Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 54
Filtrar
1.
J Bacteriol ; 206(4): e0045223, 2024 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-38551342

RESUMO

The wobble bases of tRNAs that decode split codons are often heavily modified. In bacteria, tRNAGlu, Gln, Asp contains a variety of xnm5s2U derivatives. The synthesis pathway for these modifications is complex and fully elucidated only in a handful of organisms, including the Gram-negative Escherichia coli K12 model. Despite the ubiquitous presence of mnm5s2U modification, genomic analysis shows the absence of mnmC orthologous genes, suggesting the occurrence of alternate biosynthetic schemes for the conversion of cmnm5s2U to mnm5s2U. Using a combination of comparative genomics and genetic studies, a member of the YtqA subgroup of the radical Sam superfamily was found to be involved in the synthesis of mnm5s2U in both Bacillus subtilis and Streptococcus mutans. This protein, renamed MnmL, is encoded in an operon with the recently discovered MnmM methylase involved in the methylation of the pathway intermediate nm5s2U into mnm5s2U in B. subtilis. Analysis of tRNA modifications of both S. mutans and Streptococcus pneumoniae shows that growth conditions and genetic backgrounds influence the ratios of pathway intermediates owing to regulatory loops that are not yet understood. The MnmLM pathway is widespread along the bacterial tree, with some phyla, such as Bacilli, relying exclusively on these two enzymes. Although mechanistic details of these newly discovered components are not fully resolved, the occurrence of fusion proteins, alternate arrangements of biosynthetic components, and loss of biosynthetic branches provide examples of biosynthetic diversity to retain a conserved tRNA modification in Nature.IMPORTANCEThe xnm5s2U modifications found in several tRNAs at the wobble base position are widespread in bacteria where they have an important role in decoding efficiency and accuracy. This work identifies a novel enzyme (MnmL) that is a member of a subgroup of the very versatile radical SAM superfamily and is involved in the synthesis of mnm5s2U in several Gram-positive bacteria, including human pathogens. This is another novel example of a non-orthologous displacement in the field of tRNA modification synthesis, showing how different solutions evolve to retain U34 tRNA modifications.


Assuntos
Escherichia coli K12 , RNA de Transferência , Humanos , RNA de Transferência/genética , Escherichia coli K12/genética , Bactérias/genética , Metilação , Bactérias Gram-Positivas/genética
2.
Inorg Chem ; 61(32): 12625-12634, 2022 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-35920800

RESUMO

We report the isolation and characterization of a series of three cobalt(II) bis(phosphine) complexes with varying numbers of coordinated solvent ligands in the axial position. X-ray quality crystals of [Co(dppv)2][BF4]2 (1), [Co(dppv)2(NCCH3)][BPh4]2 (2), and [Co(dppv)2(NCCH3)2][BF4]2 (3) (dppv = cis-1,2-bis(diphenylphosphino)ethylene) were grown under slightly different conditions, and their structures were compared. This analysis revealed multiple crystallization motifs for divalent cobalt(II) complexes with the same set of phosphine ligands. Notably, the 4-coordinate complex 1 is a rare example of a square-planar cobalt(II) complex, the first crystallographically characterized square-planar Co(II) complex containing only neutral, bidentate ligands. Characterization of the different axial geometries via EPR and UV-visible spectroscopies showed that there is a very shallow energy landscape for axial ligation. Ligand field angular overlap model calculations support this conclusion, and we provide a strategy for tuning other ligands to be axially labile on a phosphine scaffold. This methodology is proposed to be used for designing cobalt phosphine catalysts for a variety of oxidation and reduction reactions.


