
Stouthamer
A H
AH
van Spanning
R J
RJ
van der Oost
J
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010101:Oxygen Consumption; D010231:Paracoccus denitrificans / Q000201:enzymology*; D017494:Proton Pumps; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D017494:Proton Pumps; D010088:Oxidoreductases; C020094:duroquinol oxidase; D003576:Electron Transport Complex IV