Chloride- and Hydrosulfide-Bound 2Fe Complexes as Models of the Oxygen-Stable State of [FeFe] Hydrogenase.

[FeFe] hydrogenases demonstrate remarkable catalytic efficiency in hydrogen evolution and oxidation processes. However, susceptibility of these enzymes to oxygen-induced degradation impedes their practical deployment in hydrogen-production devices and fuel cells. Recent investigations into the oxygen-stable (Hinact) state of the H-cluster revealed its inherent capacity to resist oxygen degradation. Herein, we present findings on Cl- and SH-bound [2Fe-2S] complexes, bearing relevance to the oxygen-stable state within a biological context. A characteristic attribute of these complexes is the terminal Cl-/SH- ligation to the iron center bearing the CO bridge. Structural analysis of the t-Cl demonstrates a striking resemblance to the Hinact state of DdHydAB and CbA5H. The t-Cl/t-SH exhibit reversible oxidation, with both redox species, electronically, being the first biomimetic analogs to the Htrans and Hinact states. These complexes exhibit notable resistance against oxygen-induced decomposition, supporting the potential oxygen-resistant nature of the Htrans and Hinact states. The swift reductive release of the Cl-/SH-group demonstrates its labile and kinetically controlled binding. The findings garnered from these investigations offer valuable insights into properties of the enzymatic O2-stable state, and key factors governing deactivation and reactivation conversion. This work contributes to the advancement of bio-inspired molecular catalysts and the integration of enzymes and artificial catalysts into H2-evolution devices and fuel-cell applications.

Targeting the ALS/FTD-associated A-DNA kink with anthracene-based metal complex causes DNA backbone straightening and groove contraction.

The use of a small molecule compound to reduce toxic repeat RNA transcripts or their translated aberrant proteins to target repeat-expanded RNA/DNA with a G4C2 motif is a promising strategy to treat C9orf72-linked disorders. In this study, the crystal structures of DNA and RNA-DNA hybrid duplexes with the -GGGCCG- region as a G4C2 repeat motif were solved. Unusual groove widening and sharper bending of the G4C2 DNA duplex A-DNA conformation with B-form characteristics inside was observed. The G4C2 RNA-DNA hybrid duplex adopts a more typical rigid A form structure. Detailed structural analysis revealed that the G4C2 repeat motif of the DNA duplex exhibits a hydration shell and greater flexibility and serves as a 'hot-spot' for binding of the anthracene-based nickel complex, NiII(Chro)2 (Chro = Chromomycin A3). In addition to the original GGCC recognition site, NiII(Chro)2 has extended specificity and binds the flanked G:C base pairs of the GGCC core, resulting in minor groove contraction and straightening of the DNA backbone. We have also shown that Chro-metal complexes inhibit neuronal toxicity and suppresses locomotor deficits in a Drosophila model of C9orf72-associated ALS. The approach represents a new direction for drug discovery against ALS and FTD diseases by targeting G4C2 repeat motif DNA.

Crystal Structure Analysis of the Repair of Iron Centers Protein YtfE and Its Interaction with NO.

Molecular mechanisms underlying the repair of nitrosylated [Fe-S] clusters by the microbial protein YtfE remain poorly understood. The X-ray crystal structure of YtfE, in combination with EPR, magnetic circular dichroism (MCD), UV, and (17) O-labeling electron spin echo envelope modulation measurements, show that each iron of the oxo-bridged Fe(II) -Fe(III) diiron core is coordinatively unsaturated with each iron bound to two bridging carboxylates and two terminal histidines in addition to an oxo-bridge. Structural analysis reveals that there are two solvent-accessible tunnels, both of which converge to the diiron center and are critical for capturing substrates. The reactivity of the reduced-form Fe(II) -Fe(II) YtfE toward nitric oxide demonstrates that the prerequisite for N2 O production requires the two iron sites to be nitrosylated simultaneously. Specifically, the nitrosylation of the two iron sites prior to their reductive coupling to produce N2 O is cooperative. This result suggests that, in addition to any repair of iron centers (RIC) activity, YtfE acts as an NO-trapping scavenger to promote the NO to N2 O transformation under low NO flux, which precedes nitrosative stress.

