Purification and magneto-optical spectroscopic characterization of cytoplasmic membrane and outer membrane multiheme c-type cytochromes from Shewanella frigidimarina NCIMB400.
Two membranous c-type cytochromes from the Fe(III)-respiring bacterium Shewanella frigidimarina NCIMB400, CymA and OmcA, have been purified and characterized by UV-visible, magnetic circular dichroism, and electron paramagnetic resonance spectroscopies. The 20-kDa CymA is a member of the NapC/NirT family of multiheme cytochromes, which are invariably anchored to the cytoplasmic membrane of Gram-negative bacteria, and are postulated to mediate electron flow between quinols and periplasmic redox proteins. CymA was found to contain four low-spin c-hemes, each with bis-His axial ligation, and midpoint reduction potentials of +10, -108, -136, and -229 mV. The 85-kDa OmcA is located at the outer membrane of S. frigidimarina NCIMB400, and as such might function as a terminal reductase via interaction with insoluble Fe(III) substrates. This putative role is supported by the finding that the protein was released into solution upon incubation of harvested intact cells at 25 degrees C, suggesting an attachment to the exterior face of the outer membrane. OmcA was revealed by magneto-optical spectrocopies to contain 10 low-spin bis-His ligated c-hemes, with the redox titer indicating two sets of near iso-potential components centered at -243 and -324 mV.
Field
S J
SJ
School of Biological Sciences, Centre for Metalloprotein Biology and Spectroscopy, University of East Anglia, Norwich NR4 7TJ, United Kingdom.
Dobbin
P S
PS
Cheesman
M R
MR
Watmough
N J
NJ
Thomson
A J
AJ
Richardson
D J
DJ
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002451:Cell Compartmentation; D002458:Cell Fractionation; D002942:Circular Dichroism; D003574:Cytochrome c Group / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry; D008280:Magnetics; D008565:Membrane Proteins / Q000737:chemistry*; D010084:Oxidation-Reduction; D011199:Potentiometry; D020539:Sequence Analysis, Protein; D020592:Shewanella / Q000737:chemistry*; D013053:Spectrophotometry
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D003574:Cytochrome c Group; D008565:Membrane Proteins; C114590:omc A protein, Shewanella putrefaciens; D006418:Heme; C017165:cytochrome c(3)