The haem b558 component of the cytochrome bd quinol oxidase complex from Escherichia coli has histidine-methionine axial ligation

• Published in: Biochemical journal 1995-06, Vol.308 ( Pt 2) (Pt 2), p.641-644
• Main Authors: Spinner, F, Cheesman, M R, Thomson, A J, Kaysser, T, Gennis, R B, Peng, Q, Peterson, J
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - chemistry; Circular Dichroism; Cytochrome b Group - chemistry; Cytochromes - chemistry; Electron Spin Resonance Spectroscopy; Electron Transport Chain Complex Proteins; Escherichia coli - enzymology; Escherichia coli Proteins; Heme - chemistry; Histidine - chemistry; Methionine - chemistry; NADPH Oxidases; Oxidoreductases - chemistry; Spectrophotometry, Ultraviolet
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj3080641

The cytochrome bd ubiquinol oxidase from Escherichia coli is induced when the bacteria are cultured under microaerophilic or low-aeration conditions. This membrane-bound respiratory oxidase catalyses the two-electron oxidation of ubiquinol and the four-electron reduction of dioxygen to water. The oxidase contains three haem prosthetic groups: haem b558, haem b595 and haem d. Haem d is the oxygen binding site, and it is likely that haem d and b595 form a bimetallic site in the enzyme. Haem b558 has been previously characterized spectroscopically as being low spin and has been shown to be located within subunit I (CydA) of this two-subunit enzyme. It is likely that haem b558 is associated with the quinol oxidation site, which has also been shown to be within subunit I. In a previous effort to locate the specific amino acids axially ligated to haem b558, all six histidines within subunit I were altered by site-directed mutagenesis. Only one, histidine-186, was identified as a likely ligand to haem b558. Hence it was suggested that haem b558 could not have bis(histidine) ligation. In the current work, a combination of low-temperature near-infrared magnetic circular dichroism (NIR-MCD) and EPR spectroscopies have been employed to identify the nature of the haem b558 axial ligands. The NIR-MCD spectrum at cryogenic temperatures is dominated by the low-spin haem b558 component of the complex, and the low-energy band near 1800 nm is strong evidence for histidine-methionine ligation. It is concluded that haem b558 is ligated to histidine-186 plus one of the methionines located within subunit I of the oxidase.