THE CYTOCHROME BD QUINOL OXIDASE IN ESCHERICHIA-COLI HAS AN EXTREMELYHIGH OXYGEN-AFFINITY AND 2 OXYGEN-BINDING HEMES - IMPLICATIONS FOR REGULATION OF ACTIVITY IN-VIVO BY OXYGEN INHIBITION
R. Dmello et al., THE CYTOCHROME BD QUINOL OXIDASE IN ESCHERICHIA-COLI HAS AN EXTREMELYHIGH OXYGEN-AFFINITY AND 2 OXYGEN-BINDING HEMES - IMPLICATIONS FOR REGULATION OF ACTIVITY IN-VIVO BY OXYGEN INHIBITION, Microbiology, 142, 1996, pp. 755-763
Cytochrome bd is a respiratory oxidase in Escherichia coli and many ot
her bacteria, It contains cytochromes b(558), b(595) and d as redox ce
ntres, and is thus unrelated to the haem-copper super-family of termin
al oxidases. The apparent affinities (K-m) for oxygen uptake by respir
ing cells and membranes from a mutant lacking the alternative oxidase
cytochrome bo' were determined by deoxygenation of oxyleghaemoglobin a
s a sensitive reporter of dissolved oxygen concentration. Respiration
rates were maximal at oxygen concentrations of 25-50 nM, but the kinet
ics were complex and indicative of substrate (i.e. oxygen) inhibition.
K-m values were in the range 3-8 nM (the lowest recorded for a respir
atory oxidase), and K-i values between 0.5 and 1.8 mu M were obtained.
Low temperature photodissociation of anoxic, CO-ligated membranes con
firmed the absence of cytochrome bo' and revealed a high-spin b-type c
ytochrome identified as cytochrome b(595) of the cytochrome bd complex
. Photodissociation in the presence of oxygen revealed binding of a li
gand (presumably oxygen) to cytochrome b(595) at a rate much greater t
han that of CO binding, and formation of the oxygenated form of cytoch
rome d. The results confirm that both high-spin haems in the cytochrom
e bd complex bind CO and demonstrate that oxygen can also react with b
oth haems. Substrate inhibition of oxidase activity, in addition to tr
anscriptional regulation of oxidase synthesis, may play a crucial role
in the regulation of partitioning of electron flux between the cytoch
rome bd- and bo'-terminated respiratory pathways.