The electronic and magnetic properties of rubredoxine a low-temperature magnetic circular dichroism study

Bennett, Deborah E.; Johnson, Michael K.

doi:10.1016/0167-4838(87)90272-x

Cited by 64 publications

(51 citation statements)

References 33 publications

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“…Figure 1A shows that purified hIscA1 has an absorption peak at 315 nm, indicative of iron binding in the protein [22, 26, 32]. The iron content analysis showed that as-purified hIscA1 contained about 0.1–0.2 iron atoms per hIscA1 dimer (n = 3).…”

Section: Resultsmentioning

confidence: 99%

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

Lü

Bitoun

Tan

et al. 2010

Biochemical Journal

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SYNOPSIS A human homologue of the iron-sulfur cluster assembly protein IscA (hIscA1) has been cloned and expressed in Escherichia coli cells. The UV-visible absorption and EPR (electron paramagnetic resonance) measurements reveal that hIscA1 purified from E. coli cells contains a mononuclear iron center and that the iron binding in hIscA1 expressed in E. coli cells can be further modulated by the iron content in the cell growth medium. Additional studies show that purified hIscA1 binds iron with an iron association constant of approx. 2.0 × 1019 M−1, and that the iron-bound hIscA1 is able to provide the iron for the iron-sulfur cluster assembly in a proposed scaffold protein IscU of E. coli in vitro. The complementation experiments indicate that hIscA1 can partially substitute for IscA in restoring the cell growth of E. coli in the M9 minimal medium under aerobic conditions. The results suggest that human IscA1, like E. coli IscA, is an iron binding protein that may act as an iron chaperone for biogenesis of iron-sulfur clusters.

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Section: Resultsmentioning

confidence: 99%

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

Lü

Bitoun

Tan

et al. 2010

Biochemical Journal

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“…The offset of the low temperature isotherms from those at higher temperatures is characteristic of low lying excited states and rhombic zero-field splitting (ZFS) of non-Kramers doublet ground states (21). Ground state parameters for a non-Kramers system can be obtained through doublet fitting of the VTVH MCD intensity (Figure 5–6) using equation 2 (18, 22–24):

Δ ε = \sum_{i} true[{(A_{s a t lim})}_{i} true(\int_{0}^{\frac{π}{2}} \frac{{cos}^{2} θ sin θ}{Γ_{i}} g_{‖ i} β H α_{i} d θ - \sqrt{2} \frac{M_{z}}{M_{xy}} \int_{0}^{\frac{π}{2}} \frac{{sin}^{3} θ}{Γ_{i}} g_{⊥ i} β H α_{i} d θ true) + B_{i} H γ_{i} true]

where

\begin{matrix} {normalΓ}_{normali} = \sqrt{δ_{normali}^{2} + {({normalg}_{‖ i} β H cos θ)}^{2} + {({normalg}_{⊥ i} β H sin θ)}^{2}} \\ \begin{matrix} α_{i} = \frac{e^{- (E_{i} - {normalΓ}_{i} / 2) / kT} - e^{- (E_{i} + {normalΓ}_{i} / 2) / kT}}{\sum_{j}} \end{matrix} \end{matrix}

…”

Section: Resultsmentioning

confidence: 99%

CD and MCD Spectroscopic Studies of the Two Dps Miniferritin Proteins from Bacillus anthracis: Role of O₂ and H₂O₂ Substrates in Reactivity of the Diiron Catalytic Centers

Schwartz

Liu²,

Tosha³

et al. 2010

Biochemistry

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DNA Protection during Starvation (Dps) proteins are mini-ferritins found in bacteria and archaea that provide protection from uncontrolled Fe(II)/O radical chemistry; thus the catalytic sites are targets for antibiotics against pathogens, such as anthrax. Ferritin protein cages synthesize ferric oxymineral from Fe(II) and O 2 /H 2 O 2 , which accumulates in the large central cavity; for Dps, H 2 O 2 , is the more common Fe(II) oxidant contrasting with eukaryotic maxi-ferritins that often prefer dioxygen. To better understand the differences in the catalytic sites of maxi versus miniferritins, we used a combination of NIR circular dichroism (CD), magnetic circular dichroism (MCD), and variable-temperature, variable-field MCD (VTVH MCD) to study Fe(II) binding to the catalytic sites of the two B. anthracis mini-ferritins; one in which two Fe(II) react with O 2 exclusively (Dps1) and a second in which both O 2 or H 2 O 2 can react with two Fe(II) (Dps2). Both result in the formation of iron oxy-biomineral. The data show: a single 5 or 6-coordinate Fe(II) in the absence of oxidant; Fe(II) binding to Dps2 is 30 × more stable than Dps1; and the lower limit of K d for binding a second Fe(II), in the absence of oxidant, is 2-3 orders of magnitude weaker than for the binding of the single Fe(II). The data fit an equilibrium model where binding of oxidant facilitates formation of the catalytic site, in sharp contrast to eukaryotic M-ferritins where the binuclear Fe(II) centers are preformed before binding of O 2 . The two different binding sequences illustrate the mechanistic range possible for catalytic sites of the family of ferritins.DNA Protection during Starvation (Dps) proteins, also known as mini-ferritins, are 12-monomer spherical proteins capable of storing iron mineral and thus are part of the ferritin super-family (1-4). Unlike maxi (24-monomer) ferritins, Dps proteins have been shown to bind and protect DNA from oxidation (2-4). The paired Dps proteins in Bacillus anthracis * To whom correspondence should be addressed: Elizabeth C. Theil etheil@chori.org; and Edward I. Solomon, edward.solomon@stanford.edu;. Supporting Information Available CD spectra showing effect of temperature on Dps1 transitions; MCD spectra of Dps1 overlaid with Fe(II) control spectrum, and MCD spectrum with control subtracted; Apparent initial rates of Fe 2+ oxidation in Dps1 & 2 scaled to overlay with the calculated concentration of binuclear Fe(II) active sites occupied at a given Fe 2+ /Dps2 ratio using fixed K D1 and varied K D2 values. This material is available free of charge via the Internet at http://pubs.acs.org. NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2011 December 14. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript (Dps1 and Dps2), which share ~60% sequence homology(3), confer greatest protection when both are present. Dps proteins in pathogens, such as those from B. anthracis, are of particular interest as better knowledge of how these proteins p...