The effect of anions in the synthesis and structure of pyrazolylamidino copper(II) complexes.

Six new pyrazolylamidino Cu(II) complexes are synthesized directly from the reactions of Cu(X)2 salts (X = ClO4-, BF4-, or Cl-) and pyrazole (pzH) in nitrile solution (RCN, R = Me or Et) at 298 K via the metal-mediated coupling of RCN with pzH: [Cu(HNC(R)pz)2(X)2] (X = ClO4- or BF4-, R = Me, 1 or 7 and Et, 2 or 8, respectively) and dichloro Cu(II) complexes [Cu2Cl2(µ-Cl)2(HNC(Me)pz)2] (3) and [CuCl2(HNC(Et)pz)] (4). Four more new complexes, [Cu2(µ-Cl)2(HNC(Me)pz)2(pzH)2][X]2 (X = ClO4-, 5 and BF4-, 9) and [Cu2(µ-Cl)2(HNC(Et)pz)2(pzH)2(X)2] (X = ClO4-, 6 and BF4-, 10), are obtained indirectly from the anion substitution reaction with Cl- ions in 1 and 7, and 2 and 8, respectively. All complexes are characterized by EA, FTIR, UV-vis and EPR spectroscopy and X-ray crystallographic analyses. HNC(Et)pz or pzH is unobserved in both the nitrile-exchange reaction of 2 to d6-1 and the anion-substitution reaction of 2 to d6-5 in the CD3CN solution. The 1H NMR results reveal that the pzH-RCN coupling is intramolecular and reversible on a Cu(II) center. The crystal structures of these complexes show diverse supramolecular assemblies through imino NH⋯anion hydrogen bonds and pyrazolylamidino pz-pz (π⋯π) and pz-Cu(II) (π⋯metal) interactions. EPR results suggest weak magnetic couplings between Cu(II) centers in the polynuclear Cu(II) complexes. The yield and rate of the formation of 1 are higher in the reaction of Cu(ClO4)2 with a 4-fold molar excess of pzH compared with a 2-fold excess, indicating that [Cu(pzH)4]2+ is the more active species for pzH-RCN coupling. The highest rate for the formation of 1 is achieved when [Cu(pzH)4(ClO4)2] is used in MeCN solution. Thus, a plausible synthetic path for synthesizing pyrazolylamidino Cu(II) complexes is established. An intermediate species, [Cu(HNC(Me)pz)2(pzH)2][ClO4]2 (1a), is proposed for the synthetic process based on spectroscopic studies and DFT calculations. The reaction of [Cu(pzH)4X2] (X = ClO4-, Cl-, NO3-, or BF4-) in MeCN solution suggests that the lability of coordinated anions upon nitrile substitution affects the rate of the formation of bis-pyrazolylamidino Cu(II) complexes.

N,N'-Bis[(1H-imidazol-1-yl)meth-yl]-2,2'-(disulfanedi-yl)dianiline.

The symmetrical title compound, C(20)H(20)N(6)S(2), contains a disulfide bond of 2.0884 (6) Å. The C-S-S-C torsion angle is -59.57 (7)°. In the crystal, classical N-H⋯N and non-classical C-H⋯N hydrogen bonds link the compounds into chains along the a axis.

N-[2-(Methyl-sulfan-yl)phen-yl]-2-sulfanylbenzamide.

In the title compound, C(14)H(13)NOS(2), the S atom with the methyl group is involved in an intra-molecular hydrogen bond with the amido H atom. In the crystal, the sulfanyl H atoms form inter-molecular hydrogen bonds with the O atoms, connecting the mol-ecules into zigzag chains along the c axis. The two aromatic rings exhibit a small interplanar angle of 16.03 (9)°.

Catalytic O2 activation toward oxidative N-S bond formation by a thiolato Fe(III) complex.

Compounds with the benzisothiazol-3-one (BIT) skeleton perform excellently in the pharmaceutical field, although current synthetic methods remain limited in terms of synthetic efficiency. Herein, we report the catalytic intramolecular N-S bond formation for BITs from easily prepared disulfide precursors by an Fe(III) dithiolate through O2 activation at 298 K. Interestingly, the catalytic performance is enhanced by substituting O2 with a milder O-donor oxidant, ONMe3. Catalytic oxygenation of PPh3 to OPPh3 can also proceed under similar conditions. In addition, the first anionic monosulfenato Fe(III) species, Fe(III)-S(O)R, is isolated with structural characterization from the reaction of Fe(III) thiolate and ONMe3.

trans-Diacetonitrile-tetra-kis(1H-pyrazole-κN)nickel(II) dinitrate.

