Characterization of a new dihemic c(4)-type cytochrome isolated from Thiobacillus ferrooxidans.
A new soluble c-type cytochrome has been purified to homogeneity from the acidophilic proteobacterium Thiobacillus ferrooxidans BRGM. It is characterized by an alpha-peak wavelength of 552 nm, a molecular mass of 26 567 Da (as determined by mass spectroscopy) and a pI value of 8. Optical redox titrations at pH 4.0 revealed the presence of two distinguishable redox species with an E(m) of 510 mV and an E(m) of 430 +/- 20 mV. EPR spectra recorded for this heme protein demonstrated the presence of stoichiometric amounts of two low-spin hemes with a g(z)() of 3.08 (510 mV species) and a g(z)() of 3.22 (430 mV species). Modifications of the physicochemical properties of the cytochrome were observed on complex formation with the blue copper protein rusticyanin, another soluble electron carrier in the genus Thiobacillus. N-Terminal sequencing yielded the polypeptide sequence up to the 50th residue. The determined sequence was found to be present (at 100% amino acid identity) in the (unfinished) genome of T. ferrooxidans ATCC 23270, and the corresponding full-length protein turned out to be surprisingly similar (34.5% amino acid identity) to another c(4)-type diheme protein from T. ferrooxidans BRGM [Cavazza, C., et al. (1996) Eur. J. Biochem. 242, 308-314], the gene of which is also present (at 97% amino acid identity) in the T. ferrooxidans ATCC 23270 genome. The physicochemical properties and sequence characteristics of both c(4) cytochromes present in the same bacteria are compared, and the functional role of this new diheme protein in the iron(II)-oxidizing electron transport chain in the genus Thiobacillus is discussed.
Giudici-Orticoni
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Laboratoire de Bioénergétique et Ingénierie des Protéines, CNRS, IFR1, 31, chemin Joseph Aiguier, 13402 Marseille Cedex 20, France. giudici@ibsm.cnrs-mrs.fr
Leroy
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Nitschke
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Bruschi
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001400:Azurin / Q000031:analogs & derivatives / Q000378:metabolism; D003574:Cytochrome c Group / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D013855:Thiobacillus / Q000737:chemistry*
D000596:Amino Acids; D003574:Cytochrome c Group; C012543:rusticyanin; D001400:Azurin; C034186:cytochrome C4