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Geoffrey R. Moore - One of the best experts on this subject based on the ideXlab platform.
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The B-type Channel Is a Major Route for Iron Entry into the Ferroxidase Center and Central Cavity of Bacterioferritin
Journal of Biological Chemistry, 2014Co-Authors: Steve G. Wong, Geoffrey R. Moore, Jason C. Grigg, Nick E. Le Brun, Michael E. P. Murphy, A. Grant MaukAbstract:Bacterioferritin is a bacterial iron storage and detoxification protein that is capable of forming a ferric oxyhydroxide mineral core within its central cavity. To do this, iron must traverse the Bacterioferritin protein shell, which is expected to occur through one or more of the channels through the shell identified by structural studies. The size and negative electrostatic potential of the 24 B-type channels suggest that they could provide a route for iron into Bacterioferritin. Residues at the B-type channel (Asn-34, Glu-66, Asp-132, and Asp-139) of E. coli Bacterioferritin were substituted to determine if they are important for iron core formation. A significant decrease in the rates of initial oxidation of Fe(II) at the ferroxidase center and subsequent iron mineralization was observed for the D132F variant. The crystal structure of this variant shows that substitution of residue 132 with phenylalanine caused a steric blockage of the B-type channel and no other material structural perturbation. We conclude that the B-type channel is a major route for iron entry into both the ferroxidase center and the iron storage cavity of Bacterioferritin.
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Structural basis for iron mineralization by Bacterioferritin
Journal of the American Chemical Society, 2009Co-Authors: Allister Crow, Geoffrey R. Moore, T. Lawson, Allison Lewin, Nick E. Le BrunAbstract:Ferritin proteins function to detoxify, solubilize and store cellular iron by directing the synthesis of a ferric oxyhydroxide mineral solubilized within the protein’s central cavity. Here, through the application of X-ray crystallographic and kinetic methods, we report significant new insight into the mechanism of mineralization in a Bacterioferritin (BFR). The structures of nonheme iron-free and di-Fe2+ forms of BFR showed that the intrasubunit catalytic center, known as the ferroxidase center, is preformed, ready to accept Fe2+ ions with little or no reorganization. Oxidation of the di-Fe2+ center resulted in a di-Fe3+ center, with bridging electron density consistent with a μ-oxo or hydro bridged species. The μ-oxo bridged di-Fe3+ center appears to be stable, and there is no evidence that Fe3+species are transferred into the core from the ferroxidase center. Most significantly, the data also revealed a novel Fe2+ binding site on the inner surface of the protein, lying ∼10 A directly below the ferroxid...
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Effect of phosphate on Bacterioferritin-catalysed iron(II) oxidation
JBIC Journal of Biological Inorganic Chemistry, 2004Co-Authors: Helen Aitken-rogers, Geoffrey R. Moore, Allison Lewin, Chloe Singleton, Alice Taylor-gee, Nick E. Le BrunAbstract: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 °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 ~2 mM phosphate (for iron(II) concentrations
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effect of phosphate on Bacterioferritin catalysed iron ii oxidation
Journal of Biological Inorganic Chemistry, 2004Co-Authors: Helen Aitkenrogers, Geoffrey R. Moore, Allison Lewin, Chloe Singleton, Alice Taylorgee, Nick Le E BrunAbstract: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 °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 ~2 mM phosphate (for iron(II) concentrations <200 μM). 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.