In the title complex, [Ni(CH(3)CN)(2)(C(3)H(4)N(2))(4)](NO(3))(2), the cation lies on an inversion center and adopts an octa-hedral coordination geometry about the Ni atom. The two acetonitrile ligands are in a trans conformation. N-H⋯O hydrogen bonds between cations and anions link the complex mol-ecules into one-dimensional chains running parallel to [100].

Dioxygen activation by a dinuclear thiolate-ligated Fe(ii) complex.

Dioxygen activation by FeII thiolate complexes is relatively rare in biological and chemical systems because the sulfur site is at least as vulnerable as the iron site to oxidative modification. O2 activation by FeII-SR complexes with thiolate bound trans to the O2 binding site generally affords the FeIV[double bond, length as m-dash]O intermediate and oxidized thiolate. On the other hand, O2 activation by Fe(ii)-SR complexes with thiolate bound cis to the O2 binding site generates FeIII-O-FeIII or S-oxygenated complexes. The postulated FeIV[double bond, length as m-dash]O intermediate has only been identified in isopenicillin N synthase recently. We demonstrated here that O2 activation by a dinuclear FeII thiolate-rich complex produces a mononuclear FeIII complex and water with a supply of electron donors. The thiolate is bound cis to the postulated dioxygen binding site, and no FeIII-O-FeIII or S-oxygenated complex was observed. Although we have not detected the transient intermediate by spectroscopic measurements, the FeIV[double bond, length as m-dash]O intermediate is suggested to exist by theoretical calculation, and P-oxidation and hydride-transfer experiments. In addition, an unprecedented FeIII-O2-FeIII complex supported by thiolates was observed during the reaction by using a coldspray ionization time-of-flight mass (CSI-TOF MS) instrument. This is also supported by low-temperature UV-vis measurements. The intramolecular NHO[double bond, length as m-dash]FeIV hydrogen bonding, calculated by DFT, probably fine tunes the O2-activation process for intramolecular hydrogen abstraction, avoiding the S-oxygenation at cis-thiolate.

Assembly and disassembly of a Zn10 high-nuclearity circular helicate.

A nano-scale decanuclear Zn(II) circular helicate is synthesized without the aid of counteranions during the assembly process, and can be totally disassembled into its reactants by specific anions.

Selective capture of volatile iodine using amorphous molecular organic solids.

A simple shape-persistent organic molecular container is capable of selective absorption and storage of I(2(g)) over water vapor and NO gas even in its amorphous solid state. In addition, the strongly associated I(2) can be efficiently released from the charged container in organic solvents under ambient conditions.

Selective encapsulation of volatile and reactive methyl iodide.

A simple organic molecular container can selectively encapsulate the volatile and highly reactive MeI through hydrogen-bonding interactions in solution. The remarkable encapsulation of MeI without self-methylation of the container appears to be determined by the complementary binding sites and the rigidity of the hydrogen-bonding array constrained by the molecular framework.

Spectroscopic and kinetic studies of the reaction of bromopropanesulfonate with methyl-coenzyme M reductase.

