A novel haem compound accumulated in Escherichia coli overexpressing the cydDC operon, encoding an ABC-type transporter required for cytochrome assembly.
cydDC genes encode a heterodimeric ABC transporter required for assembly of the membrane-bound cytochrome bd quinol oxidase and periplasmic cytochromes. Here, we demonstrate that overexpression of functional cydDC genes on a multicopy plasmid results in elevated levels of cytochromes b and d, but most notably formation in anaerobically grown cells of a novel haem-containing component P-574. The pigment has a distinctive absorbance at 574-579 nm and 448 nm in reduced minus oxidised spectra and renders over-producing cells reddish in colour. The highest levels of P-574 were observed in mutants (cydAB) in the structural genes for the polypeptides of cytochrome bd. P-574 is labile; its spectral signal is reduced in cells that are frozen-thawed or subjected to mechanical disruption. P-574 was not detected in cytoplasmic or periplasmic fractions and was predominantly associated with the cell membrane. P-574 did not bind CO or cyanide. Production of P-574 was dependent on haem biosynthesis indicating that it is a haem-containing molecule or derived from haem biosynthesis. These findings suggest that P-574 may result from association of a haem compound with overexpressed transporter subunits, but not with oxidase subunits, and are consistent with an intimate link between the transporter and haem processing during oxidase assembly.
Cook
Gregory M
GM
Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, The University of Sheffield, UK.
Cruz-Ramos
Hugo
H
Moir
Arthur J G
AJ
Poole
Robert K
RK
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000235:genetics; D003580:Cytochromes / Q000096:biosynthesis*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006418:Heme / Q000096:biosynthesis / Q000378:metabolism*; D009876:Operon; D010088:Oxidoreductases / Q000096:biosynthesis; D010957:Plasmids / Q000235:genetics
D018528:ATP-Binding Cassette Transporters; D003580:Cytochromes; D006418:Heme; D010088:Oxidoreductases