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iron detoxification properties of escherichia coli Bacterioferritin attenuation of oxyradical chemistry
Journal of Biological Chemistry, 2002Co-Authors: Fadi Bouabdallah, Geoffrey R. Moore, Allison Lewin, Nick Le E Brun, Dennis N. ChasteenAbstract:Abstract Bacterioferritin (EcBFR) ofEscherichia coli is an iron-mineralizing hemoprotein composed of 24 identical subunits, each containing a dinuclear metal-binding site known as the “ferroxidase center.” The chemistry of Fe(II) binding and oxidation and Fe(III) hydrolysis using H2O2 as oxidant was studied by electrode oximetry, pH-stat, UV-visible spectrophotometry, and electron paramagnetic resonance spin trapping experiments. Absorption spectroscopy data demonstrate the oxidation of two Fe(II) per H2O2 at the ferroxidase center, thus avoiding hydroxyl radical production via Fenton chemistry. The oxidation reaction with H2O2 corresponds to [Fe(II)2-P]Z + H2O2→ [Fe(III)2O-P]Z + H2O, where [Fe(II)2-P]Z represents a diferrous ferroxidase center complex of the protein P with net charge Z and [Fe(III)2O-P]Z a μ-oxo-bridged diferric ferroxidase complex. The mineralization reaction is given by 2Fe2+ + H2O2 + 2H2O → 2FeOOH(core) + 4H+, where two Fe(II) are again oxidized by one H2O2. Hydrogen peroxide is shown to be an intermediate product of dioxygen reduction when O2 is used as the oxidant in both the ferroxidation and mineralization reactions. Most of the H2O2produced from O2 is rapidly consumed in a subsequent ferroxidase reaction with Fe(II) to produce H2O. EPR spin trapping experiments show that the presence of EcBFR greatly attenuates the production of hydroxyl radical during Fe(II) oxidation by H2O2, consistent with the ability of the Bacterioferritin to facilitate the pairwise oxidation of Fe(II) by H2O2, thus avoiding odd electron reduction products of oxygen and therefore oxidative damage to the protein and cellular components through oxygen radical chemistry.
Mario Rivera - One of the best experts on this subject based on the ideXlab platform.
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The Structure of the BfrB–Bfd Complex Reveals Protein–Protein Interactions Enabling Iron Release from Bacterioferritin
2016Co-Authors: Huili Yao, Scott Lovell, Kevin P. Battaile, Y. Wang, Ritesh Kumar, Anatoly M. Ruvinsky, Ilya A. Vakser, Mario RiveraAbstract:Ferritin-like molecules are unique to cellular iron homeostasis because they can store iron at concentrations much higher than those dictated by the solubility of Fe3+. Very little is known about the protein interactions that deliver iron for storage or promote the mobilization of stored iron from ferritin-like molecules. Here, we report the X-ray crystal structure of Pseudomonas aeruginosa Bacterioferritin (Pa-BfrB) in complex with Bacterioferritin-associated ferredoxin (Pa-Bfd) at 2.0 Å resolution. As the first example of a ferritin-like molecule in complex with a cognate partner, the structure provides unprecedented insight into the complementary interface that enables the [2Fe-2S] cluster of Pa-Bfd to promote heme-mediated electron transfer through the BfrB protein dielectric (∼18 Å), a process that is necessary to reduce the core ferric mineral and facilitate mobilization of Fe2+. The Pa-BfrB–Bfd complex also revealed the first structure of a Bfd, thus providing a first view to what appears to be a versatile metal binding domain ubiquitous to the large Fer2_BFD family of proteins and enzymes with diverse functions. Residues at the Pa-BfrB–Bfd interface are highly conserved in Bfr and Bfd sequences from a number of pathogenic bacteria, suggesting that the specific recognition between Pa-BfrB and Pa-Bfd is of widespread significance to the understanding of bacterial iron homeostasis
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characterization of the Bacterioferritin Bacterioferritin associated ferredoxin protein protein interaction in solution and determination of binding energy hot spots
Biochemistry, 2015Co-Authors: Y Wang, H. Yao, Scott Lovell, Kevin P. Battaile, Y Cheng, C R Midaugh, Mario RiveraAbstract:Mobilization of iron stored in the interior cavity of BfrB requires electron transfer from the [2Fe–2S] cluster in Bfd to the core iron in BfrB. A crystal structure of the Pseudomonas aeruginosa BfrB:Bfd complex revealed that BfrB can bind up to 12 Bfd molecules at 12 structurally identical binding sites, placing the [2Fe–2S] cluster of each Bfd immediately above a heme group in BfrB [Yao, H., et al. (2012) J. Am. Chem. Soc., 134, 13470–13481]. We report here a study aimed at characterizing the strength of the P. aeruginosa BfrB:Bfd association using surface plasmon resonance and isothermal titration calorimetry as well as determining the binding energy hot spots at the protein–protein interaction interface. The results show that the 12 Bfd-binding sites on BfrB are equivalent and independent and that the protein–protein association at each of these sites is driven entropically and is characterized by a dissociation constant (Kd) of approximately 3 μM. Determination of the binding energy hot spots was car...