Assuntos
Cobalto , Cobalto/química , Cristalografia por Raios X , Ligantes , Oxirredução , Fosfinas
3.
Inorg Chem ; 58(12): 7978-7988, 2019 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-31185562

RESUMO

Accessing covalent bonding interactions between actinides and ligating atoms remains a central problem in the field. Our current understanding of actinide bonding is limited because of a paucity of diverse classes of compounds and the lack of established models. We recently synthesized a thorium (Th)-aluminum (Al) heterobimetallic molecule that represents a new class of low-valent Th-containing compounds. To gain further insight into this system and actinide-metal bonding more generally, it is useful to study their underlying electronic structures. Here, we report characterization by electron paramagnetic resonance (EPR) and electron-nuclear double resonance (ENDOR) spectroscopy of two heterobimetallic compounds: (i) a Cptt2ThH3AlCTMS3 [TMS = Si(CH3)3; Cptt = 1,3-di- tert-butylcyclopentadienyl] complex with bridging hydrides and (ii) an actinide-free Cp2TiH3AlCTMS3 (Cp = cyclopentadienyl) analogue. Analyses of the hyperfine interactions between the paramagnetic trivalent metal centers and the surrounding magnetic nuclei, 1H and 27Al, yield spin distributions over both complexes. These results show that while the bridging hydrides in the two complexes have similar hyperfine couplings ( aiso = -9.7 and -10.7 MHz, respectively), the spin density on the Al ion in the Th3+ complex is ∼5-fold larger than that in the titanium(3+) (Ti3+) analogue. This suggests a direct orbital overlap between Th and Al, leading to a covalent interaction between Th and Al. Our quantitative investigation by a pulse EPR technique deepens our understanding of actinide bonding to main-group elements.

4.
Inorg Chem ; 58(4): 2336-2345, 2019 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-30730725

RESUMO

The modulation of the reactivity of metal oxo species by redox inactive metals has attracted much interest due to the observation of redox inactive metal effects on processes involving electron transfer both in nature (the oxygen-evolving complex of Photosystem II) and in heterogeneous catalysis (mixed-metal oxides). Studies of small-molecule models of these systems have revealed numerous instances of effects of redox inactive metals on electron- and group-transfer reactivity. However, the heterometallic species directly involved in these transformations have rarely been structurally characterized and are often generated in situ. We have previously reported the preparation and structural characterization of multiple series of heterometallic clusters based on Mn3 and Fe3 cores and described the effects of Lewis acidity of the heterometal incorporated in these complexes on cluster reduction potential. To determine the effects of Lewis acidity of redox inactive metals on group transfer reactivity in structurally well-defined complexes, we studied [Mn3MO4], [Mn3MO(OH)], and [Fe3MO(OH)] clusters in oxygen atom transfer (OAT) reactions with phosphine substrates. The qualitative rate of OAT correlates with the Lewis acidity of the redox inactive metal, confirming that Lewis acidic metal centers can affect the chemical reactivity of metal oxo species by modulating cluster electronics.

5.
Biochemistry ; 57(8): 1306-1315, 2018 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-29405700

RESUMO

The Radical SAM (RS) enzyme PqqE catalyzes the first step in the biosynthesis of the bacterial cofactor pyrroloquinoline quinone, forming a new carbon-carbon bond between two side chains within the ribosomally synthesized peptide substrate PqqA. In addition to the active site RS 4Fe-4S cluster, PqqE is predicted to have two auxiliary Fe-S clusters, like the other members of the SPASM domain family. Here we identify these sites and examine their structure using a combination of X-ray crystallography and Mössbauer and electron paramagnetic resonance (EPR) spectroscopies. X-ray crystallography allows us to identify the ligands to each of the two auxiliary clusters at the C-terminal region of the protein. The auxiliary cluster nearest the RS site (AuxI) is in the form of a 2Fe-2S cluster ligated by four cysteines, an Fe-S center not seen previously in other SPASM domain proteins; this assignment is further supported by Mössbauer and EPR spectroscopies. The second, more remote cluster (AuxII) is a 4Fe-4S center that is ligated by three cysteine residues and one aspartate residue. In addition, we examined the roles these ligands play in catalysis by the RS and AuxII clusters using site-directed mutagenesis coupled with EPR spectroscopy. Lastly, we discuss the possible functional consequences that these unique AuxI and AuxII clusters may have in catalysis for PqqE and how these may extend to additional RS enzymes catalyzing the post-translational modification of ribosomally encoded peptides.