Methyl-coenzyme M reductase (MCR) catalyzes the final step of methanogenesis in which coenzyme B and methyl-coenzyme M are converted to methane and the heterodisulfide, CoMS-SCoB. MCR also appears to initiate anaerobic methane oxidation (reverse methanogenesis). At the active site of MCR is coenzyme F430, a nickel tetrapyrrole. This paper describes the reaction of the active MCR(red1) state with the potent inhibitor, 3-bromopropanesulfonate (BPS; I50 = 50 nM) by UV-visible and EPR spectroscopy and by steady-state and rapid kinetics. BPS was shown to be an alternative substrate of MCR in an ionic reaction that is coenzyme B-independent and leads to debromination of BPS and formation of a distinct state ("MCR(PS)") with an EPR signal that was assigned to a Ni(III)-propylsulfonate species (Hinderberger, D., Piskorski, R. P., Goenrich, M., Thauer, R. K., Schweiger, A., Harmer, J., and Jaun, B. (2006) Angew. Chem. Int. Ed. Engl. 45, 3602-3607). A similar EPR signal was generated by reacting MCR(red1) with several halogenated sulfonate and carboxylate substrates. In rapid chemical quench experiments, the propylsulfonate ligand was identified by NMR spectroscopy and high performance liquid chromatography as propanesulfonic acid after protonolysis of the MCR(PS) complex. Propanesulfonate formation was also observed in steady-state reactions in the presence of Ti(III) citrate. Reaction of the alkylnickel intermediate with thiols regenerates the active MCR(red1) state and eliminates the propylsulfonate group, presumably as the thioether. MCR(PS) is catalytically competent in both the generation of propanesulfonate and reformation of MCR(red1). These results provide evidence for the intermediacy of an alkylnickel species in the final step in anaerobic methane oxidation and in the initial step of methanogenesis.

The P174L mutation in human Sco1 severely compromises Cox17-dependent metallation but does not impair copper binding.

Sco1 is a metallochaperone that is required for copper delivery to the Cu(A) site in the CoxII subunit of cytochrome c oxidase. The only known missense mutation in human Sco1, a P174L substitution in the copper-binding domain, is associated with a fatal neonatal hepatopathy; however, the molecular basis for dysfunction of the protein is unknown. Immortalized fibroblasts from a SCO1 patient show a severe deficiency in cytochrome c oxidase activity that was partially rescued by overexpression of P174L Sco1. The mutant protein retained the ability to bind Cu(I) and Cu(II) normally when expressed in bacteria, but Cox17-mediated copper transfer was severely compromised both in vitro and in a yeast cytoplasmic assay. The corresponding P153L substitution in yeast Sco1 was impaired in suppressing the phenotype of cells harboring the weakly functional C57Y allele of Cox17; however, it was functional in sco1delta yeast when the wild-type COX17 gene was present. Pulse-chase labeling of mitochondrial translation products in SCO1 patient fibroblasts showed no change in the rate of CoxII translation, but there was a specific and rapid turnover of CoxII protein in the chase. These data indicate that the P174L mutation attenuates a transient interaction with Cox17 that is necessary for copper transfer. They further suggest that defective Cox17-mediated copper metallation of Sco1, as well as the subsequent failure of Cu(A) site maturation, is the basis for the inefficient assembly of the cytochrome c oxidase complex in SCO1 patients.

Spectroscopic and computational studies of reduction of the metal versus the tetrapyrrole ring of coenzyme F430 from methyl-coenzyme M reductase.

Methyl-coenzyme M reductase (MCR) catalyzes the final step in methane biosynthesis by methanogenic archaea and contains a redox-active nickel tetrahydrocorphin, coenzyme F430, at its active site. Spectroscopic and computational methods have been used to study a novel form of the coenzyme, called F330, which is obtained by reducing F430 with sodium borohydride (NaBH4). F330 exhibits a prominent absorption peak at 330 nm, which is blue shifted by 100 nm relative to F430. Mass spectrometric studies demonstrate that the tetrapyrrole ring in F330 has undergone reduction, on the basis of the incorporation of protium (or deuterium), upon treatment of F430 with NaBH4 (or NaBD4). One- and two-dimensional NMR studies show that the site of reduction is the exocyclic ketone group of the tetrahydrocorphin. Resonance Raman studies indicate that elimination of this pi-bond increases the overall pi-bond order in the conjugative framework. X-ray absorption, magnetic circular dichroism, and computational results show that F330 contains low-spin Ni(II). Thus, conversion of F430 to F330 reduces the hydrocorphin ring but not the metal. Conversely, reduction of F430 with Ti(III) citrate to generate F380 (corresponding to the active MCR(red1) state) reduces the Ni(II) to Ni(I) but does not reduce the tetrapyrrole ring system, which is consistent with other studies [Piskorski, R., and Jaun, B. (2003) J. Am. Chem. Soc. 125, 13120-13125; Craft, J. L., et al. (2004) J. Biol. Inorg. Chem. 9, 77-89]. The distinct origins of the absorption band shifts associated with the formation of F330 and F380 are discussed within the framework of our computational results. These studies on the nature of the product(s) of reduction of F430 are of interest in the context of the mechanism of methane formation by MCR and in relation to the chemistry of hydroporphinoid systems in general. The spectroscopic and time-dependent DFT calculations add important insight into the electronic structure of the nickel hydrocorphinate in its Ni(II) and Ni(I) valence states.

