Ferroxidase kinetics of horse spleen apoferritin.

Sun, Shujun; Chasteen, N. Dennis

doi:10.1016/s0021-9258(19)74019-8

Cited by 91 publications

(81 citation statements)

References 40 publications

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“…This side chain might bind directly to the Mn(II), or protonated histidine might block access to the binding site. The temperature dependence of the lag rate (Figure C, green stars) yields an activation energy of 44 kJ/mol, close to that reported for Fe 2+ binding to apoferritin …”

Section: Resultsmentioning

confidence: 99%

Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism

Soldatova

Tao²,

Romano

et al. 2017

J. Am. Chem. Soc.

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The bacterial protein complex Mnx contains a multicopper oxidase (MCO) MnxG that, unusually, catalyzes the two-electron oxidation of Mn(II) to MnO biomineral, via a Mn(III) intermediate. Although Mn(III)/Mn(II) and Mn(IV)/Mn(III) reduction potentials are expected to be high, we find a low reduction potential, 0.38 V (vs Normal Hydrogen Electrode, pH 7.8), for the MnxG type 1 Cu, the electron acceptor. Indeed the type 1 Cu is not reduced by Mn(II) in the absence of molecular oxygen, indicating that substrate oxidation requires an activation step. We have investigated the enzyme mechanism via electronic absorption spectroscopy, using chemometric analysis to separate enzyme-catalyzed MnO formation from MnO nanoparticle aging. The nanoparticle aging time course is characteristic of nucleation and particle growth; rates for these processes followed expected dependencies on Mn(II) concentration and temperature, but exhibited different pH optima. The enzymatic time course is sigmoidal, signaling an activation step, prior to turnover. The Mn(II) concentration and pH dependence of a preceding lag phase indicates weak Mn(II) binding. The activation step is enabled by a pK > 8.6 deprotonation, which is assigned to Mn(II)-bound HO; it induces a conformation change (consistent with a high activation energy, 106 kJ/mol) that increases Mn(II) affinity. Mnx activation is proposed to decrease the Mn(III/II) reduction potential below that of type 1 Cu(II/I) by formation of a hydroxide-bridged binuclear complex, Mn(II)(μ-OH)Mn(II), at the substrate site. Turnover is found to depend cooperatively on two Mn(II) and is enabled by a pK 7.6 double deprotonation. It is proposed that turnover produces a Mn(III)(μ-OH)Mn(III) intermediate that proceeds to the enzyme product, likely Mn(IV)(μ-O)Mn(IV) or an oligomer, which subsequently nucleates MnO nanoparticles. We conclude that Mnx exploits manganese polynuclear chemistry in order to facilitate an otherwise difficult oxidation reaction, as well as biomineralization. The mechanism of the Mn(III/IV) conversion step is elucidated in an accompanying paper .

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Section: Resultsmentioning

confidence: 99%

Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism

Soldatova

Tao²,

Romano

et al. 2017

J. Am. Chem. Soc.

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“…Ferritins were reconstituted to final iron loadings of 500, 1000, 1500, 2000 or 2500 Fe/shell in both low and high iron fluxes or to a level where precipitation was observed. For the low iron flux samples, a series of additions at 50 Fe(II)/shell were made, allowing at least 10 minutes between additions for the complete oxidation of Fe(II) [69]. The 1 mL ferritin solutions were open to air and constantly stirred to maintain molecular oxygen in the sample.…”

Section: Reconstitution Of Ferritinsmentioning

confidence: 99%

“…The high iron flux samples were made using a maximal single addition of 1000 Fe(II)/shell at a protein concentration of 0.25 mg/mL to keep the sample from going anoxic during the course of the experiment. Under these conditions about 50% of the dissolved O 2 would be consumed based on the known oxidation stoichiometry of 4 Fe(II)/O 2 [69,70].…”

Section: Reconstitution Of Ferritinsmentioning

confidence: 99%

The sedimentation properties of ferritins. New insights and analysis of methods of nanoparticle preparation

May

Grady

Laue

et al. 2010

Biochimica et Biophysica Acta (BBA) - General Subjects

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Background-Ferritin exhibits complex behavior in the ultracentrifuge due to variability in iron core size among molecules. A comprehensive study was undertaken to develop procedures for obtaining more uniform cores and assessing their homogeneity.Methods-Analytical ultracentrifugation was used to measure the mineral core size distributions obtained by adding iron under high-and low-flux conditions to horse spleen (apoHoSF) and human H-chain (apoHuHF) apoferritins.Results-More uniform core sizes are obtained with the homopolymer human H-chain ferritin than with the heteropolymer horse spleen HoSF protein in which subpopulations of HoSF molecules with varying iron content are observed. A binomial probability distribution of H-and Lsubunits among protein shells qualitatively accounts for the observed subpopulations. The addition of Fe 2+ to apoHuHF produces iron core particle size diameters from 3.8 ± 0.3 to 6.2 ± 0.3 nm. Diameters from 3.4 ± 0.6 to 6.5 ± 0.6 nm are obtained with natural HoSF after sucrose gradient fractionation. The change in the sedimentation coefficient as iron accumulates in ferritin suggests that the protein shell contracts ~10% to a more compact structure, a finding consistent with published electron micrographs. The physicochemical parameters for apoHoSF (15%/85% H/L subunits) are M = 484,120 g/mol, ν̄ = 0.735mL/g, s 20,w = 17.0 S and D 20,W = 3.21 × 10 −7 cm 2 /s; and for apoHuHF M = 506,266 g/mol, ν̄ = 0.724 mL/g, s 20,w = 18.3 S and D 20,w = 3.18 × 10 −7 cm 2 /s.Significance-The methods presented here should prove useful in the synthesis of size controlled nanoparticles of other minerals.

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“…Iron biomineralization in ferritins is a multistep process involving Fe II sequestration and binding, ferroxidase catalyzed oxidation of iron (FC), Fe III hydrolysis and finally Fe III migration and nucleation of the mineral core. [10][11][12] The natural biological function of ferritins in iron sequestration and biomineralization has served as a model and inspiration for our efforts to use a biomimetic strategy for nanomaterial synthesis. 13 While the antiferromagnetic properties of iron oxide (Fe(O)OH) minerals naturally formed within the ferritin cage are not of considerable biotechnical interest; Mann and co-workers showed that Fe 3 O 4 (or c-Fe 2 O 3 ) nanoparticles could be artificially synthesized within empty ferritin (apo-ferritin) cages under conditions of elevated temperature and pH.…”

Section: Inspired By Naturementioning

confidence: 99%

Bioprospecting in high temperature environments; application of thermostable protein cages

et al. 2007

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Ferroxidase kinetics of horse spleen apoferritin.

Cited by 91 publications

References 40 publications

Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism

Mn(II) Oxidation by the Multicopper Oxidase Complex Mnx: A Binuclear Activation Mechanism

The sedimentation properties of ferritins. New insights and analysis of methods of nanoparticle preparation

Bioprospecting in high temperature environments; application of thermostable protein cages

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