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“…Rubredoxin, whose redox active site consists of a single iron ligated by four cysteinyl sulfurs, is the simplest and most extensively studied iron−sulfur protein, with published investigations by NMR, 9−16 EPR, 17,18 MCD, 18,19 and Mössbauer 18,20 spectroscopies, X-ray crystallography, 21−23 and theoretical calculations. 13,14,24−26 Rubredoxin has served as a model system for investigations of the role of the metal in protein folding and assembly of the metal center.…”

Section: Introductionmentioning

confidence: 99%

Hyperfine-Shifted ¹³C and ¹⁵N NMR Signals from Clostridium pasteurianum Rubredoxin: Extensive Assignments and Quantum Chemical Verification

et al. 2009

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Stable isotope-labeling methods, coupled with novel techniques for detecting fast-relaxing NMR signals, now permit detailed investigations of paramagnetic centers of metalloproteins. We have utilized these advances to carry out comprehensive assignments of the hyperfine-shifted 13C and 15N signals of the rubredoxin from Clostridium pasteurianum (CpRd) in both its oxidized and reduced states. We used residue-specific labeling (by chemical synthesis) and residue-type-selective labeling (by biosynthesis) to assign signals detected by one-dimensional 15N NMR spectroscopy, to nitrogen atoms near the iron center. We refined and extended these 15N assignments to the adjacent carbonyl carbons by means of one-dimensional 13C[15N] decoupling difference experiments. We collected paramagnetic-optimized SuperWEFT 13C[13C] constant time COSY (SW-CT-COSY) data to complete the assignment of 13C signals of reduced CpRd. By following these 13C signals as the protein was gradually oxidized, we transferred these assignments to carbons in the oxidized state. We have compared these assignments with hyperfine chemical shifts calculated from available X-ray structures of CpRd in its oxidized and reduced forms. The results allow the evaluation of the X-ray structural models as representative of the solution structure of the protein, and they provide a framework for future investigation of the active site of this protein. The methods developed here should be applicable to other proteins that contain a paramagnetic center with high spin and slow electron exchange.

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The electronic and magnetic properties of rubredoxine a low-temperature magnetic circular dichroism study

Cited by 64 publications

References 33 publications

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

CD and MCD Spectroscopic Studies of the Two Dps Miniferritin Proteins from Bacillus anthracis: Role of O₂ and H₂O₂ Substrates in Reactivity of the Diiron Catalytic Centers

Hyperfine-Shifted ¹³C and ¹⁵N NMR Signals from Clostridium pasteurianum Rubredoxin: Extensive Assignments and Quantum Chemical Verification

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The electronic and magnetic properties of rubredoxine a low-temperature magnetic circular dichroism study

Cited by 64 publications

References 33 publications

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

Iron-binding activity of human iron–sulfur cluster assembly protein hIscA1

CD and MCD Spectroscopic Studies of the Two Dps Miniferritin Proteins from Bacillus anthracis: Role of O2 and H2O2 Substrates in Reactivity of the Diiron Catalytic Centers

Hyperfine-Shifted 13C and 15N NMR Signals from Clostridium pasteurianum Rubredoxin: Extensive Assignments and Quantum Chemical Verification

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CD and MCD Spectroscopic Studies of the Two Dps Miniferritin Proteins from Bacillus anthracis: Role of O₂ and H₂O₂ Substrates in Reactivity of the Diiron Catalytic Centers

Hyperfine-Shifted ¹³C and ¹⁵N NMR Signals from Clostridium pasteurianum Rubredoxin: Extensive Assignments and Quantum Chemical Verification