Functional analysis of the domains in Cox11.

Cox11 is an intrinsic mitochondrial membrane protein essential for the assembly of an active cytochrome c oxidase complex. Cox11 is tethered to the mitochondrial inner membrane by a single transmembrane helix. Domain mapping was carried out to determine the functional segments of the Cox11 protein. The C-terminal 189 residue Cu(I)-binding domain is shown to be exposed within the mitochondrial intermembrane space. This orientation was demonstrated by the proteolytic susceptibility of a C-terminal Myc epitope tag in mitoplasts but not intact mitochondria. Fusion of the N terminus of Cox11 to the matrix ribosomal protein Rsm22 results in a functional protein capable of suppressing the respiratory defect of both Deltacox11 cells and Deltarsm22 cells. The functionality of the fusion protein suggests that the Cox11 N terminus projects into the matrix. The fusion of the C-terminal segment of Cox11 to Rsm22 resembles a naturally occurring fusion of Cox11 in Schizosaccharomyces pombe to a sequence homologous to the Saccharomyces cerevisiae Rsm22. Studies on a series of SCO1/COX11 chimeras reveal that the matrix domain of Cox11 lacks a specific function, whereas the Cu(I) binding/donating function requires the yeast Cox11 sequence. The Cu(I)-binding domain from human Cox11 cannot functionally replace the yeast sequence. The copper domain of Cox11 may be an important docking motif for Cox1 or a Cox1-associated protein.

Human Sco1 and Sco2 function as copper-binding proteins.

The function of human Sco1 and Sco2 is shown to be dependent on copper ion binding. Expression of soluble domains of human Sco1 and Sco2 either in bacteria or the yeast cytoplasm resulted in the recovery of copper-containing proteins. The metallation of human Sco1, but not Sco2, when expressed in the yeast cytoplasm is dependent on the co-expression of human Cox17. Two conserved cysteines and a histidyl residue, known to be important for both copper binding and in vivo function in yeast Sco1, are also critical for in vivo function of human Sco1 and Sco2. Human and yeast Sco proteins can bind either a single Cu(I) or Cu(II) ion. The Cu(II) site yields S-Cu(II) charge transfer transitions that are not bleached by weak reductants or chelators. The Cu(I) site exhibits trigonal geometry, whereas the Cu(II) site resembles a type II Cu(II) site with a higher coordination number. To identify additional potential ligands for the Cu(II) site, a series of mutant proteins with substitutions in conserved residues in the vicinity of the Cu(I) site were examined. Mutation of several conserved carboxylates did not alter either in vivo function or the presence of the Cu(II) chromophore. In contrast, replacement of Asp238 in human or yeast Sco1 abrogated the Cu(II) visible transitions and in yeast Sco1 attenuated Cu(II), but not Cu(I), binding. Both the mutant yeast and human proteins were nonfunctional, suggesting the importance of this aspartate for normal function. Taken together, these data suggest that both Cu(I) and Cu(II) binding are critical for normal Sco function.

Rapid ligand exchange in the MCRred1 form of methyl-coenzyme M reductase.