Assuntos
Proteínas de Bactérias/química , Endopeptidases/química , Proteínas Ferro-Enxofre/química , Methylobacterium extorquens/química , Cristalografia por Raios X , Espectroscopia de Ressonância de Spin Eletrônica , Modelos Moleculares , Conformação Proteica , Temperatura
6.
J Am Chem Soc ; 140(40): 12947-12963, 2018 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-30222930

RESUMO

Biotin (vitamin B7) is an enzyme cofactor required by organisms from all branches of life but synthesized only in microbes and plants. In the final step of biotin biosynthesis, a radical S-adenosyl-l-methionine (SAM) enzyme, biotin synthase (BioB), converts the substrate dethiobiotin to biotin through the stepwise formation of two C-S bonds. Previous electron paramagnetic resonance (EPR) spectroscopic studies identified a semistable intermediate in the formation of the first C-S bond as 9-mercaptodethiobiotin linked to a paramagnetic [2Fe-2S] cluster through one of its bridging sulfides. Herein, we report orientation-selected pulse EPR spectroscopic results that reveal hyperfine interactions between the [2Fe-2S] cluster and a number of magnetic nuclei (e.g., 57Fe, 15N, 13C, and 2H) introduced in a site-specific manner via biosynthetic methods. Combining these results with quantum chemical modeling gives a structural model of the intermediate showing that C6, the target of the second hydrogen-atom abstraction, is now in close proximity to the nascent thioether sulfur and is ideally positioned for the second C-S bond forming event.


Assuntos
Espectroscopia de Ressonância de Spin Eletrônica , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Sulfurtransferases/metabolismo , Biotina/análogos & derivados , Biotina/química , Biotina/metabolismo , Escherichia coli/química , Proteínas de Escherichia coli/química , Modelos Moleculares , Conformação Proteica , Sulfurtransferases/química
7.
J Am Chem Soc ; 140(40): 12798-12807, 2018 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-30208703

RESUMO

Organisms that perform the de novo biosynthesis of cobalamin (vitamin B12) do so via unique pathways depending on the presence of oxygen in the environment. The anaerobic biosynthesis pathway of 5,6-dimethylbenzimidazole, the so-called "lower ligand" to the cobalt center, has been recently identified. This process begins with the conversion of 5-aminoimidazole ribotide (AIR) to 5-hydroxybenzimidazole (HBI) by the radical S-adenosyl-l-methionine (SAM) enzyme BzaF, also known as HBI synthase. In this work we report the characterization of a radical intermediate in the reaction of BzaF using electron paramagnetic resonance spectroscopy. Using various isotopologues of AIR, we extracted hyperfine parameters for a number of nuclei, allowing us to propose plausible chemical compositions and structures for this intermediate. Specifically, we find that an aminoimidazole radical is formed in close proximity to a fragment of the ribose ring. These findings induce the revision of past proposed mechanisms and illustrate the ability of radical SAM enzymes to tightly control the radical chemistry that they engender.


Assuntos
Proteínas de Bactérias/metabolismo , Benzimidazóis/metabolismo , Vias Biossintéticas , Desulfuromonas/metabolismo , Vitamina B 12/metabolismo , Anaerobiose , Espectroscopia de Ressonância de Spin Eletrônica , S-Adenosilmetionina/metabolismo
8.
J Biol Inorg Chem ; 23(7): 1093-1104, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29968177

RESUMO

The multi-copper oxidase (MCO) MnxG from marine Bacillus bacteria plays an essential role in geochemical cycling of manganese by oxidizing Mn2+(aq) to form manganese oxide minerals at rates that are three to five orders of magnitude faster than abiotic rates. The MCO MnxG protein is isolated as part of a multi-protein complex, denoted as Mnx, which includes one MnxG unit and a hexamer of MnxE3F3 subunit. During the oxidation of Mn2+(aq) catalyzed by the Mnx protein complex, an enzyme-bound Mn(III) species was trapped recently in the presence of pyrophosphate (PP) and analyzed using parallel-mode electron paramagnetic resonance (EPR) spectroscopy. Herein, we provide a full analysis of this enzyme-bound Mn(III) intermediate via temperature dependence studies and spectral simulations. This Mnx-bound Mn(III) species is characterized by a hyperfine-coupling value of A(55Mn) = 4.2 mT (corresponding to 120 MHz) and a negative zero-field splitting (ZFS) value of D = - 2.0 cm-1. These magnetic properties suggest that the Mnx-bound Mn(III) species could be either six-coordinate with a 5B1g ground state or square-pyramidal five-coordinate with a 5B1 ground state. In addition, as a control, Mn(III)PP is also analyzed by parallel-mode EPR spectroscopy. It exhibits distinctly different magnetic properties with a hyperfine-coupling value of A(55Mn) = 4.8 mT (corresponding to 140 MHz) and a negative ZFS value of D = - 2.5 cm-1. The different ZFS values suggest differences in ligand environment of Mnx-bound Mn(III) and aqueous Mn(III)PP species. These studies provide further insights into the mechanism of biological Mn2+(aq) oxidation.