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Protein dynamics and ion traffic in Bacterioferritin.
Biochemistry, 2012Co-Authors: Huan Rui, Mario RiveraAbstract:Bacterioferritin (Bfr) is a spherical protein composed of 24 subunits and 12 heme molecules. Bfrs contribute to regulate iron homeostasis in bacteria by capturing soluble but potentially toxic Fe(2+) and by compartmentalizing it in the form of a bioavailable ferric mineral inside the protein's hollow cavity. When iron is needed, Fe(3+) is reduced and mobilized into the cytosol as Fe(2+). Hence, key to the function of Bfr is its ability to permeate iron ions in and out of its interior cavity, which is likely imparted by a flexible protein shell. To examine the conformational flexibility of Bfrs in a native-like environment and the way in which the protein shell interacts with monovalent cations, we have performed molecular dynamics (MD) simulations of BfrB from Pseudomonas aeruginosa (Pa BfrB) in K(2)HPO(4) solutions at different ionic strengths. The results indicate the presence of coupled thermal fluctuations (dynamics) in the 4-fold pores and B-pores of the protein, which is key to allowing passage of monovalent cations through the protein shell using B-pores as conduits. The MD simulations also show that Pa BfrB ferroxidase centers are highly dynamic and permanently populated by transient cations exchanging with other cations in the interior cavity, as well as the solution bathing the protein. Taken together, the findings suggest that Fe(2+) passes across the Pa BfrB shell via B-pores and that the ferroxidase pores allow the capture and oxidation of Fe(2+), followed by translocation of Fe(3+) to the interior cavity, aided by the conformationally active H130.
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The structure of the BfrB-Bfd complex reveals protein-protein interactions enabling iron release from Bacterioferritin.
Journal of the American Chemical Society, 2012Co-Authors: H. Yao, Scott Lovell, Kevin P. Battaile, Y. Wang, Ritesh Kumar, Anatoly M. Ruvinsky, Ilya A. Vakser, Mario RiveraAbstract:Ferritin-like molecules are unique to cellular iron homeostasis because they can store iron at concentrations much higher than those dictated by the solubility of Fe3+. Very little is known about the protein interactions that deliver iron for storage or promote the mobilization of stored iron from ferritin-like molecules. Here, we report the X-ray crystal structure of Pseudomonas aeruginosa Bacterioferritin (Pa-BfrB) in complex with Bacterioferritin-associated ferredoxin (Pa-Bfd) at 2.0 A resolution. As the first example of a ferritin-like molecule in complex with a cognate partner, the structure provides unprecedented insight into the complementary interface that enables the [2Fe-2S] cluster of Pa-Bfd to promote heme-mediated electron transfer through the BfrB protein dielectric (∼18 A), a process that is necessary to reduce the core ferric mineral and facilitate mobilization of Fe2+. The Pa-BfrB–Bfd complex also revealed the first structure of a Bfd, thus providing a first view to what appears to be a v...
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Two distinct ferritin-like molecules in Pseudomonas aeruginosa: the product of the bfrA gene is a bacterial ferritin (FtnA) and not a Bacterioferritin (Bfr).