Methyl-coenzyme M reductase (MCR) from Methanothermobacter marburgensis (Mtm), catalyses the final step in methane synthesis in all methanogenic organisms. Methane is produced by coenzyme B-dependent two-electron reduction of methyl-coenzyme M. At the active site of MCR is the corphin cofactor F(430), which provides four-coordination through the pyrrole nitrogens to a central Ni ion in all states of the enzyme. The important MCRox1 ("ready") and MCRred1 ("active") states contain six-coordinate Ni(I) and differ in their upper axial ligands; furthermore, red1 appears to be two-electrons more reduced than in ox1 and other Ni(II) states that have been studied. On the basis of the reactivity of MCRred1 and MCRox1 with a substrate analogue and inhibitor (3-bromopropanesulfonate) and other small molecules (chloroform, dichloromethane, mercaptoethanol, and nitric oxide), we present evidence that the six-coordinate Ni(I) centers in the MCRred1 and MCRox1 states exhibit markedly different inherent reactivities. MCRred1 reacts faster with chloroform (2100-fold or 35000-fold when corrected for temperature effects), nitric oxide (90-fold), and 3-bromopropanesulfonate (10(6)-fold) than MCRox1. MCRred1 reacts with chloroform and dichloromethane and, like F(430), can catalyze dehalogenation reactions and produce lower halogenated products. We conclude that the enhanced reactivity of MCRred1 is due to the replacement of a relatively exchange-inert thiol ligand in MCRox1 with a weakly coordinating upper axial ligand in red1 that can be easily replaced by incoming ligands.

Nickel oxidation states of F(430) cofactor in methyl-coenzyme M reductase.

Magnetic circular dichroism (MCD) spectroscopy and variable-temperature variable-field MCD are used in combination with density functional theory (DFT) and time-dependent DFT (TD-DFT) calculations to characterize the so-called ox1-silent, red1, and ox1 forms of the Ni-containing cofactor F430 in methyl-coenzyme M reductase (MCR). Previous studies concluded that the ox1 state, which is the precursor of the key reactive red1 state of MCR, is a Ni(I) species that derives from one-electron reduction of the Ni(II)-containing ox1-silent state. However, our absorption and MCD data provide compelling evidence that ox1 is actually a Ni(II) species. In support of this proposal, our DFT and TD-DFT calculations indicate that addition of an electron to the ox1-silent state leads to formation of a hydrocorphin anion radical rather than a Ni(I) center. These results and biochemical evidence suggest that ox1 is more oxidized than red1, which prompted us to test a new model for ox1 in which the ox1-silent species is oxidized by one electron to form a thiyl radical derived from coenzyme M that couples antiferromagnetically to the Ni(II) ion. This alternative ox1 model, formally corresponding to a Ni(III)/thiolate resonance form but with predicted S = 1/2 EPR parameters reminiscent of a Ni(I) (3dx2-y2)1 species, rationalizes the requirement for reduction of ox1 to yield the red1 species and the seemingly incongruent EPR and electronic spectra of the ox1 state.

Spectroscopic and computational characterization of the nickel-containing F430 cofactor of methyl-coenzyme M reductase.

Methyl-coenzyme M reductase (MCR) catalyzes the terminal reaction in methanogenesis, the formation of methane from methyl-coenzyme M and coenzyme B. The active site of MCR binds the prosthetic group F(430), a unique nickel hydrocorphin cofactor. Here, spectroscopy and computations are employed in developing detailed electronic descriptions of the Ni(II) and Ni(I) forms of the free cofactor. Absorption, magnetic circular dichroism (MCD), and variable-temperature variable-field MCD data are analyzed within the framework of time-dependent DFT computations to assign key electronic transitions. DFT calculations are further employed to evaluate possible reduced F(430) models-a one-electron reduced Ni(I)F(430) model and a three-electron reduced Ni(I)F(red430) model (possessing a reduced hydrocorphin ligand)-on the basis of excited-state spectra and published EPR/ENDOR parameters. While calculations on both models yield spectroscopic parameters that are consistent with most experimental data, overall better agreement is achieved using the Ni(I)F(430) model, particularly with respect to electronic absorption and (1)H ENDOR. The experimentally validated bonding descriptions generated herein show that in Ni(II)F(430) the occupied Ni 3d orbitals are too low in energy to significantly perturb the dominant electronic transition involving the pi and pi* frontier MOs of the macrocycle (i.e., the HOMO-->LUMO transition). Upon one-electron reduction of the Ni(II) ion, the occupied Ni 3d orbitals are raised in energy, shifting between the HOMO and the LUMO of the oxidized cofactor. These ground-state changes have a dramatic effect on the excited-state structure, causing a blue shift of the dominant pi-->pi* transition and the appearance of numerous Ni 3d-->hydrocorphin pi* charge-transfer features in the vis/near-IR region.