Assuntos
Manganês/metabolismo , Oxirredutases/metabolismo , Bacillus/enzimologia , Espectroscopia de Ressonância de Spin Eletrônica , Manganês/química , Modelos Moleculares , Oxirredutases/química , Oxirredutases/isolamento & purificação , Temperatura
9.
J Am Chem Soc ; 139(33): 11369-11380, 2017 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-28712284

RESUMO

The bacterial protein complex Mnx contains a multicopper oxidase (MCO) MnxG that, unusually, catalyzes the two-electron oxidation of Mn(II) to MnO2 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 Cu2+, the electron acceptor. Indeed the type 1 Cu2+ 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 MnO2 formation from MnO2 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 pKa > 8.6 deprotonation, which is assigned to Mn(II)-bound H2O; 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 pKa 7.6 double deprotonation. It is proposed that turnover produces a Mn(III)(µ-OH)2Mn(III) intermediate that proceeds to the enzyme product, likely Mn(IV)(µ-O)2Mn(IV) or an oligomer, which subsequently nucleates MnO2 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 .


Assuntos
Bacillus/enzimologia , Cobre/metabolismo , Manganês/metabolismo , Oxirredutases/metabolismo , Oxigênio/metabolismo , Bacillus/metabolismo , Catálise , Compostos de Manganês/metabolismo , Oxirredução , Óxidos/metabolismo
10.
J Am Chem Soc ; 139(33): 11381-11391, 2017 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-28712303

RESUMO

The bacterial manganese oxidase MnxG of the Mnx protein complex is unique among multicopper oxidases (MCOs) in carrying out a two-electron metal oxidation, converting Mn(II) to MnO2 nanoparticles. The reaction occurs in two stages: Mn(II) → Mn(III) and Mn(III) → MnO2. In a companion study , we show that the electron transfer from Mn(II) to the low-potential type 1 Cu of MnxG requires an activation step, likely forming a hydroxide bridge at a dinuclear Mn(II) site. Here we study the second oxidation step, using pyrophosphate (PP) as a Mn(III) trap. PP chelates Mn(III) produced by the enzyme and subsequently allows it to become a substrate for the second stage of the reaction. EPR spectroscopy confirms the presence of Mn(III) bound to the enzyme. The Mn(III) oxidation step does not involve direct electron transfer to the enzyme from Mn(III), which is shown by kinetic measurements to be excluded from the Mn(II) binding site. Instead, Mn(III) is proposed to disproportionate at an adjacent polynuclear site, thereby allowing indirect oxidation to Mn(IV) and recycling of Mn(II). PP plays a multifaceted role, slowing the reaction by complexing both Mn(II) and Mn(III) in solution, and also inhibiting catalysis, likely through binding at or near the active site. An overall mechanism for Mnx-catalyzed MnO2 production from Mn(II) is presented.


Assuntos
Bacillus/enzimologia , Compostos de Manganês/metabolismo , Manganês/metabolismo , Óxidos/metabolismo , Oxirredutases/metabolismo , Bacillus/metabolismo , Cobre/metabolismo , Difosfatos/metabolismo , Modelos Moleculares , Nanopartículas/metabolismo , Oxirredução
11.
J Am Chem Soc ; 139(26): 8868-8877, 2017 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-28587464

