Effect of phosphate on bacterioferritin-catalysed iron(II) oxidation.
The iron(III) mineral cores of bacterioferritins (BFRs), as isolated, contain a significant component of phosphate, with an iron-to-phosphate ratio approaching 1:1 in some cases. In order to better understand the in vivo core-formation process, the effect of phosphate on in vitro core formation in Escherichia coli BFR was investigated. Iron cores reconstituted in the presence of phosphate were found to have iron-to-phosphate ratios similar to those of native cores, and possessed electron paramagnetic resonance properties characteristic of the phosphate-rich core. Phosphate did not affect the stoichiometry of the initial iron(II) oxidation reaction that takes place at the intrasubunit dinuclear iron-binding sites (phase 2 of core formation), but did increase the rate of oxidation. Phosphate had a more significant effect on subsequent core formation (the phase 3 reaction), increasing the rate up to five-fold at pH 6.5 and 25 degrees C. The dependence of the phase 3 rate on phosphate was complex, being greatest at low phosphate and gradually decreasing until the point of saturation at approximately 2 mM phosphate (for iron(II) concentrations <200 microM). Phosphate caused a significant decrease in the absorption properties of both phase 2 and phase 3 products, and the phosphate dependence of the latter mirrored the observed rate dependence, suggesting that distinct iron(III)-phosphate species are formed at different phosphate concentrations. The effect of phosphate on absorption properties enabled the observation of previously undetected events in the phase 2 to phase 3 transition period.
Aitken-Rogers
Helen
H
Centre for Metalloprotein Spectroscopy and Biology, School of Chemical Sciences and Pharmacy, University of East Anglia, Norwich NR4 7TJ, UK.
Singleton
Chloe
C
Lewin
Allison
A
Taylor-Gee
Alice
A
Moore
Geoffrey R
GR
Le Brun
Nick E
NE
D001426:Bacterial Proteins / Q000737:chemistry*; D002384:Catalysis; D002852:Chromatography, Ion Exchange; D003573:Cytochrome b Group / Q000737:chemistry*; D005293:Ferritins / Q000737:chemistry*; D005296:Ferrous Compounds / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D010084:Oxidation-Reduction; D010710:Phosphates / Q000737:chemistry*; D011487:Protein Conformation
D001426:Bacterial Proteins; D003573:Cytochrome b Group; D005296:Ferrous Compounds; D010710:Phosphates; D005293:Ferritins; C018032:bacterioferritin