Biochemistry, 2011Co-Authors: H. Yao, Grace Jepkorir, Scott Lovell, Pavithra Vani Nama, Saroja Weeratunga, Kevin P. Battaile, Mario RiveraAbstract:Two distinct types of ferritin-like molecules often coexist in bacteria, the heme binding Bacterioferritins (Bfr) and the non-heme binding bacterial ferritins (Ftn). The early isolation of a ferritin-like molecule from Pseudomonas aeruginosa suggested the possibility of a Bacterioferritin assembled from two different subunits [Moore, G. R., et al. (1994) Biochem. J. 304, 493-497]. Subsequent studies demonstrated the presence of two genes encoding ferritin-like molecules in P. aeruginosa, designated bfrA and bfrB, and suggested that two distinct Bacterioferritins may coexist [Ma, J.-F., et al. (1999) J. Bacteriol. 181, 3730-3742]. In this report, we present structural evidence demonstrating that the product of the bfrA gene is a ferritin-like molecule not capable of binding heme that harbors a catalytically active ferroxidase center with structural properties similar to those characteristic of bacterial and archaeal Ftns and clearly distinct from those of the ferroxidase center typical of Bfrs. Consequently, the product of the bfrA gene in P. aeruginosa is a bacterial ferritin, which we propose should be termed Pa FtnA. These results, together with the previous characterization of the product of the bfrB gene as a genuine Bacterioferritin (Pa BfrB) [Weeratunga, S. J., et al. (2010) Biochemistry 49, 1160-1175], indicate the coexistence of a bacterial ferritin (Pa FtnA) and a Bacterioferritin (Pa BfrB) in P. aeruginosa. In agreement with this idea, we also obtained evidence demonstrating that release of iron from Pa BfrB and Pa FtnA is likely subject to different regulation in P. aerugionsa. Whereas the efficient release of iron stored in Pa FtnA requires only the input of electrons from a ferredoxin NADP reductase (Pa Fpr), the release of iron stored in Pa BfrB requires not only electron delivery by Pa Fpr but also the presence of a "regulator", the apo form of a Bacterioferritin-associated ferredoxin (apo Pa Bfd). Finally, structural analysis of iron uptake in crystallo suggests a possible pathway for the internalization of ferroxidase iron into the interior cavity of Pa FtnA.
Ligia M Saraiva - One of the best experts on this subject based on the ideXlab platform.
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Bacterioferritin protects the anaerobe Desulfovibrio vulgaris Hildenborough against oxygen.
Anaerobe, 2012Co-Authors: Mafalda C. O. Figueiredo, Susana A.l. Lobo, João N. Carita, Lígia S. Nobre, Ligia M SaraivaAbstract:Intracellular free iron, is under aerobic conditions and via the Fenton reaction a catalyst for the formation of harmful reactive oxygen species. In this article, we analyzed the relation between intracellular iron storage and oxidative stress response in the sulfate reducing bacterium Desulfovibrio vulgaris Hildenborough, an anaerobe that is often found in oxygenated niches. To this end, we investigated the role of the iron storage protein Bacterioferritin using transcriptomic and physiological approaches. We observed that transcription of Bacterioferritin is strongly induced upon exposure of cells to an oxygenated atmosphere. When grown in the presence of high concentrations of oxygen the D. vulgaris Bacterioferritin mutant exhibited, in comparison with the wild type strain, lower viability and a higher content of intracellular reactive oxygen species. Furthermore, the Bacterioferritin gene is under the control of the oxidative stress response regulator D. vulgaris PerR. Altogether the data revealed a previously unrecognized ability for the iron storage Bacterioferritin to contribute to the oxygen tolerance exhibited by D. vulgaris.
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the genetic organization of desulfovibrio desulphuricans atcc 27774 Bacterioferritin and rubredoxin 2 genes involvement of rubredoxin in iron metabolism
Molecular Microbiology, 2001Co-Authors: Patricia N Da Costa, Celia V Romao, Eurico Melo, Miguel Teixeira, Antonio V Xavier, Jean Legall, Ligia M SaraivaAbstract:The anaerobic bacterium Desulfovibrio desulphuricans ATCC 27774 contains a unique Bacterioferritin, isolated with a stable di-iron centre and having iron-coproporphyrin III as its haem cofactor, as well as a type 2 rubredoxin with an unusual spacing of four amino acid residues between the first two binding cysteines. The genes encoding for these two proteins were cloned and sequenced. The deduced amino acid sequence of the Bacterioferritin shows that it is among the most divergent members of this protein family. Most interestingly, the Bacterioferritin and rubredoxin-2 genes form a dicistronic operon, which reflects the direct interaction between the two proteins. Indeed, Bacterioferritin and rubredoxin-2 form a complex in vitro, as shown by the significant increase in the anisotropy and decay times of the fluorescence of rubredoxin-2 tryptophan(s) when mixed with Bacterioferritin. In addition, rubredoxin-2 donates electrons to Bacterioferritin. This is the first identification of an electron donor to a Bacterioferritin and shows the involvement of rubredoxin-2 in iron metabolism. Furthermore, analysis of the genomic data for anaerobes suggests that rubredoxins play a general role in iron metabolism and oxygen detoxification in these prokaryotes.
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The genetic organization of Desulfovibrio desulphuricans ATCC 27774 Bacterioferritin and rubredoxin‐2 genes: involvement of rubredoxin in iron metabolism
Molecular Microbiology, 2001Co-Authors: Patricia N Da Costa, Celia V Romao, Eurico Melo, Miguel Teixeira, Antonio V Xavier, Jean Legall, Ligia M SaraivaAbstract:The anaerobic bacterium Desulfovibrio desulphuricans ATCC 27774 contains a unique Bacterioferritin, isolated with a stable di-iron centre and having iron-coproporphyrin III as its haem cofactor, as well as a type 2 rubredoxin with an unusual spacing of four amino acid residues between the first two binding cysteines. The genes encoding for these two proteins were cloned and sequenced. The deduced amino acid sequence of the Bacterioferritin shows that it is among the most divergent members of this protein family. Most interestingly, the Bacterioferritin and rubredoxin-2 genes form a dicistronic operon, which reflects the direct interaction between the two proteins. Indeed, Bacterioferritin and rubredoxin-2 form a complex in vitro, as shown by the significant increase in the anisotropy and decay times of the fluorescence of rubredoxin-2 tryptophan(s) when mixed with Bacterioferritin. In addition, rubredoxin-2 donates electrons to Bacterioferritin. This is the first identification of an electron donor to a Bacterioferritin and shows the involvement of rubredoxin-2 in iron metabolism. Furthermore, analysis of the genomic data for anaerobes suggests that rubredoxins play a general role in iron metabolism and oxygen detoxification in these prokaryotes.
Patricia N Da Costa - One of the best experts on this subject based on the ideXlab platform.
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the genetic organization of desulfovibrio desulphuricans atcc 27774 Bacterioferritin and rubredoxin 2 genes involvement of rubredoxin in iron metabolism
Molecular Microbiology, 2001Co-Authors: Patricia N Da Costa, Celia V Romao, Eurico Melo, Miguel Teixeira, Antonio V Xavier, Jean Legall, Ligia M SaraivaAbstract:The anaerobic bacterium Desulfovibrio desulphuricans ATCC 27774 contains a unique Bacterioferritin, isolated with a stable di-iron centre and having iron-coproporphyrin III as its haem cofactor, as well as a type 2 rubredoxin with an unusual spacing of four amino acid residues between the first two binding cysteines. The genes encoding for these two proteins were cloned and sequenced. The deduced amino acid sequence of the Bacterioferritin shows that it is among the most divergent members of this protein family. Most interestingly, the Bacterioferritin and rubredoxin-2 genes form a dicistronic operon, which reflects the direct interaction between the two proteins. Indeed, Bacterioferritin and rubredoxin-2 form a complex in vitro, as shown by the significant increase in the anisotropy and decay times of the fluorescence of rubredoxin-2 tryptophan(s) when mixed with Bacterioferritin. In addition, rubredoxin-2 donates electrons to Bacterioferritin. This is the first identification of an electron donor to a Bacterioferritin and shows the involvement of rubredoxin-2 in iron metabolism. Furthermore, analysis of the genomic data for anaerobes suggests that rubredoxins play a general role in iron metabolism and oxygen detoxification in these prokaryotes.
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The genetic organization of Desulfovibrio desulphuricans ATCC 27774 Bacterioferritin and rubredoxin‐2 genes: involvement of rubredoxin in iron metabolism
Molecular Microbiology, 2001Co-Authors: Patricia N Da Costa, Celia V Romao, Eurico Melo, Miguel Teixeira, Antonio V Xavier, Jean Legall, Ligia M SaraivaAbstract:The anaerobic bacterium Desulfovibrio desulphuricans ATCC 27774 contains a unique Bacterioferritin, isolated with a stable di-iron centre and having iron-coproporphyrin III as its haem cofactor, as well as a type 2 rubredoxin with an unusual spacing of four amino acid residues between the first two binding cysteines. The genes encoding for these two proteins were cloned and sequenced. The deduced amino acid sequence of the Bacterioferritin shows that it is among the most divergent members of this protein family. Most interestingly, the Bacterioferritin and rubredoxin-2 genes form a dicistronic operon, which reflects the direct interaction between the two proteins. Indeed, Bacterioferritin and rubredoxin-2 form a complex in vitro, as shown by the significant increase in the anisotropy and decay times of the fluorescence of rubredoxin-2 tryptophan(s) when mixed with Bacterioferritin. In addition, rubredoxin-2 donates electrons to Bacterioferritin. This is the first identification of an electron donor to a Bacterioferritin and shows the involvement of rubredoxin-2 in iron metabolism. Furthermore, analysis of the genomic data for anaerobes suggests that rubredoxins play a general role in iron metabolism and oxygen detoxification in these prokaryotes.
Nick Le E Brun - One of the best experts on this subject based on the ideXlab platform.
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effect of phosphate on Bacterioferritin catalysed iron ii oxidation
Journal of Biological Inorganic Chemistry, 2004Co-Authors: Helen Aitkenrogers, Geoffrey R. Moore, Allison Lewin, Chloe Singleton, Alice Taylorgee, Nick Le E BrunAbstract: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 °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 ~2 mM phosphate (for iron(II) concentrations <200 μM). 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.
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iron detoxification properties of escherichia coli Bacterioferritin attenuation of oxyradical chemistry
Journal of Biological Chemistry, 2002Co-Authors: Fadi Bouabdallah, Geoffrey R. Moore, Allison Lewin, Nick Le E Brun, Dennis N. ChasteenAbstract:Abstract Bacterioferritin (EcBFR) ofEscherichia coli is an iron-mineralizing hemoprotein composed of 24 identical subunits, each containing a dinuclear metal-binding site known as the “ferroxidase center.” The chemistry of Fe(II) binding and oxidation and Fe(III) hydrolysis using H2O2 as oxidant was studied by electrode oximetry, pH-stat, UV-visible spectrophotometry, and electron paramagnetic resonance spin trapping experiments. Absorption spectroscopy data demonstrate the oxidation of two Fe(II) per H2O2 at the ferroxidase center, thus avoiding hydroxyl radical production via Fenton chemistry. The oxidation reaction with H2O2 corresponds to [Fe(II)2-P]Z + H2O2→ [Fe(III)2O-P]Z + H2O, where [Fe(II)2-P]Z represents a diferrous ferroxidase center complex of the protein P with net charge Z and [Fe(III)2O-P]Z a μ-oxo-bridged diferric ferroxidase complex. The mineralization reaction is given by 2Fe2+ + H2O2 + 2H2O → 2FeOOH(core) + 4H+, where two Fe(II) are again oxidized by one H2O2. Hydrogen peroxide is shown to be an intermediate product of dioxygen reduction when O2 is used as the oxidant in both the ferroxidation and mineralization reactions. Most of the H2O2produced from O2 is rapidly consumed in a subsequent ferroxidase reaction with Fe(II) to produce H2O. EPR spin trapping experiments show that the presence of EcBFR greatly attenuates the production of hydroxyl radical during Fe(II) oxidation by H2O2, consistent with the ability of the Bacterioferritin to facilitate the pairwise oxidation of Fe(II) by H2O2, thus avoiding odd electron reduction products of oxygen and therefore oxidative damage to the protein and cellular components through oxygen radical chemistry.
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an epr investigation of non haem iron sites in escherichia coli Bacterioferritin and their interaction with phosphate a study using nitric oxide as a spin probe
FEBS Letters, 1993Co-Authors: Nick Le E Brun, Geoffrey R. Moore, Simon C. Andrews, John R. Guest, Andrew J. Thomson, Myles R. Cheesman, Pauline M. HarrisonAbstract:EPR studies of Bacterioferritin (BFR), an iron-storage protein of Escherichia coli [1993, Biochem. J. 292, 47-56.], have revealed the presence of non-haem iron (III) (NHI) sites within the protein coat which may be involved in iron uptake and release. When nitric oxide was used as an EPR spin probe of the Fe(II) state of the NHI sites, two distinct mononuclear NHI species were found. Under certain conditions, an iron dimer was also observed. The reaction of phosphate with NHI species has been investigated. Results point to a function for this anion in core nucleation.