RESUMO

Manganese-oxide minerals (MnOx) are widely distributed over the Earth's surface, and their geochemical cycling is globally important. A multicopper oxidase (MCO) MnxG protein from marine Bacillus bacteria plays an essential role in producing MnOx minerals by oxidizing Mn2+(aq) at rates that are 3 to 5 orders of magnitude faster than abiotic rates. The MnxG protein is isolated as part of a multiprotein complex denoted as "Mnx" that includes accessory protein subunits MnxE and MnxF, with an estimated stoichiometry of MnxE3F3G and corresponding molecular weight of ≈211 kDa. Herein, we report successful expression and isolation of the MCO MnxG protein without the E3F3 hexamer. This isolated MnxG shows activity for Mn2+(aq) oxidation to form manganese oxides. The complement of paramagnetic Cu(II) ions in the Mnx protein complex was examined by electron paramagnetic resonance (EPR) spectroscopy. Two distinct classes of type 2 Cu sites were detected. One class of Cu(II) site (denoted as T2Cu-A), located in the MnxG subunit, is identified by the magnetic parameters g∥ = 2.320 and A∥ = 510 MHz. The other class of Cu(II) sites (denoted as T2Cu-B) is characterized by g∥ = 2.210 and A∥ = 615 MHz and resides in the putative hexameric MnxE3F3 subunit. These different magnetic properties correlate with the differences in the reduction potentials of the respective Cu(II) centers. These studies provide new insights into the molecular mechanism of manganese biomineralization.


Assuntos
Cobre/química , Compostos de Manganês/química , Manganês/química , Óxidos/química , Bacillus/enzimologia , Sítios de Ligação , Ferro/química , Compostos de Manganês/isolamento & purificação , Compostos de Manganês/metabolismo , Oxirredução , Óxidos/isolamento & purificação , Óxidos/metabolismo
12.
Inorg Chem ; 56(23): 14596-14604, 2017 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-29130691

RESUMO

A simple exchange reaction between [AriPr4Sn(µ-Cl)]2 (1) and sodium azide afforded the doubly bridged Sn(II) azide, [AriPr4Sn(µ-N3)]2 (2) (AriPr4 = C6H3-2,6(C6H3-2,6-iPr2)2) in 85% yield. Photolysis of a diethyl ether solution of 2 for ca. 16 h yielded an azepinyl-substituted insertion product, [C6H3-2-(C6H3-2,6-iPr2)-6-(C6H3N-3,7-iPr2)Sn]2 (3). The reaction of the Lewis acid, B(C6F5)3 (BCF), or the Lewis base, pyridine, with 2 dissociates the dimer to afford the corresponding complexed monomeric Sn(II) azide, AriPr4SnN3BCF (4) in which BCF coordinates the α-nitrogen, or AriPr4Sn(pyridine)N3 (6) in which pyridine coordinates to the tin atom. Photolysis of 4 in diethyl ether for 12 h results in the insertion of the α-nitrogen of the azide group into one of the B-C bonds of the BCF acceptor to yield the tin(II) amide, AriPr4SnN(C6F5)B(C6F5)2 (5). In contrast, photolysis of 6 for over 36 h afforded no apparent reaction. A highly reactive Sn nitride intermediate, AriPr4Sn≡N, is proposed as part of the mechanistic pathway for the formation of 3 and 5 as a result of trapping the tin-centered radical isomers. This was effected by immediate freezing the samples of 2 or 4 after ca. 30 min of UV photolysis and recording their electron paramagnetic resonance spectra. These exhibited a rhombic g tensor of [g1, g2, g3] = [2.029, 1.978, 1.933]. This radical intermediate could be related to the valence isomers of the nitride [-SnIV≡N] intermediate, in isomeric equilibrium with the nitrene [-SnII-N] and nitridyl [-SnIII═N·] forms, but with the spin density on the nitrogen being quenched, possibly by the H atom abstraction to form an S = 1/2 species of formula -Sn·â•N(H).

