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Spectroscopic and Redox Studies of Valence-Delocalized [Fe2S2]+ Centers in Thioredoxin-like Ferredoxins
journal contribution
posted on 2015-04-08, 00:00 authored by Sowmya Subramanian, Evert C. Duin, Sarah E. J. Fawcett, Fraser
A. Armstrong, Jacques Meyer, Michael K. JohnsonReduced
forms of the C56S and C60S variants of the thioredoxin-like Clostridium pasteurianum [Fe2S2] ferredoxin
(CpFd) provide the only known examples of valence-delocalized
[Fe2S2]+ clusters, which constitute
a fundamental building block of all higher nuclearity Fe–S
clusters. In this work, we have revisited earlier work on the CpFd variants and carried out redox and spectroscopic studies
on the [Fe2S2]2+,+ centers in wild-type
and equivalent variants of the highly homologous and structurally
characterized Aquifex aeolicus ferredoxin 4 (AaeFd4) using EPR, UV–visible–NIR absorption,
CD and variable-temperature MCD, and protein–film electrochemistry.
The results indicate that the [Fe2S2]+ centers in the equivalent AaeFd4 and CpFd variants reversibly interconvert between similar valence-localized S = 1/2 and valence-delocalized S = 9/2 forms as a function of pH, with pKa values in the range 8.3–9.0, because of protonation
of the coordinated serinate residue. However, freezing high-pH samples
results in partial or full conversion from valence-delocalized S = 9/2 to valence-localized S = 1/2 [Fe2S2]+ clusters. MCD saturation magnetization
data for valence-delocalized S = 9/2 [Fe2S2]+ centers facilitated determination of transition
polarizations and thereby assignments of low-energy MCD bands associated
with the Fe–Fe interaction. The assignments provide experimental
assessment of the double exchange parameter, B, for
valence-delocalized [Fe2S2]+ centers
and demonstrate that variable-temperature MCD spectroscopy provides
a means of detecting and investigating the properties of valence-delocalized S = 9/2 [Fe2S2]+ fragments
in higher nuclearity Fe–S clusters. The origin of valence delocalization
in thioredoxin-like ferredoxin Cys-to-Ser variants and Fe–S
clusters in general is discussed in light of these results.
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EPRFe 2S Centersequivalent AaeFd 4spectroscopic studiesCpFd variantsCpFd variants reversibly interconvertAquifex aeolicus ferredoxin 4Fe 2S ferredoxinUVFe 2S fragmentsC 56Stransition polarizationsFe 2S centersvalence delocalizationMCD saturation magnetization dataFe 2S clustersRedox Studiesbuilding blockequivalent variantsserinate residuepKa valuesexchange parameterC 60S variants
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