13.
J Am Chem Soc ; 138(25): 7951-64, 2016 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-27276098

RESUMO

Fluorinated tyrosines (FnY's, n = 2 and 3) have been site-specifically incorporated into E. coli class Ia ribonucleotide reductase (RNR) using the recently evolved M. jannaschii Y-tRNA synthetase/tRNA pair. Class Ia RNRs require four redox active Y's, a stable Y radical (Y·) in the ß subunit (position 122 in E. coli), and three transiently oxidized Y's (356 in ß and 731 and 730 in α) to initiate the radical-dependent nucleotide reduction process. FnY (3,5; 2,3; 2,3,5; and 2,3,6) incorporation in place of Y122-ß and the X-ray structures of each resulting ß with a diferric cluster are reported and compared with wt-ß2 crystallized under the same conditions. The essential diferric-FnY· cofactor is self-assembled from apo FnY-ß2, Fe(2+), and O2 to produce ∼1 Y·/ß2 and ∼3 Fe(3+)/ß2. The FnY· are stable and active in nucleotide reduction with activities that vary from 5% to 85% that of wt-ß2. Each FnY·-ß2 has been characterized by 9 and 130 GHz electron paramagnetic resonance and high-field electron nuclear double resonance spectroscopies. The hyperfine interactions associated with the (19)F nucleus provide unique signatures of each FnY· that are readily distinguishable from unlabeled Y·'s. The variability of the abiotic FnY pKa's (6.4 to 7.8) and reduction potentials (-30 to +130 mV relative to Y at pH 7.5) provide probes of enzymatic reactions proposed to involve Y·'s in catalysis and to investigate the importance and identity of hopping Y·'s within redox active proteins proposed to protect them from uncoupled radical chemistry.


Assuntos
Proteínas de Escherichia coli/química , Escherichia coli/enzimologia , Exorribonucleases/química , Flúor/química , Methanocaldococcus/enzimologia , Ribonucleotídeo Redutases/química , Tirosina/química , Aminoacil-tRNA Sintetases/metabolismo , Sítios de Ligação , Catálise , Simulação por Computador , Cristalização , Cristalografia por Raios X , Espectroscopia de Ressonância de Spin Eletrônica , Escherichia coli/genética , Radicais Livres/química , Ligação de Hidrogênio , Cinética , Espectroscopia de Ressonância Magnética , Methanocaldococcus/genética , Modelos Moleculares , Oxirredução , Oxigênio/química , Fosforilação , RNA de Transferência/química , Temperatura
14.
Inorg Chem ; 55(24): 12728-12736, 2016 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-27989208

RESUMO

A variety of metal oxides can catalyze the oxidation of water to molecular oxygen when polarized by a sufficiently high electrochemical potential. Minimizing the overpotential and increasing the rate of the oxygen-evolving reaction (OER) are key goals in making such materials a component of viable energy storage devices. However, the structural factors that imbue the metal oxides with their catalytic power are difficult to assess as these solids contain many distinct metal-ion sites, have a varying amount of defect sites within the lattice, and can be composed of multiple phases. In the present study, we determined the magnetic properties for a series of dimeric cobalt complexes in which the two metal centers are bridged by a dioxygen moiety. Our spectroscopically validated electronic structure description indicates that each species is best described as two Co(III) ions that are bound to a µ-η1η1 superoxide ligand. Intriguingly, we found evidence that the two compounds that possess oxygen-evolving activity coordinate the superoxide ion in an unusual, nonplanar fashion. It appears as if the intermediately long Co···Co distance of 3.9 Å is responsible for the unusual superoxide binding geometry. This structural factor may be an important element in the design of solid-state OER catalysts.

15.
Angew Chem Int Ed Engl ; 55(35): 10444-7, 2016 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-27416899

RESUMO

The synthesis of the first linear coordinated Cu(II) complex Cu{N(SiMe3 )Dipp}2 (1 Dipp=C6 H5 -2,6Pr(i) 2 ) and its Cu(I) counterpart [Cu{N(SiMe3 )Dipp}2 ](-) (2) is described. The formation of 1 proceeds through a dispersion force-driven disproportionation, and is the reaction product of a Cu(I) halide and LiN(SiMe3 )Dipp in a non-donor solvent. The synthesis of 2 is accomplished by preventing the disproportionation into 1 by using the complexing agent 15-crown-5. EPR spectroscopy of 1 provides the first detailed study of a two-coordinate transition-metal complex indicating strong covalency in the Cu-N bonds.

16.
J Am Chem Soc ; 137(27): 8829-37, 2015 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-26083545

RESUMO

High-resolution X-ray structures of photosystem II reveal several potential substrate binding sites at the water-oxidizing/oxygen-evolving 4MnCa cluster. Aspartate-61 of the D1 protein hydrogen bonds with one such water (W1), which is bound to the dangler Mn4A of the oxygen-evolving complex. Comparison of pulse EPR spectra of (14)NH3 and (15)NH3 bound to wild-type Synechocystis PSII and a D1-D61A mutant lacking this hydrogen-bonding interaction demonstrates that ammonia binds as a terminal NH3 at this dangler Mn4A site and not as a partially deprotonated bridge between two metal centers. The implications of this finding on identifying the binding sites of the substrate and the subsequent mechanism of dioxygen formation are discussed.


Assuntos
Amônia/metabolismo , Proteínas de Bactérias/metabolismo , Manganês/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Synechocystis/metabolismo , Proteínas de Bactérias/química , Sítios de Ligação , Cristalografia por Raios X , Espectroscopia de Ressonância de Spin Eletrônica , Ligação de Hidrogênio , Manganês/química , Modelos Moleculares , Complexo de Proteína do Fotossistema II/química , Conformação Proteica , Synechocystis/química
17.
J Am Chem Soc ; 137(10): 3525-32, 2015 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-25686300

RESUMO

Reductive dehalogenases play a critical role in the microbial detoxification of aquifers contaminated with chloroethenes and chlorethanes by catalyzing the reductive elimination of a halogen. We report here the first heterologous production of vinyl chloride reductase VcrA from Dehalococcoides mccartyi strain VS. Heterologously expressed VcrA was reconstituted to its active form by addition of hydroxocobalamin/adenosylcobalamin, Fe(3+), and sulfide in the presence of mercaptoethanol. The kinetic properties of reconstituted VcrA catalyzing vinyl chloride reduction with Ti(III)-citrate as reductant and methyl viologen as mediator were similar to those obtained previously for VcrA as isolated from D. mccartyi strain VS. VcrA was also found to catalyze a novel reaction, the environmentally important dihaloelimination of 1,2-dichloroethane to ethene. Electron paramagnetic resonance (EPR) spectroscopic studies with reconstituted VcrA in the presence of mercaptoethanol revealed the presence of Cob(II)alamin. Addition of Ti(III)-citrate resulted in the appearance of a new signal characteristic of a reduced [4Fe-4S] cluster and the disappearance of the Cob(II)alamin signal. UV-vis absorption spectroscopy of Ti(III)citrate-treated samples revealed the formation of two new absorption maxima characteristic of Cob(I)alamin. No evidence for the presence of a [3Fe-4S] cluster was found. We postulate that during the reaction cycle of VcrA, a reduced [4Fe-4S] cluster reduces Co(II) to Co(I) of the enzyme-bound cobalamin. Vinyl chloride reduction to ethene would be initiated when Cob(I)alamin transfers an electron to the substrate, generating a vinyl radical as a potential reaction intermediate.


Assuntos
Chloroflexi/enzimologia , Hidrolases/genética , Hidrolases/metabolismo , Cloreto de Vinil/metabolismo , Chloroflexi/genética , Espectroscopia de Ressonância de Spin Eletrônica , Escherichia coli/genética , Expressão Gênica , Halogenação , Hidrolases/química , Oxirredução , Especificidade por Substrato
18.
J Am Chem Soc ; 137(33): 10563-75, 2015 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-26244911

RESUMO

The dynamics of manganese solid formation (as MnOx) by the multicopper oxidase (MCO)-containing Mnx protein complex were examined by electron paramagnetic resonance (EPR) spectroscopy. Continuous-wave (CW) EPR spectra of samples of Mnx, prepared in atmosphere and then reacted with Mn(II) for times ranging from 7 to 600 s, indicate rapid oxidation of the substrate manganese (with two-phase pseudo-first-order kinetics modeled using rate coefficients of: k(1obs) = 0.205 ± 0.001 s(-1) and k(2obs) = 0.019 ± 0.001 s(-1)). This process occurs on approximately the same time scale as in vitro solid MnOx formation when there is a large excess of Mn(II). We also found CW and pulse EPR spectroscopic evidence for at least three classes of Mn(II)-containing species in the reaction mixtures: (i) aqueous Mn(II), (ii) a specifically bound mononuclear Mn(II) ion coordinated to the Mnx complex by one nitrogenous ligand, and (iii) a weakly exchange-coupled dimeric Mn(II) species. These findings provide new insights into the molecular mechanism of manganese mineralization.


Assuntos
Manganês/metabolismo , Oxirredutases/metabolismo , Animais , Bacillus/enzimologia , Bovinos , Dimerização , Espectroscopia de Ressonância de Spin Eletrônica , Cinética , Manganês/química , Oxirredução , Óxidos/química , Óxidos/metabolismo , Oxirredutases/química , Ligação Proteica
19.
J Am Chem Soc ; 137(8): 3004-16, 2015 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-25597447

RESUMO

The antimicrobial protein calprotectin (CP), a hetero-oligomer of the S100 family members S100A8 and S100A9, is the only identified mammalian Mn(II)-sequestering protein. Human CP uses Ca(II) ions to tune its Mn(II) affinity at a biologically unprecedented hexahistidine site that forms at the S100A8/S100A9 interface, and the molecular basis for this phenomenon requires elucidation. Herein, we investigate the remarkable Mn(II) coordination chemistry of human CP using X-ray crystallography as well as continuous-wave (CW) and pulse electron paramagnetic resonance (EPR) spectroscopies. An X-ray crystallographic structure of Mn(II)-CP containing one Mn(II), two Ca(II), and two Na(I) ions per CP heterodimer is reported. The CW EPR spectrum of Ca(II)- and Mn(II)-bound CP prepared with a 10:0.9:1 Ca(II):Mn(II):CP ratio is characterized by an unusually low zero-field splitting of 485 MHz (E/D = 0.30) for the S = 5/2 Mn(II) ion, consistent with the high symmetry of the His6 binding site observed crystallographically. Results from electron spin-echo envelope modulation and electron-nuclear double resonance experiments reveal that the six Mn(II)-coordinating histidine residues of Ca(II)- and Mn(II)-bound CP are spectroscopically equivalent. The observed (15)N (I = 1/2) hyperfine couplings (A) arise from two distinct classes of nitrogen atoms: the coordinating ε-nitrogen of the imidazole ring of each histidine ligand (A = [3.45, 3.71, 5.91] MHz) and the distal δ-nitrogen (A = [0.11, 0.18, 0.42] MHz). In the absence of Ca(II), the binding affinity of CP for Mn(II) drops by two to three orders of magnitude and coincides with Mn(II) binding at the His6 site as well as other sites. This study demonstrates the role of Ca(II) in enabling high-affinity and specific binding of Mn(II) to the His6 site of human calprotectin.


Assuntos
Cálcio/metabolismo , Complexo Antígeno L1 Leucocitário/química , Complexo Antígeno L1 Leucocitário/metabolismo , Manganês/metabolismo , Cristalografia por Raios X , Espectroscopia de Ressonância de Spin Eletrônica , Histidina/química , Humanos , Imidazóis/química , Complexo Antígeno L1 Leucocitário/genética , Modelos Moleculares , Mutação , Oligopeptídeos/química , Ligação Proteica , Estrutura Secundária de Proteína , Prótons
20.
Photosynth Res ; 124(1): 7-18, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25663565

RESUMO

In exchange-coupled mixed-valence spin systems, the magnitude and sign of the effective ligand hyperfine interaction (HFI) can be useful in determining the formal oxidation state of the coordinating metal ion, as well as provide information about the coordination geometry. This is due to the fact that the observed ligand HFI is a function of the projection factor (Clebsch-Gordon coefficient) that maps the site spin value S i of the local paramagnetic center onto the total spin of the exchange-coupled system, S T. Recently, this relationship has been successfully exploited in identifying the oxidation state of the Mn ion coordinated by the sole nitrogenous ligand to the oxygen-evolving complex in certain states of photosystem II. The origin and evolution of these efforts is described.


Assuntos
Metais/metabolismo , Espectroscopia de Ressonância de Spin Eletrônica , Íons , Ligantes , Oxirredução , Complexo de Proteína do Fotossistema II/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA