Carrageenans as a new source of drugs with metal binding properties.

PubMed

Khotimchenko, Yuri S; Khozhaenko, Elena V; Khotimchenko, Maxim Y; Kolenchenko, Elena A; Kovalev, Valeri V

2010-04-01

Carrageenans are abundant and safe non-starch polysaccharides exerting their biological effects in living organisms. Apart from their known pro-inflammation properties and some pharmacological activity, carrageenans can also strongly bind and hold metal ions. This property can be used for creation of the new drugs for elimination of metals from the body or targeted delivery of these metal ions for healing purposes. Metal binding activity of different carrageenans in aqueous solutions containing Y(3+) or Pb(2+) ions was studied in a batch sorption system. The metal uptake by carrageenans is not affected by the change of the pH within the range from 2.0 to 6.0. The rates and binding capacities of carrageenans regarding metal ions were evaluated. The Langmuir, Freundlich and BET sorption models were applied to describe the isotherms and constants, and the sorption isothermal data could be explained well by the Langmuir equation. The results obtained through the study suggest that kappa-, iota-, and lambda-carrageenans are favorable sorbents. The largest amount of Y(3+) and Pb(2+) ions are bound by iota-carrageenan. Therefore, it can be concluded that this type of polysaccharide is the more appropriate substance for elaboration of the drugs with high selective metal binding properties.

Metalloregulatory Proteins: Metal Selectivity and Allosteric Switching

PubMed Central

Caballero, Hermes Reyes; Campanello, Gregory C.; Giedroc, David P.

2011-01-01

Prokaryotic organisms have evolved an impressive capacity to quickly adapt to a changing and challenging microenvironment in which the availability of both biologically required and non-essential transition metal ions can vary dramatically. In all bacteria, a panel of metalloregulatory proteins control the expression of genes encoding membrane transporters and metal trafficking proteins, that collectively manage metal homeostasis and resistance. These “metal sensors” are specialized allosteric proteins, in which the direct binding of a specific or small number of “cognate” metal ion(s) drives a conformational change in the regulator that allosterically activates or inhibits operator DNA binding, or alternatively, distorts the promoter structure thereby converting a poor promoter to a strong one. In this review, we discuss our current understanding of the features that control metal specificity of the allosteric response in these systems, and the role that structure, thermodynamics and conformational dynamics play in mediating allosteric activation or inhibition of DNA binding. PMID:21511390

Requirement for transient metal ions revealed through computational analysis for DNA polymerase going in reverse

PubMed Central

Perera, Lalith; Freudenthal, Bret D.; Beard, William A.; Shock, David D.; Pedersen, Lee G.; Wilson, Samuel H.

2015-01-01

DNA polymerases facilitate faithful insertion of nucleotides, a central reaction occurring during DNA replication and repair. DNA synthesis (forward reaction) is “balanced,” as dictated by the chemical equilibrium by the reverse reaction of pyrophosphorolysis. Two closely spaced divalent metal ions (catalytic and nucleotide-binding metals) provide the scaffold for these reactions. The catalytic metal lowers the pKa of O3′ of the growing primer terminus, and the nucleotide-binding metal facilitates substrate binding. Recent time-lapse crystallographic studies of DNA polymerases have identified an additional metal ion (product metal) associated with pyrophosphate formation, leading to the suggestion of its possible involvement in the reverse reaction. Here, we establish a rationale for a role of the product metal using quantum mechanical/molecular mechanical calculations of the reverse reaction in the confines of the DNA polymerase β active site. Additionally, site-directed mutagenesis identifies essential residues and metal-binding sites necessary for pyrophosphorolysis. The results indicate that the catalytic metal site must be occupied by a magnesium ion for pyrophosphorolysis to occur. Critically, the product metal site is occupied by a magnesium ion early in the pyrophosphorolysis reaction path but must be removed later. The proposed dynamic nature of the active site metal ions is consistent with crystallographic structures. The transition barrier for pyrophosphorolysis was estimated to be significantly higher than that for the forward reaction, consistent with kinetic activity measurements of the respective reactions. These observations provide a framework to understand how ions and active site changes could modulate the internal chemical equilibrium of a reaction that is central to genome stability. PMID:26351676

Direct Measurement of the Nanomechanical Stability of a Redox Protein Active Site and Its Dependence upon Metal Binding.

PubMed

Giannotti, Marina I; Cabeza de Vaca, Israel; Artés, Juan M; Sanz, Fausto; Guallar, Victor; Gorostiza, Pau

2015-09-10

The structural basis of the low reorganization energy of cupredoxins has long been debated. These proteins reconcile a conformationally heterogeneous and exposed metal-chelating site with the highly rigid copper center required for efficient electron transfer. Here we combine single-molecule mechanical unfolding experiments with statistical analysis and computer simulations to show that the metal-binding region of apo-azurin is mechanically flexible and that high mechanical stability is imparted by copper binding. The unfolding pathway of the metal site depends on the pulling residue and suggests that partial unfolding of the metal-binding site could be facilitated by the physical interaction with certain regions of the redox protein.

Subunit Dissociation and Metal Binding by Escherichia coli apo-Manganese Superoxide Dismutase

PubMed Central

Whittaker, Mei M.; Lerch, Thomas F.; Kirillova, Olga; Chapman, Michael S.; Whittaker, James W.

2010-01-01

Metal binding by apo-manganese superoxide dismutase (apo-MnSOD) is essential for functional maturation of the enzyme. Previous studies have demonstrated that metal binding by apo-MnSOD is conformationally gated, requiring protein reorganization for the metal to bind. We have now solved the X-ray crystal structure of apo-MnSOD at 1.9 Å resolution. The organization of active site residues is independent of the presence of the metal cofactor, demonstrating that protein itself templates the unusual metal coordination geometry. Electrophoretic analysis of mixtures of apo- and (Mn2)-MnSOD, dye-conjugated protein, or C-terminal Strep-tag II fusion protein reveals a dynamic subunit exchange process associated with cooperative metal binding by the two subunits of the dimeric protein. In contrast, (S126C) (SS) apo-MnSOD, which contains an inter-subunit covalent disulfide crosslink, exhibits anticooperative metal binding. The protein concentration dependence of metal uptake kinetics implies that protein dissociation is involved in metal binding by the wild type apo-protein, although other processes may also contribute to gating metal uptake. Protein concentration dependent small-zone size exclusion chromatography is consistent with apo-MnSOD dimer dissociation at low protein concentration (KD = 1×10−6 M). Studies on metal uptake by apo-MnSOD in Escherichia coli cells show that the protein exhibits similar behavior in vivo and in vitro. PMID:21044611

Functional Analysis of the Citrate Activator CitO from Enterococcus faecalis Implicates a Divalent Metal in Ligand Binding

PubMed Central

Blancato, Víctor S.; Pagliai, Fernando A.; Magni, Christian; Gonzalez, Claudio F.; Lorca, Graciela L.

2016-01-01

The regulator of citrate metabolism, CitO, from Enterococcus faecalis belongs to the FCD family within the GntR superfamily. In the presence of citrate, CitO binds to cis-acting sequences located upstream of the cit promoters inducing the expression of genes involved in citrate utilization. The quantification of the molecular binding affinities, performed by isothermal titration calorimetry (ITC), indicated that CitO has a high affinity for citrate (KD = 1.2 ± 0.2 μM), while it did not recognize other metabolic intermediates. Based on a structural model of CitO where a putative small molecule and a metal binding site were identified, it was hypothesized that the metal ion is required for citrate binding. In agreement with this model, citrate binding to CitO sharply decreased when the protein was incubated with EDTA. This effect was reverted by the addition of Ni2+, and Zn2+ to a lesser extent. Structure-based site-directed mutagenesis was conducted and it was found that changes to alanine in residues Arg97 and His191 resulted in decreased binding affinities for citrate, as determined by EMSA and ITC. Further assays using lacZ fusions confirmed that these residues in CitO are involved in sensing citrate in vivo. These results indicate that the molecular modifications induced by a ligand and a metal binding in the C-terminal domain of CitO are required for optimal DNA binding activity, and consequently, transcriptional activation. PMID:26903980

Theoretical Insights into Methane C–H Bond Activation on Alkaline Metal Oxides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aljama, Hassan; Nørskov, Jens K.; Abild-Pedersen, Frank

Here, we investigate the role of alkaline metal oxides (AMO) (MgO, CaO, and SrO) in activating the C–H bond in methane. We also use Density Functional Theory (DFT) and microkinetic modeling to study the catalytic elementary steps in breaking the C–H bond in methane and creating the methyl radical, a precursor prior to creating C2 products. We also study the effects of surface geometry on the catalytic activity of AMO by examining terrace and step sites. We observe that the process of activating methane depends strongly on the structure of the AMO. When the AMO surface is doped with anmore » alkali metal, the transition state (TS) structure has a methyl radical-like behavior, where the methyl radical interacts weakly with the AMO surface. In this case, the TS energy scales with the hydrogen binding energy. On pure AMO, the TS interacts with AMO surface oxygen as well as the metal atom on the surface, and consequently the TS energy scales with the binding energy of hydrogen and methyl. We study the activity of AMO using a mean-field microkinetic model. The results indicate that terrace sites have similar catalytic activity, with the exception of MgO(100). Step sites bind hydrogen more strongly, making them more active, and this confirms previously reported experimental results. We map the catalytic activity of AMO using a volcano plot with two descriptors: the methyl and the hydrogen binding energies, with the latter being a more significant descriptor. The microkinetic model results suggest that C–H bond dissociation is not always the rate-limiting step. At weak hydrogen binding, the reaction is limited by C–H bond activation. At strong hydrogen binding, the reaction is limited due to poisoning of the active site. We found an increase in activity of AMO as the basicity increased. Finally, the developed microkinetic model allows screening for improved catalysts using simple calculations of the hydrogen binding energy.« less

Theoretical Insights into Methane C–H Bond Activation on Alkaline Metal Oxides

DOE PAGES

Aljama, Hassan; Nørskov, Jens K.; Abild-Pedersen, Frank

2017-07-17

Here, we investigate the role of alkaline metal oxides (AMO) (MgO, CaO, and SrO) in activating the C–H bond in methane. We also use Density Functional Theory (DFT) and microkinetic modeling to study the catalytic elementary steps in breaking the C–H bond in methane and creating the methyl radical, a precursor prior to creating C2 products. We also study the effects of surface geometry on the catalytic activity of AMO by examining terrace and step sites. We observe that the process of activating methane depends strongly on the structure of the AMO. When the AMO surface is doped with anmore » alkali metal, the transition state (TS) structure has a methyl radical-like behavior, where the methyl radical interacts weakly with the AMO surface. In this case, the TS energy scales with the hydrogen binding energy. On pure AMO, the TS interacts with AMO surface oxygen as well as the metal atom on the surface, and consequently the TS energy scales with the binding energy of hydrogen and methyl. We study the activity of AMO using a mean-field microkinetic model. The results indicate that terrace sites have similar catalytic activity, with the exception of MgO(100). Step sites bind hydrogen more strongly, making them more active, and this confirms previously reported experimental results. We map the catalytic activity of AMO using a volcano plot with two descriptors: the methyl and the hydrogen binding energies, with the latter being a more significant descriptor. The microkinetic model results suggest that C–H bond dissociation is not always the rate-limiting step. At weak hydrogen binding, the reaction is limited by C–H bond activation. At strong hydrogen binding, the reaction is limited due to poisoning of the active site. We found an increase in activity of AMO as the basicity increased. Finally, the developed microkinetic model allows screening for improved catalysts using simple calculations of the hydrogen binding energy.« less

N-Acetylcysteine interacts with copper to generate hydrogen peroxide and selectively induce cancer cell death

PubMed Central

Zheng, Jie; Lou, Jessica R.; Zhang, Xiao-Xi; Benbrook, Doris M.; Hanigan, Marie H.; Lind, Stuart E.; Ding, Wei-Qun

2013-01-01

A variety of metal-binding compounds have been found to exert anti-cancer activity. We postulated that N-acetylcysteine (NAC), which is a membrane-permeable metal-binding compound, might have anti-cancer activity in the presence of metals. We found that NAC/Cu(II) significantly alters growth and induces apoptosis in human cancer lines, yet NAC/Zn(II) and NAC/Fe(III) do not. We further confirmed that this cytotoxicity of NAC/Cu(II) is attributed to reactive oxygen species (ROS). These findings indicate that the combination of Cu(II) and thiols generates cytotoxic ROS that induce apoptosis in cancer cells. They also indicate a fourth class of anti-neoplastic metal-binding compounds, the “ROS generator”. PMID:20667650

Distinct roles of beta1 metal ion-dependent adhesion site (MIDAS), adjacent to MIDAS (ADMIDAS), and ligand-associated metal-binding site (LIMBS) cation-binding sites in ligand recognition by integrin alpha2beta1.

PubMed

Valdramidou, Dimitra; Humphries, Martin J; Mould, A Paul

2008-11-21

Integrin-ligand interactions are regulated in a complex manner by divalent cations, and previous studies have identified ligand-competent, stimulatory, and inhibitory cation-binding sites. In collagen-binding integrins, such as alpha2beta1, ligand recognition takes place exclusively at the alpha subunit I domain. However, activation of the alphaI domain depends on its interaction with a structurally similar domain in the beta subunit known as the I-like or betaI domain. The top face of the betaI domain contains three cation-binding sites: the metal-ion dependent adhesion site (MIDAS), the ADMIDAS (adjacent to MIDAS), and LIMBS (ligand-associated metal-binding site). The role of these sites in controlling ligand binding to the alphaI domain has yet to be elucidated. Mutation of the MIDAS or LIMBS completely blocked collagen binding to alpha2beta1; in contrast mutation of the ADMIDAS reduced ligand recognition but this effect could be overcome by the activating monoclonal antibody TS2/16. Hence, the MIDAS and LIMBS appear to be essential for the interaction between alphaI and betaI, whereas occupancy of the ADMIDAS has an allosteric effect on the conformation of betaI. An activating mutation in the alpha2 I domain partially restored ligand binding to the MIDAS and LIMBS mutants. Analysis of the effects of Ca(2+), Mg(2+), and Mn(2+) on ligand binding to these mutants showed that the MIDAS is a ligand-competent site through which Mn(2+) stimulates ligand binding, whereas the LIMBS is a stimulatory Ca(2+)-binding site, occupancy of which increases the affinity of Mg(2+) for the MIDAS.

Spectroscopic characterization of metal bound phytochelatin analogue (Glu-Cys)4-Gly.

PubMed

Cheng, Yongsheng; Yan, Yong-Bin; Liu, Jinyuan

2005-10-01

The metal ion binding properties of a phytochelatin (PC) analogue, (Glu-Cys)4-Gly (named as EC4), have been studied by a divalent metal ion binding assay monitored by UV-visible spectroscopy, circular dichroism and NMR spectroscopy. Spectro- photometric titration with different divalent metal ions have revealed that the stiochoimetry of metal-bound EC4 was 1:1, and its metal binding affinities with different divalent metal ions in the order of Cd(II)>Cu(II)>Zn(II)>Pb(II)>Ni(II)>Co(II). UV-visible spectroscopic analysis of metal complexes indicated that four sulfur atoms in cysteine residues are attributable to ligand-to-metal charge transfer (LMCT) between divalent metal ions and EC4, and further confirmed by 1D H1 NMR study and Circular Dichroism. In addition, Circular Dichroism spectra of both free and metal-bound forms of EC4 revealed that metal coordination drives the nonapeptide chain to fold into a turned conformation. The comprehensive analysis of spectroscopic properties of the nonapeptide complexed with metal ions not only provides a fundamental description of the metal ion binding properties of PC analogue, but also shows a correlation between metal binding affinity of PC analogue and the induction activity of metal ions.

Engineered Single-Chain, Antiparallel, Coiled Coil Mimics the MerR Metal Binding Site

PubMed Central

Song, Lingyun; Caguiat, Jonathan; Li, Zhongrui; Shokes, Jacob; Scott, Robert A.; Olliff, Lynda; Summers, Anne O.

2004-01-01

The repressor-activator MerR that controls transcription of the mercury resistance (mer) operon is unusual for its high sensitivity and specificity for Hg(II) in in vivo and in vitro transcriptional assays. The metal-recognition domain of MerR resides at the homodimer interface in a novel antiparallel arrangement of α-helix 5 that forms a coiled-coil motif. To facilitate the study of this novel metal binding motif, we assembled this antiparallel coiled coil into a single chain by directly fusing two copies of the 48-residue α-helix 5 of MerR. The resulting 107-residue polypeptide, called the metal binding domain (MBD), and wild-type MerR were overproduced and purified, and their metal-binding properties were determined in vivo and in vitro. In vitro MBD bound ca. 1.0 equivalent of Hg(II) per pair of binding sites, just as MerR does, and it showed only a slightly lower affinity for Hg(II) than did MerR. Extended X-ray absorption fine structure data showed that MBD has essentially the same Hg(II) coordination environment as MerR. In vivo, cells overexpressing MBD accumulated 70 to 100% more 203Hg(II) than cells bearing the vector alone, without deleterious effects on cell growth. Both MerR and MBD variously bound other thiophilic metal ions, including Cd(II), Zn(II), Pb(II), and As(III), in vitro and in vivo. We conclude that (i) it is possible to simulate in a single polypeptide chain the in vitro and in vivo metal-binding ability of dimeric, full-length MerR and (ii) MerR's specificity in transcriptional activation does not reside solely in the metal-binding step. PMID:14996817

Crystal Structure of Phosphatidylglycerophosphatase (PGPase), a Putative Membrane-Bound Lipid Phosphatase, Reveals a Novel Binuclear Metal Binding Site and Two Proton Wires

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kumaran,D.; Bonnano, J.; Burley, S.

2006-01-01

Phosphatidylglycerophosphatase (PGPase), an enzyme involved in lipid metabolism, catalyzes formation of phosphatidylglycerol from phosphatidylglycerophosphate. Phosphatidylglycerol is a multifunctional phospholipid, found in the biological membranes of many organisms. Here, we report the crystal structure of Listeria monocytogenes PGPase at 1.8 Angstroms resolution. PGPase, an all-helical molecule, forms a homotetramer. Each protomer contains an independent active site with two metal ions, Ca{sup 2+} and Mg{sup 2+}, forming a hetero-binuclear center located in a hydrophilic cavity near the surface of the molecule. The binuclear center, conserved ligands, metal-bound water molecules, and an Asp-His dyad form the active site. The catalytic mechanism of thismore » enzyme is likely to proceed via binuclear metal activated nucleophilic water. The binuclear metal-binding active-site environment of this structure should provide insights into substrate binding and metal-dependent catalysis. A long channel with inter-linked linear water chains, termed 'proton wires', is observed at the tetramer interface. Comparison of similar water chain structures in photosynthetic reaction centers (RCs), Cytochrome f, gramicidin, and bacteriorhodopsin, suggests that PGPase may conduct protons via proton wires.« less

Effects of metal on the biochemical properties of Helicobacter pylori HypB, a maturation factor of [NiFe]-hydrogenase and urease.

PubMed

Sydor, Andrew M; Liu, Jenny; Zamble, Deborah B

2011-03-01

The biosyntheses of the [NiFe]-hydrogenase and urease enzymes in Helicobacter pylori require several accessory proteins for proper construction of the nickel-containing metallocenters. The hydrogenase accessory proteins HypA and HypB, a GTPase, have been implicated in the nickel delivery steps of both enzymes. In this study, the metal-binding properties of H. pylori HypB were characterized, and the effects of metal binding on the biochemical behavior of the protein were examined. The protein can bind stoichiometric amounts of Zn(II) or Ni(II), each with nanomolar affinity. Mutation of Cys106 and His107, which are located between two major GTPase motifs, results in undetectable Ni(II) binding, and the Zn(II) affinity is weakened by 2 orders of magnitude. These two residues are also required for the metal-dependent dimerization observed in the presence of Ni(II) but not Zn(II). The addition of metals to the protein has distinct impacts on GTPase activity, with zinc significantly reducing GTP hydrolysis to below detectable levels and nickel only slightly altering the k(cat) and K(m) of the reaction. The regulation of HypB activities by metal binding may contribute to the maturation of the nickel-containing enzymes.

Metal selectivity of the E. coli nickel metallochaperone, SlyD

PubMed Central

Kaluarachchi, Harini; Siebel, Judith F.; Kaluarachchi-Duffy, Supipi; Krecisz, Sandra; Sutherland, Duncan E. K.; Stillman, Martin J.; Zamble, Deborah B.

2012-01-01

SlyD is a Ni(II)-binding protein that contributes to nickel homeostasis in Escherichia coli. The C-terminal domain of SlyD contains a rich variety of metal-binding amino acids, suggesting broader metal-binding capabilities, and previous work demonstrated that the protein can coordinate several types of first row transition metals. However, the binding of SlyD to metals other than Ni(II) has not been previously characterized. To further our understanding of the in vitro metal-binding activity of SlyD and how it correlates with the in vivo function of this protein, the interactions between SlyD and the series of biologically relevant transition metals Mn(II), Fe(II), Co(II), Cu(I) and Zn(II) were examined by using a combination of optical spectroscopy and mass spectrometry. SlyD binding to Mn(II) or to Fe(II) ions was not detected but the protein coordinates multiple ions of Co(II), Zn(II) and Cu(I) with appreciable affinities (KD ≤ nM), highlighting the promiscuous nature of this protein. The order of affinities of SlyD for the metals examined is Mn(II), Fe(II) < Co(II) < Ni(II) ~ Zn(II) ≪ Cu(I). Although the purified protein is unable to overcome the large thermodynamic preference for Cu(I) and exclude Zn(II) chelation in the presence of Ni(II), in vivo studies reveal a Ni(II)-specific function for the protein. Furthermore, these latter experiments support a specific role for SlyD as a [NiFe]-hydrogenase enzyme maturation factor. The implications of the divergence between the metal selectivity of SlyD in vitro and the specific activity in vivo are discussed. PMID:22047179

Metal Ion Binding at the Catalytic Site Induces Widely Distributed Changes in a Sequence Specific Protein–DNA Complex

PubMed Central

2016-01-01

Metal ion cofactors can alter the energetics and specificity of sequence specific protein–DNA interactions, but it is unknown if the underlying effects on structure and dynamics are local or dispersed throughout the protein–DNA complex. This work uses EcoRV endonuclease as a model, and catalytically inactive lanthanide ions, which replace the Mg2+ cofactor. Nuclear magnetic resonance (NMR) titrations indicate that four Lu3+ or two La3+ cations bind, and two new crystal structures confirm that Lu3+ binding is confined to the active sites. NMR spectra show that the metal-free EcoRV complex with cognate (GATATC) DNA is structurally distinct from the nonspecific complex, and that metal ion binding sites are not assembled in the nonspecific complex. NMR chemical shift perturbations were determined for 1H–15N amide resonances, for 1H–13C Ile-δ-CH3 resonances, and for stereospecifically assigned Leu-δ-CH3 and Val-γ-CH3 resonances. Many chemical shifts throughout the cognate complex are unperturbed, so metal binding does not induce major conformational changes. However, some large perturbations of amide and side chain methyl resonances occur as far as 34 Å from the metal ions. Concerted changes in specific residues imply that local effects of metal binding are propagated via a β-sheet and an α-helix. Both amide and methyl resonance perturbations indicate changes in the interface between subunits of the EcoRV homodimer. Bound metal ions also affect amide hydrogen exchange rates for distant residues, including a distant subdomain that contacts DNA phosphates and promotes DNA bending, showing that metal ions in the active sites, which relieve electrostatic repulsion between protein and DNA, cause changes in slow dynamics throughout the complex. PMID:27786446

The 1.3 A resolution structure of the RNA tridecamer r(GCGUUUGAAACGC): metal ion binding correlates with base unstacking and groove contraction.

PubMed

Timsit, Youri; Bombard, Sophie

2007-12-01

Metal ions play a key role in RNA folding and activity. Elucidating the rules that govern the binding of metal ions is therefore an essential step for better understanding the RNA functions. High-resolution data are a prerequisite for a detailed structural analysis of ion binding on RNA and, in particular, the observation of monovalent cations. Here, the high-resolution crystal structures of the tridecamer duplex r(GCGUUUGAAACGC) crystallized under different conditions provides new structural insights on ion binding on GAAA/UUU sequences that exhibit both unusual structural and functional properties in RNA. The present study extends the repertory of RNA ion binding sites in showing that the two first bases of UUU triplets constitute a specific site for sodium ions. A striking asymmetric pattern of metal ion binding in the two equivalent halves of the palindromic sequence demonstrates that sequence and its environment act together to bind metal ions. A highly ionophilic half that binds six metal ions allows, for the first time, the observation of a disodium cluster in RNA. The comparison of the equivalent halves of the duplex provides experimental evidences that ion binding correlates with structural alterations and groove contraction.

The Mismetallation of Enzymes during Oxidative Stress*

PubMed Central

Imlay, James A.

2014-01-01

Mononuclear iron enzymes can tightly bind non-activating metals. How do cells avoid mismetallation? The model bacterium Escherichia coli may control its metal pools so that thermodynamics favor the correct metallation of each enzyme. This system is disrupted, however, by superoxide and hydrogen peroxide. These species oxidize ferrous iron and thereby displace it from many iron-dependent mononuclear enzymes. Ultimately, zinc binds in its place, confers little activity, and imposes metabolic bottlenecks. Data suggest that E. coli compensates by using thiols to extract the zinc and by importing manganese to replace the catalytic iron atom. Manganese resists oxidants and provides substantial activity. PMID:25160623

Zinc can increase the activity of protein kinase C and contributes to its binding to plasma membranes in T lymphocytes.

PubMed

Csermely, P; Szamel, M; Resch, K; Somogyi, J

1988-05-15

In the primary structure of protein kinase C, the presence of a putative metal-binding site has been suggested (Parker, P.J., Coussens, L., Totty, N., Rhee, L., Young, S., Chen, E., Stabel, S., Waterfield, M.D., and Ullrich, A. (1986) Science 233, 853-859). In the present report, we demonstrate that the most abundant intracellular heavy metal, zinc, can increase the activity of cytosolic protein kinase C. Zinc reversibly binds the enzyme to plasma membranes, and it may contribute to the calcium-induced binding as well. The intracellular heavy metal chelator N,N,N',N'-tetrakis(2-pyridylmethyl) ethylenediamine prevents the phorbol ester- and antigen-induced translocation of protein kinase C. This effect can be totally reversed by the concomitant addition of Zn2+, while Fe2+ and Mn2+ are only partially counteractive. Our results suggest that zinc can activate protein kinase C and contributes to its binding to plasma membranes in T lymphocytes induced by Ca2+, phorbol ester, or antigen.

Spectro Analytical, Computational and In Vitro Biological Studies of Novel Substituted Quinolone Hydrazone and it's Metal Complexes.

PubMed

Nagula, Narsimha; Kunche, Sudeepa; Jaheer, Mohmed; Mudavath, Ravi; Sivan, Sreekanth; Ch, Sarala Devi

2018-01-01

Some novel transition metal [Cu (II), Ni (II) and Co (II)] complexes of nalidixic acid hydrazone have been prepared and characterized by employing spectro-analytical techniques viz: elemental analysis, 1 H-NMR, Mass, UV-Vis, IR, TGA-DTA, SEM-EDX, ESR and Spectrophotometry studies. The HyperChem 7.5 software was used for geometry optimization of title compound in its molecular and ionic forms. Quantum mechanical parameters, contour maps of highest occupied molecular orbitals (HOMO) and lowest unoccupied molecular orbitals (LUMO) and corresponding binding energy values were computed using semi empirical single point PM3 method. The stoichiometric equilibrium studies of metal complexes carried out spectrophotometrically using Job's continuous variation and mole ratio methods inferred formation of 1:2 (ML 2 ) metal complexes in respective systems. The title compound and its metal complexes screened for antibacterial and antifungal properties, exemplified improved activity in metal complexes. The studies of nuclease activity for the cleavage of CT- DNA and MTT assay for in vitro cytotoxic properties involving metal complexes exhibited high activity. In addition, the DNA binding properties of Cu (II), Ni (II) and Co (II) complexes investigated by electronic absorption and fluorescence measurements revealed their good binding ability and commended agreement of K b values obtained from both the techniques. Molecular docking studies were also performed to find the binding affinity of synthesized compounds with DNA (PDB ID: 1N37) and "Thymidine phosphorylase from E.coli" (PDB ID: 4EAF) protein targets.

Affinity to bovine serum albumin and anticancer activity of some new water-soluble metal Schiff base complexes

NASA Astrophysics Data System (ADS)

Asadi, Mozaffar; Asadi, Zahra; Zarei, Leila; Sadi, Somaye Barzegar; Amirghofran, Zahra

2014-12-01

Metal Schiff-base complexes show biological activity but they are usually insoluble in water so four new water-soluble metal Schiff base complexes of Na2[M(5-SO3-1,2-salben]; (5-SO3-1,2-salben denoted N,N";-bis(5-sulphosalicyliden)-1,2-diaminobenzylamine and M = Mg, Mn, Cu, Zn) were synthesized and characterized. The formation constants of the metal complexes were determined by UV-Vis absorption spectroscopy. The interaction of these complexes with bovine serum albumin (BSA) was studied by fluorescence spectroscopy. Type of quenching, binding constants, number of binding sites and binding stoichiometries were determined by fluorescence quenching method. The results showed that the mentioned complexes strongly bound to BSA. Thermodynamic parameters indicated that hydrophobic association was the major binding force and that the interaction was entropy driven and enthalpically disfavoured. The displacement experiment showed that these complexes could bind to the subdomain IIA (site I) of albumin. Furthermore the synchronous fluorescence spectra showed that the microenvironment of the tryptophan residues was not apparently changed. Based on the Förster theory of non-radiation energy transfer, the distance between the donor (Trp residues) and the acceptor metal complexes was obtained. The growth inhibitory effect of complexes toward the K562 cancer cell line was measured.

Metal site occupancy and allosteric switching in bacterial metal sensor proteins.

PubMed

Guerra, Alfredo J; Giedroc, David P

2012-03-15

All prokaryotes encode a panel of metal sensor or metalloregulatory proteins that govern the expression of genes that allows an organism to quickly adapt to toxicity or deprivation of both biologically essential transition metal ions, e.g., Zn, Cu, Fe, and heavy metal pollutants. As such, metal sensor proteins can be considered arbiters of intracellular transition metal bioavailability and thus potentially control the metallation state of the metalloproteins in the cell. Metal sensor proteins are specialized allosteric proteins that regulate transcription as a result direct binding of one or two cognate metal ions, to the exclusion of all others. In most cases, the binding of the cognate metal ion induces a structural change in a protein oligomer that either activates or inhibits operator DNA binding. A quantitative measure of the degree to which a particular metal drives metalloregulation of operator DNA-binding is the allosteric coupling free energy, ΔGc. In this review, we summarize recent work directed toward understanding metal occupancy and metal selectivity of these allosteric switches in selected families of metal sensor proteins and examine the structural origins of ΔGc in the functional context a thermodynamic "set-point" model of intracellular metal homeostasis. Copyright © 2011 Elsevier Inc. All rights reserved.

Metal-Ion Effects on the Polarization of Metal-Bound Water and Infrared Vibrational Modes of the Coordinated Metal Center of Mycobacterium tuberculosis Pyrazinamidase via Quantum Mechanical Calculations

PubMed Central

2014-01-01

Mycobacterium tuberculosis pyrazinamidase (PZAse) is a key enzyme to activate the pro-drug pyrazinamide (PZA). PZAse is a metalloenzyme that coordinates in vitro different divalent metal cofactors in the metal coordination site (MCS). Several metals including Co2+, Mn2+, and Zn2+ are able to reactivate the metal-depleted PZAse in vitro. We use quantum mechanical calculations to investigate the Zn2+, Fe2+, and Mn2+ metal cofactor effects on the local MCS structure, metal–ligand or metal–residue binding energy, and charge distribution. Results suggest that the major metal-dependent changes occur in the metal–ligand binding energy and charge distribution. Zn2+ shows the highest binding energy to the ligands (residues). In addition, Zn2+ and Mn2+ within the PZAse MCS highly polarize the O–H bond of coordinated water molecules in comparison with Fe2+. This suggests that the coordination of Zn2+ or Mn2+ to the PZAse protein facilitates the deprotonation of coordinated water to generate a nucleophile for catalysis as in carboxypeptidase A. Because metal ion binding is relevant to enzymatic reaction, identification of the metal binding event is important. The infrared vibrational mode shift of the C=Nε (His) bond from the M. tuberculosis MCS is the best IR probe to metal complexation. PMID:25055049

Structural basis of PP2A activation by PTPA, an ATP-dependent activation chaperone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Feng; Stanevich, Vitali; Wlodarchak, Nathan

Proper activation of protein phosphatase 2A (PP2A) catalytic subunit is central for the complex PP2A regulation and is crucial for broad aspects of cellular function. The crystal structure of PP2A bound to PP2A phosphatase activator (PTPA) and ATPγS reveals that PTPA makes broad contacts with the structural elements surrounding the PP2A active site and the adenine moiety of ATP. PTPA-binding stabilizes the protein fold of apo-PP2A required for activation, and orients ATP phosphoryl groups to bind directly to the PP2A active site. This allows ATP to modulate the metal-binding preferences of the PP2A active site and utilize the PP2A activemore » site for ATP hydrolysis. In vitro, ATP selectively and drastically enhances binding of endogenous catalytic metal ions, which requires ATP hydrolysis and is crucial for acquisition of pSer/Thr-specific phosphatase activity. Furthermore, both PP2A- and ATP-binding are required for PTPA function in cell proliferation and survival. Our results suggest novel mechanisms of PTPA in PP2A activation with structural economy and a unique ATP-binding pocket that could potentially serve as a specific therapeutic target.« less

Occupancy of the Zinc-binding Site by Transition Metals Decreases the Substrate Affinity of the Human Dopamine Transporter by an Allosteric Mechanism*

PubMed Central

Li, Yang; Mayer, Felix P.; Hasenhuetl, Peter S.; Burtscher, Verena; Schicker, Klaus; Sitte, Harald H.; Freissmuth, Michael; Sandtner, Walter

2017-01-01

The human dopamine transporter (DAT) has a tetrahedral Zn2+-binding site. Zn2+-binding sites are also recognized by other first-row transition metals. Excessive accumulation of manganese or of copper can lead to parkinsonism because of dopamine deficiency. Accordingly, we examined the effect of Mn2+, Co2+, Ni2+, and Cu2+ on transport-associated currents through DAT and DAT-H193K, a mutant with a disrupted Zn2+-binding site. All transition metals except Mn2+ modulated the transport cycle of wild-type DAT with affinities in the low micromolar range. In this concentration range, they were devoid of any action on DAT-H193K. The active transition metals reduced the affinity of DAT for dopamine. The affinity shift was most pronounced for Cu2+, followed by Ni2+ and Zn2+ (= Co2+). The extent of the affinity shift and the reciprocal effect of substrate on metal affinity accounted for the different modes of action: Ni2+ and Cu2+ uniformly stimulated and inhibited, respectively, the substrate-induced steady-state currents through DAT. In contrast, Zn2+ elicited biphasic effects on transport, i.e. stimulation at 1 μm and inhibition at 10 μm. A kinetic model that posited preferential binding of transition metal ions to the outward-facing apo state of DAT and a reciprocal interaction of dopamine and transition metals recapitulated all experimental findings. Allosteric activation of DAT via the Zn2+-binding site may be of interest to restore transport in loss-of-function mutants. PMID:28096460

Three-dimensional metal-intercalated covalent organic frameworks for near-ambient energy storage

PubMed Central

Gao, Fei; Ding, Zijing; Meng, Sheng

2013-01-01

A new form of nanoporous material, metal intercalated covalent organic framework (MCOF) is proposed and its energy storage property revealed. Employing density functional and thermodynamical analysis, we find that stable, chemically active, porous materials could form by stacking covalent organic framework (COF) layers with metals as a gluing agent. Metal acts as active sites, while its aggregation is suppressed by a binding energy significantly larger than the corresponding cohesive energy of bulk metals. Two important parameters, metal binding and metal-metal separation, are tuned by selecting suitable building blocks and linkers when constructing COF layers. Systematic searches among a variety of elements and organic molecules identify Ca-intercalated COF with diphenylethyne units as optimal material for H2 storage, reaching a striking gravimetric density ~ 5 wt% at near-ambient conditions (300 K, 20 bar), in comparison to < 0.1 wt% for bare COF-1 under the same condition. PMID:23698018

Metal binding of flavonoids and their distinct inhibition mechanisms toward the oxidation activity of Cu2+-β-amyloid: not just serving as suicide antioxidants!

PubMed

Tay, William Maung; da Silva, Giordano F Z; Ming, Li-June

2013-01-18

The accumulation of plagues of β-amyloid (Aβ) peptides in the brain is a hallmark of Alzheimer's disease (AD). The redox-active Cu and Fe complexes of Aβ can cause damage to the neurons potentially via reactive oxygen species (ROS). The significant metal-mediated oxidative activity of CuAβ suggests that its presence can be chemically devastating regardless whether it is a cause or a result of AD. Flavonoids exhibit various benefits to human health, attributable to their metal-binding and antioxidation activities to certain extents. Despite broad interests and extensive studies of their metal-binding properties and anti/pro-oxidation activities, these properties and the mechanisms of the activities toward metal-centered oxidation reactions have not been fully revealed and concluded. We report herein distinctive antioxidation mechanisms between two flavonoid families toward the oxidation reactions by CuAβ(1-20), wherein the flavonols quercetin (Qr) and myricetin (Mr) competitively inhibit the oxidation of catechol by CuAβ(1-20) with K(i) of 11.2 and 32.6 μM, respectively, whereas the flavanols catechin (Ct) and epicatechin (Et) are substrates with k(cat) = 1.01 × 10(-2) and 1.55 × 10(-3) s(-1) and K(m) = 0.94 and 0.55 mM, respectively. Qr has a nearly 10-fold higher antioxidative efficacy than Ct against the oxidation activity of CuAβ, while Ct is effectively oxidized, which further decreases its antioxidant capacity. Similar inhibition patterns are observed toward oxidation of the catecholamine neurotransmitter dopamine by CuAβ(1-20). Metal ions and CuAβ bind Qr with a 1:1 ratio under our experimental conditions through the α-ketoenolate moiety as determined by the use of Co(2+) and Yb(3+) as paramagnetic NMR probes. Unlike flavanols, which are merely suicide antioxidative substrates, flavonols bind to the metal center and prevent metal-mediated redox reactions. We suggest flavonols may serve as leads for drug discovery and/or as agents toward preventing metal-mediated oxidative stress due to AD and other disorders. Moreover, CuAβ shows 8.6- and 4.2-fold higher kinetic regioselectivity in terms of k(cat) and k(cat)/K(m), respectively, toward the peroxidation of Ct than that of the enantiomer Et, suggesting potential development of metallo-catalysts in regioselective catalysis by the use of metallopeptides as templates.

A new metal binding domain involved in cadmium, cobalt and zinc transport

DOE PAGES

Smith, Aaron T.; Barupala, Dulmini; Stemmler, Timothy L.; ...

2015-07-20

The P 1B-ATPases, which couple cation transport across membranes to ATP hydrolysis, are central to metal homeostasis in all organisms. An important feature of P 1B-ATPases is the presence of soluble metal binding domains (MBDs) that regulate transport activity. Only one type of MBD has been characterized extensively, but bioinformatics analyses indicate that a diversity of MBDs may exist in nature. In this paper, we report the biochemical, structural and functional characterization of a new MBD from the Cupriavidus metallidurans P 1B-4-ATPase CzcP (CzcP MBD). The CzcP MBD binds two Cd 2+, Co 2+ or Zn 2+ ions in distinctmore » and unique sites and adopts an unexpected fold consisting of two fused ferredoxin-like domains. Both in vitro and in vivo activity assays using full-length CzcP, truncated CzcP and several variants indicate a regulatory role for the MBD and distinct functions for the two metal binding sites. Finally, taken together, these findings elucidate a previously unknown MBD and suggest new regulatory mechanisms for metal transport by P 1B-ATPases.« less

A ternary metal binding site in the C2 domain of phosphoinositide-specific phospholipase C-delta1.

PubMed

Essen, L O; Perisic, O; Lynch, D E; Katan, M; Williams, R L

1997-03-11

We have determined the crystal structures of complexes of phosphoinositide-specific phospholipase C-delta1 from rat with calcium, barium, and lanthanum at 2.5-2.6 A resolution. Binding of these metal ions is observed in the active site of the catalytic TIM barrel and in the calcium binding region (CBR) of the C2 domain. The C2 domain of PLC-delta1 is a circularly permuted topological variant (P-variant) of the synaptotagmin I C2A domain (S-variant). On the basis of sequence analysis, we propose that both the S-variant and P-variant topologies are present among other C2 domains. Multiple adjacent binding sites in the C2 domain were observed for calcium and the other metal/enzyme complexes. The maximum number of binding sites observed was for the calcium analogue lanthanum. This complex shows an array-like binding of three lanthanum ions (sites I-III) in a crevice on one end of the C2 beta-sandwich. Residues involved in metal binding are contained in three loops, CBR1, CBR2, and CBR3. Sites I and II are maintained in the calcium and barium complexes, whereas sites II and III coincide with a binary calcium binding site in the C2A domain of synaptotagmin I. Several conformers for CBR1 are observed. The conformation of CBR1 does not appear to be strictly dependent on metal binding; however, metal binding may stabilize certain conformers. No significant structural changes are observed for CBR2 or CBR3. The surface of this ternary binding site provides a cluster of freely accessible liganding positions for putative phospholipid ligands of the C2 domain. It may be that the ternary metal binding site is also a feature of calcium-dependent phospholipid binding in solution. A ternary metal binding site might be a conserved feature among C2 domains that contain the critical calcium ligands in their CBR's. The high cooperativity of calcium-mediated lipid binding by C2 domains described previously is explained by this novel type of calcium binding site.

North Carolina Biomolecular Engineering and Materials Applications Center (NC-BEMAC).

DTIC Science & Technology

1987-12-29

enzyme has been replaced with cobalt(II). A further objective was to investigate Co2 activation by low molecular weight transition metal complexes as...Characterization of Low Molecular Weight Metal Complexes as Potential Models for IBio-Catalytic Processes. A number of transit ion met~~il oom~pi cxe; hive...binding, the enzyme suffered loss of activity during radiation polymerization. When covalent binding was u:sed it was necessary to introduce suitably

Biochemical, Kinetic, and Spectroscopic Characterization of Ruegeria pomeroyi DddW—A Mononuclear Iron-Dependent DMSP Lyase

PubMed Central

Brummett, Adam E.; Schnicker, Nicholas J.; Crider, Alexander; Todd, Jonathan D.; Dey, Mishtu

2015-01-01

The osmolyte dimethylsulfoniopropionate (DMSP) is a key nutrient in marine environments and its catabolism by bacteria through enzymes known as DMSP lyases generates dimethylsulfide (DMS), a gas of importance in climate regulation, the sulfur cycle, and signaling to higher organisms. Despite the environmental significance of DMSP lyases, little is known about how they function at the mechanistic level. In this study we biochemically characterize DddW, a DMSP lyase from the model roseobacter Ruegeria pomeroyi DSS-3. DddW is a 16.9 kDa enzyme that contains a C-terminal cupin domain and liberates acrylate, a proton, and DMS from the DMSP substrate. Our studies show that as-purified DddW is a metalloenzyme, like the DddQ and DddP DMSP lyases, but contains an iron cofactor. The metal cofactor is essential for DddW DMSP lyase activity since addition of the metal chelator EDTA abolishes its enzymatic activity, as do substitution mutations of key metal-binding residues in the cupin motif (His81, His83, Glu87, and His121). Measurements of metal binding affinity and catalytic activity indicate that Fe(II) is most likely the preferred catalytic metal ion with a nanomolar binding affinity. Stoichiometry studies suggest DddW requires one Fe(II) per monomer. Electronic absorption and electron paramagnetic resonance (EPR) studies show an interaction between NO and Fe(II)-DddW, with NO binding to the EPR silent Fe(II) site giving rise to an EPR active species (g = 4.29, 3.95, 2.00). The change in the rhombicity of the EPR signal is observed in the presence of DMSP, indicating that substrate binds to the iron site without displacing bound NO. This work provides insight into the mechanism of DMSP cleavage catalyzed by DddW. PMID:25993446

A structural role for the PHP domain in E. coli DNA polymerase III.

PubMed

Barros, Tiago; Guenther, Joel; Kelch, Brian; Anaya, Jordan; Prabhakar, Arjun; O'Donnell, Mike; Kuriyan, John; Lamers, Meindert H

2013-05-14

In addition to the core catalytic machinery, bacterial replicative DNA polymerases contain a Polymerase and Histidinol Phosphatase (PHP) domain whose function is not entirely understood. The PHP domains of some bacterial replicases are active metal-dependent nucleases that may play a role in proofreading. In E. coli DNA polymerase III, however, the PHP domain has lost several metal-coordinating residues and is likely to be catalytically inactive. Genomic searches show that the loss of metal-coordinating residues in polymerase PHP domains is likely to have coevolved with the presence of a separate proofreading exonuclease that works with the polymerase. Although the E. coli Pol III PHP domain has lost metal-coordinating residues, the structure of the domain has been conserved to a remarkable degree when compared to that of metal-binding PHP domains. This is demonstrated by our ability to restore metal binding with only three point mutations, as confirmed by the metal-bound crystal structure of this mutant determined at 2.9 Å resolution. We also show that Pol III, a large multi-domain protein, unfolds cooperatively and that mutations in the degenerate metal-binding site of the PHP domain decrease the overall stability of Pol III and reduce its activity. While the presence of a PHP domain in replicative bacterial polymerases is strictly conserved, its ability to coordinate metals and to perform proofreading exonuclease activity is not, suggesting additional non-enzymatic roles for the domain. Our results show that the PHP domain is a major structural element in Pol III and its integrity modulates both the stability and activity of the polymerase.

Activity of N-coordinated multi-metal-atom active site structures for Pt-free oxygen reduction reaction catalysis: Role of *OH ligands

DOE PAGES

Holby, Edward F.; Taylor, Christopher D.

2015-03-19

We report calculated oxygen reduction reaction energy pathways on multi-metal-atom structures that have previously been shown to be thermodynamically favorable. We predict that such sites have the ability to spontaneously cleave the O₂ bond and then will proceed to over-bind reaction intermediates. In particular, the *OH bound state has lower energy than the final 2 H₂O state at positive potentials. Contrary to traditional surface catalysts, this *OH binding does not poison the multi-metal-atom site but acts as a modifying ligand that will spontaneously form in aqueous environments leading to new active sites that have higher catalytic activities. These *OH boundmore » structures have the highest calculated activity to date.« less

Docking modes of BB-3497 into the PDF active site--a comparison of the pure MM and QM/MM based docking strategies.

PubMed

Kumari, Tripti; Issar, Upasana; Kakkar, Rita

2014-01-01

Peptide deformylase (PDF) has emerged as an important antibacterial drug target. Considerable effort is being directed toward developing peptidic and non-peptidic inhibitors for this metalloprotein. In this work, the known peptidic inhibitor BB-3497 and its various ionization and tautomeric states are evaluated for their inhibition efficiency against PDF using a molecular mechanics (MM) approach as well as a mixed quantum mechanics/molecular mechanics (QM/MM) approach, with an aim to understand the interactions in the binding site. The evaluated Gibbs energies of binding with the mixed QM/MM approach are shown to have the best predictive power. The experimental pose is found to have the most negative Gibbs energy of binding, and also the smallest strain energy. A quantum mechanical evaluation of the active site reveals the requirement of strong chelation by the ligand with the metal ion. The investigated ligand chelates the metal ion through the two oxygens of its reverse hydroxamate moiety, particularly the N-O(-) oxygen, forming strong covalent bonds with the metal ion, which is penta-coordinated. In the uninhibited state, the metal ion is tetrahedrally coordinated, and hence chelation with the inhibitor is associated with an increase of the metal ion coordination. Thus, the strong binding of the ligand at the binding site is accounted for.

Universal dependence of hydrogen oxidation and evolution reaction activity of platinum-group metals on pH and hydrogen binding energy

PubMed Central

Zheng, Jie; Sheng, Wenchao; Zhuang, Zhongbin; Xu, Bingjun; Yan, Yushan

2016-01-01

Understanding how pH affects the activity of hydrogen oxidation reaction (HOR) and hydrogen evolution reaction (HER) is key to developing active, stable, and affordable HOR/HER catalysts for hydroxide exchange membrane fuel cells and electrolyzers. A common linear correlation between hydrogen binding energy (HBE) and pH is observed for four supported platinum-group metal catalysts (Pt/C, Ir/C, Pd/C, and Rh/C) over a broad pH range (0 to 13), suggesting that the pH dependence of HBE is metal-independent. A universal correlation between exchange current density and HBE is also observed on the four metals, indicating that they may share the same elementary steps and rate-determining steps and that the HBE is the dominant descriptor for HOR/HER activities. The onset potential of CO stripping on the four metals decreases with pH, indicating a stronger OH adsorption, which provides evidence against the promoting effect of adsorbed OH on HOR/HER. PMID:27034988

Universal dependence of hydrogen oxidation and evolution reaction activity of platinum-group metals on pH and hydrogen binding energy.

PubMed

Zheng, Jie; Sheng, Wenchao; Zhuang, Zhongbin; Xu, Bingjun; Yan, Yushan

2016-03-01

Understanding how pH affects the activity of hydrogen oxidation reaction (HOR) and hydrogen evolution reaction (HER) is key to developing active, stable, and affordable HOR/HER catalysts for hydroxide exchange membrane fuel cells and electrolyzers. A common linear correlation between hydrogen binding energy (HBE) and pH is observed for four supported platinum-group metal catalysts (Pt/C, Ir/C, Pd/C, and Rh/C) over a broad pH range (0 to 13), suggesting that the pH dependence of HBE is metal-independent. A universal correlation between exchange current density and HBE is also observed on the four metals, indicating that they may share the same elementary steps and rate-determining steps and that the HBE is the dominant descriptor for HOR/HER activities. The onset potential of CO stripping on the four metals decreases with pH, indicating a stronger OH adsorption, which provides evidence against the promoting effect of adsorbed OH on HOR/HER.

Probing the acidic residue within the integrin binding site of laminin-511 that interacts with the metal ion-dependent adhesion site of α6β1 integrin.

PubMed

Taniguchi, Yukimasa; Li, Shaoliang; Takizawa, Mamoru; Oonishi, Eriko; Toga, Junko; Yagi, Emiko; Sekiguchi, Kiyotoshi

2017-06-03

Laminins are major cell-adhesive proteins of basement membranes that interact with integrins in a divalent cation-dependent manner. Laminin-511 consists of α5, β1, and γ1 chains, of which three laminin globular domains of the α5 chain (α5/LG1-3) and a Glu residue in the C-terminal tail of chain γ1 (γ1-Glu1607) are required for binding to integrins. However, it remains unsettled whether the Glu residue in the γ1 tail is involved in integrin binding by coordinating the metal ion in the metal ion-dependent adhesion site of β1 integrin (β1-MIDAS), or by stabilizing the conformation of α5/LG1-3. To address this issue, we examined whether α5/LG1-3 contain an acidic residue required for integrin binding that is as critical as the Glu residue in the γ1 tail; to achieve this, we undertook exhaustive alanine substitutions of the 54 acidic residues present in α5/LG1-3 of the E8 fragment of laminin-511 (LM511E8). Most of the alanine mutants possessed α6β1 integrin binding activities comparable with wild-type LM511E8. Alanine substitution for α5-Asp3198 and Asp3219 caused mild reduction in integrin binding activity, and that for α5-Asp3218 caused severe reduction, possibly resulting from conformational perturbation of α5/LG1-3. When α5-Asp3218 was substituted with asparagine, the resulting mutant possessed significant binding activity to α6β1 integrin, indicating that α5-Asp3218 is not directly involved in integrin binding through coordination with the metal ion in β1-MIDAS. Given that substitution of γ1-Glu1607 with glutamine nullified the binding activity to α6β1 integrin, these results, taken together, support the possibility that the critical acidic residue coordinating the metal ion in β1-MIDAS is Glu1607 in the γ1 tail, but no such residue is present in α5/LG1-3. Copyright © 2017 Elsevier Inc. All rights reserved.

Polymeric brushes as functional templates for immobilizing ribonuclease A: study of binding kinetics and activity.

PubMed

Cullen, Sean P; Liu, Xiaosong; Mandel, Ian C; Himpsel, Franz J; Gopalan, Padma

2008-02-05

The ability to immobilize proteins with high binding capacities on surfaces while maintaining their activity is critical for protein microarrays and other biotechnological applications. We employed poly(acrylic acid) (PAA) brushes as templates to immobilize ribonuclease A (RNase A), which is commonly used to remove RNA from plasmid DNA preparations. The brushes are grown by surface-anchored atom-transfer radical polymerization (ATRP) initiators. RNase A was immobilized by both covalent esterification and a high binding capacity metal-ion complexation method to PAA brushes. The polymer brushes immobilized 30 times more enzyme compared to self-assembled monolayers. As the thickness of the brush increases, the surface density of the RNase A increases monotonically. The immobilization was investigated by ellipsometry, X-ray photoelectron spectroscopy (XPS), thermogravimetric analysis (TGA), and near-edge X-ray absorption fine structure spectroscopy (NEXAFS). The activity of the immobilized RNase A was determined using UV absorbance. As much as 11.0 microg/cm(2) of RNase A was bound to PAA brushes by metal-ion complexation compared to 5.8 microg/cm(2) by covalent immobilization which is 30 and 16 times the estimated mass bound in a monolayer. The calculated diffusion coefficient D was 0.63 x 10(-14) cm(2)/s for metal-ion complexation and 0.71 x 10(-14) cm(2)/s for covalent immobilization. Similar values of D indicate that the binding kinetics is similar, but the thermodynamic equilibrium coverage varies with the binding chemistry. Immobilization kinetics and thermodynamics were characterized by ellipsometry for both methods. A maximum relative activity of 0.70-0.80 was reached between five and nine monolayers of the immobilized enzyme. However, the relative activity for covalent immobilization was greater than that of metal-ion complexation. Covalent esterification resulted in similar temperature dependence as free enzyme, whereas metal-ion complexation showed no temperature dependence indicating a significant change in conformation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Totoritis, Rachel; Duraiswami, Chaya; Taylor, Amy N.

The continual bacterial adaptation to antibiotics creates an ongoing medical need for the development of novel therapeutics. Polypeptide deformylase (PDF) is a highly conserved bacterial enzyme, which is essential for viability. It has previously been shown that PDF inhibitors represent a promising new area for the development of antimicrobial agents, and that many of the best PDF inhibitors demonstrate slow, time-dependent binding. To improve our understanding of the mechanistic origin of this time-dependent inhibition, we examined in detail the kinetics of PDF catalysis and inhibition by several different PDF inhibitors. Varying pH and solvent isotope led to clear changes inmore » time-dependent inhibition parameters, as did inclusion of NaCl, which binds to the active site metal of PDF. Quantitative analysis of these results demonstrated that the observed time dependence arises from slow binding of the inhibitors to the active site metal. However, we also found several metal binding inhibitors that exhibited rapid, non-time-dependent onset of inhibition. By a combination of structural and chemical modification studies, we show that metal binding is only slow when the rest of the inhibitor makes optimal hydrogen bonds within the subsites of PDF. Both of these interactions between the inhibitor and enzyme were found to be necessary to observe time-dependent inhibition, as elimination of either leads to its loss.« less

A structural role for the PHP domain in E. coli DNA polymerase III

PubMed Central

2013-01-01

Background In addition to the core catalytic machinery, bacterial replicative DNA polymerases contain a Polymerase and Histidinol Phosphatase (PHP) domain whose function is not entirely understood. The PHP domains of some bacterial replicases are active metal-dependent nucleases that may play a role in proofreading. In E. coli DNA polymerase III, however, the PHP domain has lost several metal-coordinating residues and is likely to be catalytically inactive. Results Genomic searches show that the loss of metal-coordinating residues in polymerase PHP domains is likely to have coevolved with the presence of a separate proofreading exonuclease that works with the polymerase. Although the E. coli Pol III PHP domain has lost metal-coordinating residues, the structure of the domain has been conserved to a remarkable degree when compared to that of metal-binding PHP domains. This is demonstrated by our ability to restore metal binding with only three point mutations, as confirmed by the metal-bound crystal structure of this mutant determined at 2.9 Å resolution. We also show that Pol III, a large multi-domain protein, unfolds cooperatively and that mutations in the degenerate metal-binding site of the PHP domain decrease the overall stability of Pol III and reduce its activity. Conclusions While the presence of a PHP domain in replicative bacterial polymerases is strictly conserved, its ability to coordinate metals and to perform proofreading exonuclease activity is not, suggesting additional non-enzymatic roles for the domain. Our results show that the PHP domain is a major structural element in Pol III and its integrity modulates both the stability and activity of the polymerase. PMID:23672456

Mechanistic insights into metal ion activation and operator recognition by the ferric uptake regulator

NASA Astrophysics Data System (ADS)

Deng, Zengqin; Wang, Qing; Liu, Zhao; Zhang, Manfeng; Machado, Ana Carolina Dantas; Chiu, Tsu-Pei; Feng, Chong; Zhang, Qi; Yu, Lin; Qi, Lei; Zheng, Jiangge; Wang, Xu; Huo, Xinmei; Qi, Xiaoxuan; Li, Xiaorong; Wu, Wei; Rohs, Remo; Li, Ying; Chen, Zhongzhou

2015-07-01

Ferric uptake regulator (Fur) plays a key role in the iron homeostasis of prokaryotes, such as bacterial pathogens, but the molecular mechanisms and structural basis of Fur-DNA binding remain incompletely understood. Here, we report high-resolution structures of Magnetospirillum gryphiswaldense MSR-1 Fur in four different states: apo-Fur, holo-Fur, the Fur-feoAB1 operator complex and the Fur-Pseudomonas aeruginosa Fur box complex. Apo-Fur is a transition metal ion-independent dimer whose binding induces profound conformational changes and confers DNA-binding ability. Structural characterization, mutagenesis, biochemistry and in vivo data reveal that Fur recognizes DNA by using a combination of base readout through direct contacts in the major groove and shape readout through recognition of the minor-groove electrostatic potential by lysine. The resulting conformational plasticity enables Fur binding to diverse substrates. Our results provide insights into metal ion activation and substrate recognition by Fur that suggest pathways to engineer magnetotactic bacteria and antipathogenic drugs.

A two-metal ion mechanism operates in the hammerhead ribozyme-mediated cleavage of an RNA substrate

PubMed Central

Lott, William B.; Pontius, Brian W.; von Hippel, Peter H.

1998-01-01

Evidence for a two-metal ion mechanism for cleavage of the HH16 hammerhead ribozyme is provided by monitoring the rate of cleavage of the RNA substrate as a function of La3+ concentration in the presence of a constant concentration of Mg2+. We show that a bell-shaped curve of cleavage activation is obtained as La3+ is added in micromolar concentrations in the presence of 8 mM Mg2+, with a maximal rate of cleavage being attained in the presence of 3 μM La3+. These results show that two-metal ion binding sites on the ribozyme regulate the rate of the cleavage reaction and, on the basis of earlier estimates of the Kd values for Mg2+ of 3.5 mM and >50 mM, that these sites bind La3+ with estimated Kd values of 0.9 and >37.5 μM, respectively. Furthermore, given the very different effects of these metal ions at the two binding sites, with displacement of Mg2+ by La3+ at the stronger (relative to Mg2+) binding site activating catalysis and displacement of Mg2+ by La3+ at the weaker (relative to Mg2+) (relative to Mg2+) binding site inhibiting catalysis, we show that the metal ions at these two sites play very different roles. We argue that the metal ion at binding site 1 coordinates the attacking 2′-oxygen species in the reaction and lowers the pKa of the attached proton, thereby increasing the concentration of the attacking alkoxide nucleophile in an equilibrium process. In contrast, the role of the metal ion at binding site 2 is to catalyze the reaction by absorbing the negative charge that accumulates at the leaving 5′-oxygen in the transition state. We suggest structural reasons why the Mg2+–La3+ ion combination is particularly suited to demonstrating these different roles of the two-metal ions in the ribozyme cleavage reaction. PMID:9435228

MamO Is a Repurposed Serine Protease that Promotes Magnetite Biomineralization through Direct Transition Metal Binding in Magnetotactic Bacteria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hershey, David M.; Ren, Xuefeng; Melnyk, Ryan A.

2016-03-16

Many living organisms transform inorganic atoms into highly ordered crystalline materials. An elegant example of such biomineralization processes is the production of nano-scale magnetic crystals in magnetotactic bacteria. Previous studies have implicated the involvement of two putative serine proteases, MamE and MamO, during the early stages of magnetite formation in Magnetospirillum magneticum AMB-1. Here, using genetic analysis and X-ray crystallography, we show that MamO has a degenerate active site, rendering it incapable of protease activity. Instead, MamO promotes magnetosome formation through two genetically distinct, noncatalytic activities: activation of MamE-dependent proteolysis of biomineralization factors and direct binding to transition metal ions.more » By solving the structure of the protease domain bound to a metal ion, we identify a surface-exposed di-histidine motif in MamO that contributes to metal binding and show that it is required to initiate biomineralization in vivo. Finally, we find that pseudoproteases are widespread in magnetotactic bacteria and that they have evolved independently in three separate taxa. In conclusion, our results highlight the versatility of protein scaffolds in accommodating new biochemical activities and provide unprecedented insight into the earliest stages of biomineralization.« less

Comparative study on effects of four energy plants growth on chemical fractions of heavy metals and activity of soil enzymes in copper mine tailings.

PubMed

Zhang, Jie; Yang, Shiyong; Yang, Hongfei; Huang, Yongjie; Zheng, Liming; Yuan, Jing; Zhou, Shoubiao

2018-05-12

Four gramineous energy plants, Miscanthus sacchariflorus, M. floridulus, Phragmites australis, and Arundo donax were grown on copper tailings in the field for four years. Their phytoremediation potential was examined in terms of their effects on the fractions of heavy metals and soil enzyme activities. Results showed that plantation of these four gramineous plants has improved the proportion of organic material (OM)-binding fraction of heavy metals in copper tailings as a whole, and reduced the proportion of exchangeable and residual fractions. In particular, M. sacchariflorus growth improved significantly the proportion of the OM-binding fractions of Cu (1.73 times), Cd (1.71 times), Zn (1.18 times), and Pb (3.14 times) (P < 0.05) and reduced markedly the residual fractions of Cu (64.45%), Cd (82.38%), Zn (61.43%), and Pb (73.41%) (P < 0.05). Except for A. donax, the growth of other three energy plants improved the activity of phosphatase, urease and dehydrogenase in copper tailings to some extent. In particular, the activity of soil phosphatase and urease in planted tailings differed significantly from that of control (P < 0.05). The effect of M. sacchariflorus growth on soil enzyme was the highest, followed by P. australis, M. floridulus, and A. donax. The content of each heavy metal fraction in soil was correlated with soil enzyme activities, especially the content of OM-binding fraction, which correlated significantly with the activities of phosphatase, urease and dehydrogenase in soil. According to the effects of four gramineous plants growth on activity of soil enzymes and fractions of heavy metals, M. sacchariflorus had the optimal effects for phytoremediation. Therefore, M. sacchariflorus was a candidate plant with great potential for the revegetation of heavy metal tailings.

An Iron Reservoir to the Catalytic Metal

PubMed Central

Liu, Fange; Geng, Jiafeng; Gumpper, Ryan H.; Barman, Arghya; Davis, Ian; Ozarowski, Andrew; Hamelberg, Donald; Liu, Aimin

2015-01-01

The rubredoxin motif is present in over 74,000 protein sequences and 2,000 structures, but few have known functions. A secondary, non-catalytic, rubredoxin-like iron site is conserved in 3-hydroxyanthranilate 3,4-dioxygenase (HAO), from single cellular sources but not multicellular sources. Through the population of the two metal binding sites with various metals in bacterial HAO, the structural and functional relationship of the rubredoxin-like site was investigated using kinetic, spectroscopic, crystallographic, and computational approaches. It is shown that the first metal presented preferentially binds to the catalytic site rather than the rubredoxin-like site, which selectively binds iron when the catalytic site is occupied. Furthermore, an iron ion bound to the rubredoxin-like site is readily delivered to an empty catalytic site of metal-free HAO via an intermolecular transfer mechanism. Through the use of metal analysis and catalytic activity measurements, we show that a downstream metabolic intermediate can selectively remove the catalytic iron. As the prokaryotic HAO is often crucial for cell survival, there is a need for ensuring its activity. These results suggest that the rubredoxin-like site is a possible auxiliary iron source to the catalytic center when it is lost during catalysis in a pathway with metabolic intermediates of metal-chelating properties. A spare tire concept is proposed based on this biochemical study, and this concept opens up a potentially new functional paradigm for iron-sulfur centers in iron-dependent enzymes as transient iron binding and shuttling sites to ensure full metal loading of the catalytic site. PMID:25918158

How Native and Alien Metal Cations Bind ATP: Implications for Lithium as a Therapeutic Agent

NASA Astrophysics Data System (ADS)

Dudev, Todor; Grauffel, Cédric; Lim, Carmay

2017-02-01

Adenosine triphosphate (ATP), the major energy currency of the cell, exists in solution mostly as ATP-Mg. Recent experiments suggest that Mg2+ interacts with the highly charged ATP triphosphate group and Li+ can co-bind with the native Mg2+ to form ATP-Mg-Li and modulate the neuronal purine receptor response. However, it is unclear how the negatively charged ATP triphosphate group binds Mg2+ and Li+ (i.e. which phosphate group(s) bind Mg2+/Li+) and how the ATP solution conformation depends on the type of metal cation and the metal-binding mode. Here, we reveal the preferred ATP-binding mode of Mg2+/Li+ alone and combined: Mg2+ prefers to bind ATP tridentately to each of the three phosphate groups, but Li+ prefers to bind bidentately to the terminal two phosphates. We show that the solution ATP conformation depends on the cation and its binding site/mode, but it does not change significantly when Li+ binds to Mg2+-loaded ATP. Hence, ATP-Mg-Li, like Mg2+-ATP, can fit in the ATP-binding site of the host enzyme/receptor, activating specific signaling pathways.

Site Selective Binding of Zn(ll) ot Metallo-b-Lactamase L1 from Stenotrophomonas Maltophilia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Costello,A.; Periyannan, G.; Yang, K.

2006-01-01

Extended X-ray absorption fine structure studies of the metallo-{beta}-lactamase L1 from Stenotrophomonas maltophilia containing 1 and 2 equiv of Zn(II) and containing 2 equiv of Zn(II) plus hydrolyzed nitrocefin are presented. The data indicate that the first, catalytically dominant metal ion is bound by L1 at the consensus Zn1 site. The data further suggest that binding of the first metal helps preorganize the ligands for binding of the second metal ion. The di-Zn enzyme displays a well-defined metal-metal interaction at 3.42 Angstroms. Reaction with the {beta}-lactam antibiotic nitrocefin results in a product-bound species, in which the ring-opened lactam rotates inmore » the active site to present the S1 sulfur atom of nitrocefin to one of the metal ions for coordination. The product bridges the two metal ions, with a concomitant lengthening of the Zn-Zn interaction to 3.62 Angstroms.« less

Uncertainty analysis of the nonideal competitive adsorption-donnan model: effects of dissolved organic matter variability on predicted metal speciation in soil solution.

PubMed

Groenenberg, Jan E; Koopmans, Gerwin F; Comans, Rob N J

2010-02-15

Ion binding models such as the nonideal competitive adsorption-Donnan model (NICA-Donnan) and model VI successfully describe laboratory data of proton and metal binding to purified humic substances (HS). In this study model performance was tested in more complex natural systems. The speciation predicted with the NICA-Donnan model and the associated uncertainty were compared with independent measurements in soil solution extracts, including the free metal ion activity and fulvic (FA) and humic acid (HA) fractions of dissolved organic matter (DOM). Potentially important sources of uncertainty are the DOM composition and the variation in binding properties of HS. HS fractions of DOM in soil solution extracts varied between 14 and 63% and consisted mainly of FA. Moreover, binding parameters optimized for individual FA samples show substantial variation. Monte Carlo simulations show that uncertainties in predicted metal speciation, for metals with a high affinity for FA (Cu, Pb), are largely due to the natural variation in binding properties (i.e., the affinity) of FA. Predictions for metals with a lower affinity (Cd) are more prone to uncertainties in the fraction FA in DOM and the maximum site density (i.e., the capacity) of the FA. Based on these findings, suggestions are provided to reduce uncertainties in model predictions.

Structural and Biochemical Characterization of a Copper-Binding Mutant of the Organomercurial Lyase MerB: Insight into the Key Role of the Active Site Aspartic Acid in Hg-Carbon Bond Cleavage and Metal Binding Specificity.

PubMed

Wahba, Haytham M; Lecoq, Lauriane; Stevenson, Michael; Mansour, Ahmed; Cappadocia, Laurent; Lafrance-Vanasse, Julien; Wilkinson, Kevin J; Sygusch, Jurgen; Wilcox, Dean E; Omichinski, James G

2016-02-23

In bacterial resistance to mercury, the organomercurial lyase (MerB) plays a key role in the detoxification pathway through its ability to cleave Hg-carbon bonds. Two cysteines (C96 and C159; Escherichia coli MerB numbering) and an aspartic acid (D99) have been identified as the key catalytic residues, and these three residues are conserved in all but four known MerB variants, where the aspartic acid is replaced with a serine. To understand the role of the active site serine, we characterized the structure and metal binding properties of an E. coli MerB mutant with a serine substituted for D99 (MerB D99S) as well as one of the native MerB variants containing a serine residue in the active site (Bacillus megaterium MerB2). Surprisingly, the MerB D99S protein copurified with a bound metal that was determined to be Cu(II) from UV-vis absorption, inductively coupled plasma mass spectrometry, nuclear magnetic resonance, and electron paramagnetic resonance studies. X-ray structural studies revealed that the Cu(II) is bound to the active site cysteine residues of MerB D99S, but that it is displaced following the addition of either an organomercurial substrate or an ionic mercury product. In contrast, the B. megaterium MerB2 protein does not copurify with copper, but the structure of the B. megaterium MerB2-Hg complex is highly similar to the structure of the MerB D99S-Hg complexes. These results demonstrate that the active site aspartic acid is crucial for both the enzymatic activity and metal binding specificity of MerB proteins and suggest a possible functional relationship between MerB and its only known structural homologue, the copper-binding protein NosL.

Crystal Structure of the Zorbamycin-Binding Protein ZbmA, the Primary Self-Resistance Element in Streptomyces flavoviridis ATCC21892

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rudolf, Jeffrey D.; Bigelow, Lance; Chang, Changsoo

The bleomycins (BLMs), tallysomycins (TLMs), phleomycin, and zorbamycin (ZBM) are members of the BLM family of glycopeptide-derived antitumor antibiotics. The BLM-producing Streptomyces verticillus ATCC15003 and the TLM-producing Streptoalloteichus hindustanus E465-94 ATCC31158 both possess at least two self-resistance elements, an N-acetyltransferase and a binding protein. The N-acetyltransferase provides resistance by disrupting the metal-binding domain of the antibiotic that is required for activity, while the binding protein confers resistance by sequestering the metal-bound antibiotic and preventing drug activation via molecular oxygen. We recently established that the ZBM producer, Streptomyces flavoviridis ATCC21892, lacks the N-acetyltransferase resistance gene and that the ZBM-binding protein, ZbmA,more » is sufficient to confer resistance in the producing strain. To investigate the resistance mechanism attributed to ZbmA, we determined the crystal structures of apo and Cu(II)-ZBM-bound ZbmA at high resolutions of 1.90 and 1.65 angstrom, respectively. A comparison and contrast with other structurally characterized members of the BLM-binding protein family revealed key differences in the protein ligand binding environment that fine-tunes the ability of ZbmA to sequester metal-bound ZBM and supports drug sequestration as the primary resistance mechanism in the producing organisms of the BLM family of antitumor antibiotics.« less

Histone Deacetylase 8: Characterization of Physiological Divalent Metal Catalysis.

PubMed

Nechay, Michael R; Gallup, Nathan M; Morgenstern, Amanda; Smith, Quentin A; Eberhart, Mark E; Alexandrova, Anastassia N

2016-07-07

Histone deacetylases (HDACs) are responsible for the removal of acetyl groups from histones, resulting in gene silencing. Overexpression of HDACs is associated with cancer, and their inhibitors are of particular interest as chemotherapeutics. However, HDACs remain a target of mechanistic debate. HDAC class 8 is the most studied HDAC, and of particular importance due to its human oncological relevance. HDAC8 has traditionally been considered to be a Zn-dependent enzyme. However, recent experimental assays have challenged this assumption and shown that HDAC8 is catalytically active with a variety of different metals, and that it may be a Fe-dependent enzyme in vivo. We studied two opposing mechanisms utilizing a series of divalent metal ions in physiological abundance (Zn(2+), Fe(2+), Co(2+), Mn(2+), Ni(2+), and Mg(2+)). Extensive sampling of the entire protein with different bound metals was done with the mixed quantum-classical QM/DMD method. Density functional theory (DFT) on an unusually large cluster model was used to describe the active site and reaction mechanism. We have found that the reaction profile of HDAC8 is similar among all metals tested, and follows one of the previously published mechanisms, but the rate-determining step is different from the one previously claimed. We further provide a scheme for estimating the metal binding affinities to the protein. We use the quantum theory of atoms in molecules (QTAIM) to understand the different binding affinities for each metal in HDAC8 as well as the ability of each metal to bind and properly orient the substrate for deacetylation. The combination of this data with the catalytic rate constants is required to reproduce the experimentally observed trend in metal-depending performance. We predict Co(2+) and Zn(2+) to be the most active metals in HDAC8, followed by Fe(2+), and Mn(2+) and Mg(2+) to be the least active.

Structural changes at the metal ion binding site during the phosphoglucomutase reaction.

PubMed

Ray, W J; Post, C B; Liu, Y; Rhyu, G I

1993-01-12

An electron density map of the reactive, Cd2+ form of crystalline phosphoglucomutase from X-ray diffraction studies shows that the enzymic phosphate donates a nonbridging oxygen to the ligand sphere of the bound metal ion, which appears to be tetracoordinate. 31P and 113Cd NMR spectroscopy are used to assess changes in the properties of bound Cd2+ produced by substrate/product and by substrate/product analog inhibitors. The approximately 50 ppm downfield shift of the 113Cd resonance on formation of the complex of dephosphoenzyme and glucose 1,6-bisphosphate is associated with the initial sugar-phosphate binding step and likely involves a change in the geometry of the coordinating ligands. This interpretation is supported by spectral studies involving various complexes of the active Co2+ and Ni(2+)-enzyme. In addition, there is a loss of the 31P-113Cd J coupling that characterizes the monophosphate complexes of the Cd2+ enzyme either during or immediately after the PO3- transfer step that produces the bisphosphate complex, indicating a further change at the metal binding site. The implications of these observations with respect to the PO3- transfer process in the phosphoglucomutase reaction are considered. The apparent plasticity of the ligand sphere of the active site metal ion in this system may allow a single metal ion to act as a chaperone for a nonbridging oxygen during PO3- transfer or to allow a change in metal ion coordination during catalysis. A general NMR line shape/chemical-exchange analysis for evaluating binding in protein-ligand systems when exchange is intermediate to fast on the NMR time scale is described. Its application to the present system involves multiple exchange sites that depend on a single binding rate, thereby adding further constraints to the analysis.

Nickel binding and [NiFe]-hydrogenase maturation by the metallochaperone SlyD with a single metal-binding site in Escherichia coli.

PubMed

Kaluarachchi, Harini; Altenstein, Matthias; Sugumar, Sonia R; Balbach, Jochen; Zamble, Deborah B; Haupt, Caroline

2012-03-16

SlyD (sensitive to lysis D) is a nickel metallochaperone involved in the maturation of [NiFe]-hydrogenases in Escherichia coli (E. coli) and specifically contributes to the nickel delivery step during enzyme biosynthesis. This protein contains a C-terminal metal-binding domain that is rich in potential metal-binding residues that enable SlyD to bind multiple nickel ions with high affinity. The SlyD homolog from Thermus thermophilus does not contain the extended cysteine- and histidine-rich C-terminal tail of the E. coli protein, yet it binds a single Ni(II) ion tightly. To investigate whether a single metal-binding motif can functionally replace the full-length domain, we generated a truncation of E. coli SlyD, SlyD155. Ni(II) binding to SlyD155 was investigated by using isothermal titration calorimetry, NMR and electrospray ionization mass spectrometry measurements. This in vitro characterization revealed that SlyD155 contains a single metal-binding motif with high affinity for nickel. Structural characterization by X-ray absorption spectroscopy and NMR indicated that nickel was coordinated in an octahedral geometry with at least two histidines as ligands. Heterodimerization between SlyD and another hydrogenase accessory protein, HypB, is essential for optimal hydrogenase maturation and was confirmed for SlyD155 via cross-linking experiments and NMR titrations, as were conserved chaperone and peptidyl-prolyl isomerase activities. Although these properties of SlyD are preserved in the truncated version, it does not modulate nickel binding to HypB in vitro or contribute to the maturation of [NiFe]-hydrogenases in vivo, unlike the full-length protein. This study highlights the importance of the unusual metal-binding domain of E. coli SlyD in hydrogenase biogenesis. Copyright © 2012 Elsevier Ltd. All rights reserved.

Biochemical characterization of P-type copper ATPases

PubMed Central

Inesi, Giuseppe; Pilankatta, Rajendra; Tadini-Buoninsegni, Francesco

2014-01-01

Copper ATPases, in analogy with other members of the P-ATPase superfamily, contain a catalytic headpiece including an aspartate residue reacting with ATP to form a phosphoenzyme intermediate, and transmembrane helices containing cation-binding sites [TMBS (transmembrane metal-binding sites)] for catalytic activation and cation translocation. Following phosphoenzyme formation by utilization of ATP, bound copper undergoes displacement from the TMBS to the lumenal membrane surface, with no H+ exchange. Although PII-type ATPases sustain active transport of alkali/alkali-earth ions (i.e. Na+, Ca2+) against electrochemical gradients across defined membranes, PIB-type ATPases transfer transition metal ions (i.e. Cu+) from delivery to acceptor proteins and, prominently in mammalian cells, undergo trafficking from/to various membrane compartments. A specific component of copper ATPases is the NMBD (N-terminal metal-binding domain), containing up to six copper-binding sites in mammalian (ATP7A and ATP7B) enzymes. Copper occupancy of NMBD sites and interaction with the ATPase headpiece are required for catalytic activation. Furthermore, in the presence of copper, the NMBD allows interaction with protein kinase D, yielding phosphorylation of serine residues, ATP7B trafficking and protection from proteasome degradation. A specific feature of ATP7A is glycosylation and stabilization on plasma membranes. Cisplatin, a platinum-containing anti-cancer drug, binds to copper sites of ATP7A and ATP7B, and undergoes vectorial displacement in analogy with copper. PMID:25242165

Characterizing multiple metal ion binding sites within a ribozyme by cadmium-induced EPR silencing

PubMed Central

Kisseleva, Natalia; Kraut, Stefanie; Jäschke, Andres; Schiemann, Olav

2007-01-01

In ribozyme catalysis, metal ions are generally known to make structural and∕or mechanistic contributions. The catalytic activity of a previously described Diels-Alderase ribozyme was found to depend on the concentration of divalent metal ions, and crystallographic data revealed multiple binding sites. Here, we elucidate the interactions of this ribozyme with divalent metal ions in solution using electron paramagnetic resonance (EPR) spectroscopy. Manganese ion titrations revealed five high-affinity Mn2+ binding sites with an upper Kd of 0.6±0.2 μM. In order to characterize each binding site individually, EPR-silent Cd2+ ions were used to saturate the other binding sites. This cadmium-induced EPR silencing showed that the Mn2+ binding sites possess different affinities. In addition, these binding sites could be assigned to three different types, including innersphere, outersphere, and a Mn2+ dimer. Based on simulations, the Mn2+-Mn2+ distance within the dimer was found to be ∼6 Å, which is in good agreement with crystallographic data. The EPR-spectroscopic characterization reveals no structural changes upon addition of a Diels-Alder product, supporting the concept of a preorganized catalytic pocket in the Diels-Alder ribozyme and the structural role of these ions. PMID:19404418

ROLE OF IRON AND MANGANESE OXIDES IN BIOSOLIDS AND BIOSOLIDS-AMENDED SOILS ON METAL BINDING

EPA Science Inventory

Biosolids contain high levels of Fe, Mn, and Al. Surfaces of freshly precipitated metal oxides, especially Fe and Mn, are known to be highly active sites for most dissolved metal ion species. We nw have metal sorption/desorption data that illustrate the importance of Fe and Mn fr...

Template-directed covalent conjugation of DNA to native antibodies, transferrin and other metal-binding proteins

NASA Astrophysics Data System (ADS)

Rosen, Christian B.; Kodal, Anne L. B.; Nielsen, Jesper S.; Schaffert, David H.; Scavenius, Carsten; Okholm, Anders H.; Voigt, Niels V.; Enghild, Jan J.; Kjems, Jørgen; Tørring, Thomas; Gothelf, Kurt V.

2014-09-01

DNA-protein conjugates are important in bioanalytical chemistry, molecular diagnostics and bionanotechnology, as the DNA provides a unique handle to identify, functionalize or otherwise manipulate proteins. To maintain protein activity, conjugation of a single DNA handle to a specific location on the protein is often needed. However, preparing such high-quality site-specific conjugates often requires genetically engineered proteins, which is a laborious and technically challenging approach. Here we demonstrate a simpler method to create site-selective DNA-protein conjugates. Using a guiding DNA strand modified with a metal-binding functionality, we directed a second DNA strand to the vicinity of a metal-binding site of His6-tagged or wild-type metal-binding proteins, such as serotransferrin, where it subsequently reacted with lysine residues at that site. This method, DNA-templated protein conjugation, facilitates the production of site-selective protein conjugates, and also conjugation to IgG1 antibodies via a histidine cluster in the constant domain.

Anthranilate phosphoribosyltransferase: Binding determinants for 5'-phospho-alpha-d-ribosyl-1'-pyrophosphate (PRPP) and the implications for inhibitor design.

PubMed

Evans, Genevieve L; Furkert, Daniel P; Abermil, Nacim; Kundu, Preeti; de Lange, Katrina M; Parker, Emily J; Brimble, Margaret A; Baker, Edward N; Lott, J Shaun

2018-02-01

Phosphoribosyltransferases (PRTs) bind 5'-phospho-α-d-ribosyl-1'-pyrophosphate (PRPP) and transfer its phosphoribosyl group (PRib) to specific nucleophiles. Anthranilate PRT (AnPRT) is a promiscuous PRT that can phosphoribosylate both anthranilate and alternative substrates, and is the only example of a type III PRT. Comparison of the PRPP binding mode in type I, II and III PRTs indicates that AnPRT does not bind PRPP, or nearby metals, in the same conformation as other PRTs. A structure with a stereoisomer of PRPP bound to AnPRT from Mycobacterium tuberculosis (Mtb) suggests a catalytic or post-catalytic state that links PRib movement to metal movement. Crystal structures of Mtb-AnPRT in complex with PRPP and with varying occupancies of the two metal binding sites, complemented by activity assay data, indicate that this type III PRT binds a single metal-coordinated species of PRPP, while an adjacent second metal site can be occupied due to a separate binding event. A series of compounds were synthesized that included a phosphonate group to probe PRPP binding site. Compounds containing a "bianthranilate"-like moiety are inhibitors with IC 50 values of 10-60μM, and K i values of 1.3-15μM. Structures of Mtb-AnPRT in complex with these compounds indicate that their phosphonate moieties are unable to mimic the binding modes of the PRib or pyrophosphate moieties of PRPP. The AnPRT structures presented herein indicated that PRPP binds a surface cleft and becomes enclosed due to re-positioning of two mobile loops. Copyright © 2017 Elsevier B.V. All rights reserved.

The metal-binding domain of wheat heavy metal ATPase 2 (TaHMA2) is involved in zinc/cadmium tolerance and translocation in Arabidopsis.

PubMed

Qiao, Kun; Gong, Liang; Tian, Yanbao; Wang, Hong; Chai, Tuanyao

2018-06-23

Cysteine in the N-terminal metal-binding domain (N-MBD) of TaHMA2 participates in Zn 2+ /Cd 2+ binding and translocation in Arabidopsis. Wheat heavy metal ATPase 2 (TaHMA2) can transport Zn 2+ and Cd 2+ across membranes. A previous study showed that cysteine (Cys) and glutamate residues in the N-terminal metal-binding domain (N-MBD) were necessary for metal-binding and translocation of TaHMA2 in yeast. However, the function of TaHMA2 in plants was not fully revealed. In this study, we investigated the roles of the CCxxE and CPC motifs in the N-MBD and the N/C-terminal regions of TaHMA2 in Zn 2+ /Cd 2+ translocation in root and shoot of Arabidopsis. Compared with the wild type, overexpression of TaHMA2 and the TaHMA2 derivative (glutamic substituted for alanine from CCxxE) in Arabidopsis increased root length, fresh weight and enhanced Zn 2+ /Cd 2+ root-to-shoot translocation. The plants with a truncated N/C-terminal of TaHMA2 were impaired in Zn 2+ /Cd 2+ tolerance and translocation, while mutagenesis of Cys in the N-MBD reduced the tolerance and transport activity of TaHMA2, suggesting the involvement of Cys in Zn 2+ /Cd 2+ binding and translocation in Arabidopsis. This study therefore provides a theoretical possibility for the application of TaHMA2 in transgenic breeding to regulate metal element balance in crop plants.

Heavy metal-binding proteins from metal-stimulated bacteria as a novel adsorbent for metal removal technology.

PubMed

Sano, D; Myojo, K; Omura, T

2006-01-01

Water pollution with toxic heavy metals is of growing concern because heavy metals could bring about serious problems for not only ecosystems in the water environment but also human health. Some metal removal technologies have been in practical use, but much energy and troublesome treatments for chemical wastes are required to operate these conventional technologies. In this study, heavy metal-binding proteins (HMBPs) were obtained from metal-stimulated activated sludge culture with affinity chromatography using copper ion as a ligand. Two-dimensional electrophoresis revealed that a number of proteins in activated sludge culture were recovered as HMBPs for copper ion. N-termini of five HMBPs were determined, and two of them were found to be newly discovered proteins for which no amino acid sequences in protein databases were retrieved at more than 80% identities. Metal-coordinating amino acids occupied 38% of residues in one of the N-terminal sequences of the newly discovered HMBPs. Since these HMBPs were expected to be stable under conditions of water and wastewater treatments, it would be possible to utilize HMBPs as novel adsorbents for heavy metal removal if mass volume of HMBPs can be obtained with protein cloning techniques.

Structural and Biochemical Characterization of Organotin and Organolead Compounds Binding to the Organomercurial Lyase MerB Provide New Insights into Its Mechanism of Carbon–Metal Bond Cleavage

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wahba, Haytham M.; Stevenson, Michael J.; Mansour, Ahmed

2017-01-03

The organomercurial lyase MerB has the unique ability to cleave carbon–Hg bonds, and structural studies indicate that three residues in the active site (C96, D99, and C159 in E. coli MerB) play important roles in the carbon–Hg bond cleavage. However, the role of each residue in carbon–metal bond cleavage has not been well-defined. To do so, we have structurally and biophysically characterized the interaction of MerB with a series of organotin and organolead compounds. Studies with two known inhibitors of MerB, dimethyltin (DMT) and triethyltin (TET), reveal that they inhibit by different mechanisms. In both cases the initial binding ismore » to D99, but DMT subsequently binds to C96, which induces a conformation change in the active site. In contrast, diethyltin (DET) is a substrate for MerB and the SnIV product remains bound in the active site in a coordination similar to that of HgII following cleavage of organomercurial compounds. The results with analogous organolead compounds are similar in that trimethyllead (TML) is not cleaved and binds only to D99, whereas diethyllead (DEL) is a substrate and the PbIV product remains bound in the active site. Binding and cleavage is an exothermic reaction, while binding to D99 has negligible net heat flow. These results show that initial binding of organometallic compounds to MerB occurs at D99 followed, in some cases, by cleavage and loss of the organic moieties and binding of the metal ion product to C96, D99, and C159. The N-terminus of MerA is able to extract the bound PbVI but not the bound SnIV. These results suggest that MerB could be utilized for bioremediation applications, but certain organolead and organotin compounds may present an obstacle by inhibiting the enzyme.« less

[Biological activity of selenorganic compounds at heavy metal salts intoxication].

PubMed

Rusetskaya, N Y; Borodulin, V B

2015-01-01

Possible mechanisms of the antitoxic action of organoselenium compounds in heavy metal poisoning have been considered. Heavy metal toxicity associated with intensification of free radical oxidation, suppression of the antioxidant system, damage to macromolecules, mitochondria and the genetic material can cause apoptotic cell death or the development of carcinogenesis. Organic selenium compounds are effective antioxidants during heavy metal poisoning; they exhibit higher bioavailability in mammals than inorganic ones and they are able to activate antioxidant defense, bind heavy metal ions and reactive oxygen species formed during metal-induced oxidative stress. One of promising organoselenium compounds is diacetophenonyl selenide (DAPS-25), which is characterized by antioxidant and antitoxic activity, under conditions including heavy metal intoxication.

Effects of structural modifications on the metal binding, anti-amyloid activity, and cholinesterase inhibitory activity of chalcones.

PubMed

Fosso, Marina Y; LeVine, Harry; Green, Keith D; Tsodikov, Oleg V; Garneau-Tsodikova, Sylvie

2015-09-28

As the number of individuals affected with Alzheimer's disease (AD) increases and the availability of drugs for AD treatment remains limited, the need to develop effective therapeutics for AD becomes more and more pressing. Strategies currently pursued include inhibiting acetylcholinesterase (AChE) and targeting amyloid-β (Aβ) peptides and metal-Aβ complexes. This work presents the design, synthesis, and biochemical evaluation of a series of chalcones, and assesses the relationship between their structures and their ability to bind metal ions and/or Aβ species, and inhibit AChE/BChE activity. Several chalcones were found to exhibit potent disaggregation of pre-formed N-biotinyl Aβ1-42 (bioAβ42) aggregates in vitro in the absence and presence of Cu(2+)/Zn(2+), while others were effective at inhibiting the action of AChE.

Synthesis of trimethoprim metal complexes: Spectral, electrochemical, thermal, DNA-binding and surface morphology studies.

PubMed

Demirezen, Nihat; Tarınç, Derya; Polat, Duygu; Ceşme, Mustafa; Gölcü, Ayşegül; Tümer, Mehmet

2012-08-01

Complexes of trimethoprim (TMP), with Cu(II), Zn(II), Pt(II), Ru(III) and Fe(III) have been synthesized. Then, these complexes have been characterized by spectroscopic techniques involving UV-vis, IR, mass and (1)H NMR. CHN elemental analysis, electrochemical and thermal behavior of complexes have also been investigated. The electrochemical properties of all complexes have been investigated by cyclic voltammetry (CV) using glassy carbon electrode. The biological activity of the complexes has been evaluated by examining their ability to bind to calf-thymus DNA (CT DNA) with UV spectroscopy and cyclic voltammetry. UV studies of the interaction of the complexes with DNA have shown that these compounds can bind to CT DNA. The binding constants of the complexes with CT DNA have also been calculated. The cyclic voltammograms of the complexes in the presence of CT DNA have shown that the complexes can bind to CT DNA by both the intercalative and the electrostatic binding mode. The antimicrobial activity of these complexes has been evaluated against three Gram-positive and four Gram-negative bacteria. Antifungal activity against two different fungi has been evaluated and compared with the reference drug TMP. Almost all types of complexes show excellent activity against all type of bacteria and fungi. The morphology of the CT DNA, TMP, metal ions and metal complexes has been investigated by scanning electron microscopy (SEM). To get the SEM images, the interaction of compounds with CT DNA has been studied by means of differential pulse voltammetry (DPV) at CT DNA modified pencil graphite electrode (PGE). The decrease in intensity of the guanine oxidation signals has been used as an indicator for the interaction mechanism. Copyright © 2012 Elsevier B.V. All rights reserved.

Structural and biochemical analysis of nuclease domain of clustered regularly interspaced short palindromic repeat (CRISPR)-associated protein 3 (Cas3).

PubMed

Mulepati, Sabin; Bailey, Scott

2011-09-09

RNA transcribed from clustered regularly interspaced short palindromic repeats (CRISPRs) protects many prokaryotes from invasion by foreign DNA such as viruses, conjugative plasmids, and transposable elements. Cas3 (CRISPR-associated protein 3) is essential for this CRISPR protection and is thought to mediate cleavage of the foreign DNA through its N-terminal histidine-aspartate (HD) domain. We report here the 1.8 Å crystal structure of the HD domain of Cas3 from Thermus thermophilus HB8. Structural and biochemical studies predict that this enzyme binds two metal ions at its active site. We also demonstrate that the single-stranded DNA endonuclease activity of this T. thermophilus domain is activated not by magnesium but by transition metal ions such as manganese and nickel. Structure-guided mutagenesis confirms the importance of the metal-binding residues for the nuclease activity and identifies other active site residues. Overall, these results provide a framework for understanding the role of Cas3 in the CRISPR system.

Design and characterization of α-melanotropin peptide analogs cyclized through rhenium and technetium metal coordination

PubMed Central

Giblin, Michael F.; Wang, Nannan; Hoffman, Timothy J.; Jurisson, Silvia S.; Quinn, Thomas P.

1998-01-01

α-Melanocyte stimulating hormone (α-MSH) analogs, cyclized through site-specific rhenium (Re) and technetium (Tc) metal coordination, were structurally characterized and analyzed for their abilities to bind α-MSH receptors present on melanoma cells and in tumor-bearing mice. Results from receptor-binding assays conducted with B16 F1 murine melanoma cells indicated that receptor-binding affinity was reduced to approximately 1% of its original levels after Re incorporation into the cyclic Cys4,10, d-Phe7–α-MSH4-13 analog. Structural analysis of the Re–peptide complex showed that the disulfide bond of the original peptide was replaced by thiolate–metal–thiolate cyclization. A comparison of the metal-bound and metal-free structures indicated that metal complexation dramatically altered the structure of the receptor-binding core sequence. Redesign of the metal binding site resulted in a second-generation Re–peptide complex (ReCCMSH) that displayed a receptor-binding affinity of 2.9 nM, 25-fold higher than the initial Re–α-MSH analog. Characterization of the second-generation Re–peptide complex indicated that the peptide was still cyclized through Re coordination, but the structure of the receptor-binding sequence was no longer constrained. The corresponding 99mTc- and 188ReCCMSH complexes were synthesized and shown to be stable in phosphate-buffered saline and to challenges from diethylenetriaminepentaacetic acid (DTPA) and free cysteine. In vivo, the 99mTcCCMSH complex exhibited significant tumor uptake and retention and was effective in imaging melanoma in a murine-tumor model system. Cyclization of α-MSH analogs via 99mTc and 188Re yields chemically stable and biologically active molecules with potential melanoma-imaging and therapeutic properties. PMID:9788997

Comparative analysis of activator-Eσ54 complexes formed with nucleotide-metal fluoride analogues

PubMed Central

Burrows, Patricia C.; Joly, Nicolas; Nixon, B. Tracy; Buck, Martin

2009-01-01

Bacterial RNA polymerase (RNAP) containing the major variant σ54 factor forms open promoter complexes in a reaction in which specialized activator proteins hydrolyse ATP. Here we probe binding interactions between σ54-RNAP (Eσ54) and the ATPases associated with various cellular activities (AAA+) domain of the Escherichia coli activator protein, PspF, using nucleotide-metal fluoride (BeF and AlF) analogues representing ground and transition states of ATP, which allow complexes (that are otherwise too transient with ATP) to be captured. We show that the organization and functionality of the ADP–BeF- and ADP–AlF-dependent complexes greatly overlap. Our data support an activation pathway in which the initial ATP-dependent binding of the activator to the Eσ54 closed complex results in the re-organization of Eσ54 with respect to the transcription start-site. However, the nucleotide-dependent binding interactions between the activator and the Eσ54 closed complex are in themselves insufficient for forming open promoter complexes when linear double-stranded DNA is present in the initial closed complex. PMID:19553192

Spectroscopic and metal-binding properties of DF3: an artificial protein able to accommodate different metal ions

PubMed Central

Torres Martin de Rosales, Rafael; Faiella, Marina; Farquhar, Erik; Que, Lawrence; Andreozzi, Concetta; Pavone, Vincenzo; Maglio, Ornella; Nastri, Flavia

2010-01-01

The design, synthesis, and metal-binding properties of DF3, a new de novo designed di-iron protein model are described (“DF” represents due ferri, Italian for “two iron,” “di-iron”). DF3 is the latest member of the DF family of synthetic proteins. They consist of helix–loop–helix hairpins, designed to dimerize and form an antiparallel four-helix bundle that encompasses a metal-binding site similar to those of non-heme carboxylate-bridged di-iron proteins. Unlike previous DF proteins, DF3 is highly soluble in water (up to 3 mM) and forms stable complexes with several metal ions (Zn, Co, and Mn), with the desired secondary structure and the expected stoichiometry of two ions per protein. UV–vis studies of Co(II) and Fe(III) complexes confirm a metal-binding environment similar to previous di-Co(II)- and di-Fe(III)-DF proteins, including the presence of a µ-oxo-di-Fe(III) unit. Interestingly, UV–vis, EPR, and resonance Raman studies suggest the interaction of a tyro-sine adjacent to the di-Fe(III) center. The design of DF3 was aimed at increasing the accessibility of small molecules to the active site of the four-helix bundle. Indeed, binding of azide to the di-Fe(III) site demonstrates a more accessible metal site compared with previous DFs. In fact, fitting of the binding curve to the Hill equation allows us to quantify a 150% accessibility enhancement, with respect to DF2. All these results represent a significant step towards the development of a functional synthetic DF metalloprotein. PMID:20225070

A Single Serine Residue Determines Selectivity to Monovalent Metal Ions in Metalloregulators of the MerR Family

PubMed Central

Ibáñez, María M.

2015-01-01

ABSTRACT MerR metalloregulators alleviate toxicity caused by an excess of metal ions, such as copper, zinc, mercury, lead, cadmium, silver, or gold, by triggering the expression of specific efflux or detoxification systems upon metal detection. The sensor protein binds the inducer metal ion by using two conserved cysteine residues at the C-terminal metal-binding loop (MBL). Divalent metal ion sensors, such as MerR and ZntR, require a third cysteine residue, located at the beginning of the dimerization (α5) helix, for metal coordination, while monovalent metal ion sensors, such as CueR and GolS, have a serine residue at this position. This serine residue was proposed to provide hydrophobic and steric restrictions to privilege the binding of monovalent metal ions. Here we show that the presence of alanine at this position does not modify the activation pattern of monovalent metal sensors. In contrast, GolS or CueR mutant sensors with a substitution of cysteine for the serine residue respond to monovalent metal ions or Hg(II) with high sensitivities. Furthermore, in a mutant deleted of the Zn(II) exporter ZntA, they also trigger the expression of their target genes in response to either Zn(II), Cd(II), Pb(II), or Co(II). IMPORTANCE Specificity in a stressor's recognition is essential for mounting an appropriate response. MerR metalloregulators trigger the expression of specific resistance systems upon detection of heavy metal ions. Two groups of these metalloregulators can be distinguished, recognizing either +1 or +2 metal ions, depending on the presence of a conserved serine in the former or a cysteine in the latter. Here we demonstrate that the serine residue in monovalent metal ion sensors excludes divalent metal ion detection, as its replacement by cysteine renders a pan-metal ion sensor. Our results indicate that the spectrum of signals detected by these sensors is determined not only by the metal-binding ligand availability but also by the metal-binding cavity flexibility. PMID:25691529

Tailoring oxide properties: An impact on adsorption characteristics of molecules and metals

NASA Astrophysics Data System (ADS)

Honkala, Karoliina

2014-12-01

Both density functional theory calculations and numerous experimental studies demonstrate a variety of unique features in metal supported oxide films and transition metal doped simple oxides, which are markedly different from their unmodified counterparts. This review highlights, from the computational perspective, recent literature on the properties of the above mentioned surfaces and how they adsorb and activate different species, support metal aggregates, and even catalyse reactions. The adsorption of Au atoms and clusters on metal-supported MgO films are reviewed together with the cluster's theoretically predicted ability to activate and dissociate O2 at the Au-MgO(100)/Ag(100) interface, as well as the impact of an interface vacancy to the binding of an Au atom. In contrast to a bulk MgO surface, an Au atom binds strongly on a metal-supported ultra-thin MgO film and becomes negatively charged. Similarly, Au clusters bind strongly on a supported MgO(100) film and are negatively charged favouring 2D planar structures. The adsorption of other metal atoms is briefly considered and compared to that of Au. Existing computational literature of adsorption and reactivity of simple molecules including O2, CO, NO2, and H2O on mainly metal-supported MgO(100) films is discussed. Chemical reactions such as CO oxidation and O2 dissociation are discussed on the bare thin MgO film and on selected Au clusters supported on MgO(100)/metal surfaces. The Au atoms at the perimeter of the cluster are responsible for catalytic activity and calculations predict that they facilitate dissociative adsorption of oxygen even at ambient conditions. The interaction of H2O with a flat and stepped Ag-supported MgO film is summarized and compared to bulk MgO. The computational results highlight spontaneous dissociation on MgO steps. Furthermore, the impact of water coverage on adsorption and dissociation is addressed. The modifications, such as oxygen vacancies and dopants, at the oxide-metal interface and their effect on the adsorption characteristics of water and Au are summarized. Finally, more limited computational literature on transition metal (TM) doped CaO(100) and MgO(100) surfaces is presented. Again, Au is used as a probe species. Similar to metal-supported MgO films, Au binds more strongly than on undoped CaO(100) and becomes negatively charged. The discussion focuses on rationalization of Au adsorption with the help of Born-Haber cycle, which reveals that the so-called redox energy including the electron transfer from the dopant to the Au atom together with the simultaneous structural relaxation of lattice atoms is responsible for enhanced binding. In addition, adsorption energy dependence on the position and type of the dopant is summarized.

Sodium and Potassium Ions in Proteins and Enzyme Catalysis.

PubMed

Vašák, Milan; Schnabl, Joachim

2016-01-01

The group I alkali metal ions Na(+) and K(+) are ubiquitous components of biological fluids that surround biological macromolecules. They play important roles other than being nonspecific ionic buffering agents or mediators of solute exchange and transport. Molecular evolution and regulated high intracellular and extracellular M(+) concentrations led to incorporation of selective Na(+) and K(+) binding sites into enzymes to stabilize catalytic intermediates or to provide optimal positioning of substrates. The mechanism of M(+) activation, as derived from kinetic studies along with structural analysis, has led to the classification of cofactor-like (type I) or allosteric effector (type II) activated enzymes. In the type I mechanism substrate anchoring to the enzyme active site is mediated by M(+), often acting in tandem with a divalent cation like Mg(2+), Mn(2+) or Zn(2+). In the allosteric type II mechanism, M(+) binding enhances enzyme activity through conformational transitions triggered upon binding to a distant site. In this chapter, following the discussion of the coordination chemistry of Na(+) and K(+) ions and the structural features responsible for the metal binding site selectivity in M(+)-activated enzymes, well-defined examples of M(+)-activated enzymes are used to illustrate the structural basis for type I and type II activation by Na(+) and K(+).

Transition Metal Intercalators as Anticancer Agents—Recent Advances

PubMed Central

Deo, Krishant M.; Pages, Benjamin J.; Ang, Dale L.; Gordon, Christopher P.; Aldrich-Wright, Janice R.

2016-01-01

The diverse anticancer utility of cisplatin has stimulated significant interest in the development of additional platinum-based therapies, resulting in several analogues receiving clinical approval worldwide. However, due to structural and mechanistic similarities, the effectiveness of platinum-based therapies is countered by severe side-effects, narrow spectrum of activity and the development of resistance. Nonetheless, metal complexes offer unique characteristics and exceptional versatility, with the ability to alter their pharmacology through facile modifications of geometry and coordination number. This has prompted the search for metal-based complexes with distinctly different structural motifs and non-covalent modes of binding with a primary aim of circumventing current clinical limitations. This review discusses recent advances in platinum and other transition metal-based complexes with mechanisms of action involving intercalation. This mode of DNA binding is distinct from cisplatin and its derivatives. The metals focused on in this review include Pt, Ru and Cu along with examples of Au, Ni, Zn and Fe complexes; these complexes are capable of DNA intercalation and are highly biologically active. PMID:27809241

Chemical rescue of the post-translationally carboxylated lysine mutant of allantoinase and dihydroorotase by metal ions and short-chain carboxylic acids.

PubMed

Ho, Ya-Yeh; Huang, Yen-Hua; Huang, Cheng-Yang

2013-04-01

Bacterial allantoinase (ALLase) and dihydroorotase (DHOase) are members of the cyclic amidohydrolase family. ALLase and DHOase possess similar binuclear metal centers in the active site in which two metals are bridged by a post-translationally carboxylated lysine. In this study, we determined the effects of carboxylated lysine and metal binding on the activities of ALLase and DHOase. Although DHOase is a metalloenzyme, purified DHOase showed high activity without additional metal supplementation in a reaction mixture or bacterial culture. However, unlike DHOase, ALLase had no activity unless some specific metal ions were added to the reaction mixture or culture. Substituting the metal binding sites H59, H61, K146, H186, H242, or D315 with alanine completely abolished the activity of ALLase. However, the K146C, K146D and K146E mutants of ALLase were still active with about 1-6% activity of the wild-type enzyme. These ALLase K146 mutants were found to have 1.4-1.7 mol metal per mole enzyme subunit, which may indicate that they still contained the binuclear metal center in the active site. The activity of the K146A mutant of the ALLase and the K103A mutant of DHOase can be chemically rescued by short-chain carboxylic acids, such as acetic, propionic, and butyric acids, but not by ethanol, propan-1-ol, and imidazole, in the presence of Co2+ or Mn2+ ions. However, the activity was still ~10-fold less than that of wild-type ALLase. Overall, these results indicated that the 20 natural basic amino acid residues were not sufficiently able to play the role of lysine. Accordingly, we proposed that during evolution, the post-translational modification of carboxylated lysine in the cyclic amidohydrolase family was selected for promoting binuclear metal center self-assembly and increasing the nucleophilicity of the hydroxide at the active site for enzyme catalysis. This kind of chemical rescue combined with site-directed mutagenesis may also be used to identify a binuclear metal center in the active site for other metalloenzymes.

A scale of metal ion binding strengths correlating with ionic charge, Pauling electronegativity, toxicity, and other physiological effects.

PubMed

Kinraide, Thomas B; Yermiyahu, Uri

2007-09-01

Equilibrium constants for binding to plant plasma membranes have been reported for several metal ions, based upon adsorption studies and zeta-potential measurements. LogK values for the ions are these: Al(3+), 4.30; La(3+), 3.34; Cu(2+), 2.60; Ca(2+) and Mg(2+), 1.48; Na(+) and K(+), 0 M(-1). These values correlate well with logK values for ion binding to many organic and inorganic ligands. LogK values for metal ion binding to 12 ligands were normalized and averaged to produce a scale for the binding of 49 ions. The scale correlates well with the values presented above (R(2)=0.998) and with ion binding to cell walls and other biomass. The scale is closely related to the charge (Z) and Pauling electronegativity (PE) of 48 ions (all but Hg(2+)); R(2)=0.969 for the equation (Scale values)=-1.68+Z(1.22+0.444PE). Minimum rhizotoxicity of metal ions appears to be determined by binding strengths: log a(PM,M)=1.60-2.41exp[0.238(Scale values)] determines the value of ion activities at the plasma membrane surface (a(PM,M)) that will ensure inhibition of root elongation. Additional toxicity appears to be related to softness, accounting for the great toxicity of Ag(+), for example. These binding-strength values correlate with additional physiological effects and are suitable for the computation of cell-surface electrical potentials.

Application of a mixed metal oxide catalyst to a metallic substrate

NASA Technical Reports Server (NTRS)

Sevener, Kathleen M. (Inventor); Lohner, Kevin A. (Inventor); Mays, Jeffrey A. (Inventor); Wisner, Daniel L. (Inventor)

2009-01-01

A method for applying a mixed metal oxide catalyst to a metallic substrate for the creation of a robust, high temperature catalyst system for use in decomposing propellants, particularly hydrogen peroxide propellants, for use in propulsion systems. The method begins by forming a prepared substrate material consisting of a metallic inner substrate and a bound layer of a noble metal intermediate. Alternatively, a bound ceramic coating, or frit, may be introduced between the metallic inner substrate and noble metal intermediate when the metallic substrate is oxidation resistant. A high-activity catalyst slurry is applied to the surface of the prepared substrate and dried to remove the organic solvent. The catalyst layer is then heat treated to bind the catalyst layer to the surface. The bound catalyst layer is then activated using an activation treatment and calcinations to form the high-activity catalyst system.

Synthesis, structure, DNA/BSA binding and antibacterial studies of NNO tridentate Schiff base metal complexes

NASA Astrophysics Data System (ADS)

Sakthi, Marimuthu; Ramu, Andy

2017-12-01

A new salicylaldehyde derived 2,4-diiodo-6-((2-phenylaminoethylimino)methyl)phenol Schiff base(L) and its transition metal complexes of the type MLCl where, M = Cu(II), Ni(II), Co(II), Mn(II) and Zn(II) have been synthesized. The coordination mode of Schiff base holding NNO donor atoms with metal ions was well investigated by elemental analysis, ESI-mass as well as IR, UV-vis, CV and NMR spectral studies. The binding efficiency and mode of these complexes with biological macromolecules viz., herring sperm DNA (HS- DNA) and bovine serum albumin (BSA) have been explored through various spectroscopic techniques. The characteristic changes in absorption, emission and, circular dichroism spectra of the complexes with DNA indicate the noticeable interaction between them. From the all spectral information complexes could interact with DNA via non-intercalation mode of binding. The hyperchromisim in absorption band and hypochromisim in emission intensity of BSA with different complex concentrations shown significant information, and the binding affinity value has been predicted from Stern-Volmer plots. Further, all the complexes could cleave the circular plasmid pUC19 DNA efficiently by using an activator H2O2. The ligand and all metal(II) complexes showed good antibacterial activities. The molecular docking studies of the complexes with DNA were performed in order to make a comparison and conclusion with spectral technic results.

Studies on DNA binding behaviour of biologically active transition metal complexes of new tetradentate N2O2 donor Schiff bases: inhibitory activity against bacteria.

PubMed

Sobha, S; Mahalakshmi, R; Raman, N

2012-06-15

A series of Cu(II), Ni(II) and Zn(II) complexes of the type ML have been synthesized with Schiff bases derived from o-acetoacetotoluidide, 2-hydroxybenzaldehyde and o-phenylenediamine/1,4-diaminobutane. The complexes are insoluble in common organic solvents but soluble in DMF and DMSO. The measured molar conductance values in DMSO indicate that the complexes are non-electrolytic in nature. All the six metal complexes have been fully characterized with the help of elemental analyses, molecular weights, molar conductance values, magnetic moments and spectroscopic data. The analytical data helped to elucidate the structure of the metal complexes. The Schiff bases are found to act as tetradentate ligands using N(2)O(2) donor set of atoms leading to a square-planar geometry for the complexes around all the metal ions. The binding properties of metal complexes with DNA were investigated by absorption spectra, viscosity measurements and cyclic voltammetry. Detailed analysis reveals that the metal complexes intercalate into the DNA base stack as intercalators. All the metal complexes cleave the pUC19 DNA in presence of H(2)O(2.) The Schiff bases and their complexes have been screened for their antibacterial activity against five bacterial strains (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus epidermidis, Klebsiella pneumoniae) by disk diffusion method. All the metal complexes have potent biocidal activity than the free ligands. Copyright © 2012 Elsevier B.V. All rights reserved.

Molecular Mechanism of Action for Allosteric Modulators and Agonists in CC-chemokine Receptor 5 (CCR5).

PubMed

Karlshøj, Stefanie; Amarandi, Roxana Maria; Larsen, Olav; Daugvilaite, Viktorija; Steen, Anne; Brvar, Matjaž; Pui, Aurel; Frimurer, Thomas Michael; Ulven, Trond; Rosenkilde, Mette Marie

2016-12-23

The small molecule metal ion chelators bipyridine and terpyridine complexed with Zn 2+ (ZnBip and ZnTerp) act as CCR5 agonists and strong positive allosteric modulators of CCL3 binding to CCR5, weak modulators of CCL4 binding, and competitors for CCL5 binding. Here we describe their binding site using computational modeling, binding, and functional studies on WT and mutated CCR5. The metal ion Zn 2+ is anchored to the chemokine receptor-conserved Glu-283 VII:06/7.39 Both chelators interact with aromatic residues in the transmembrane receptor domain. The additional pyridine ring of ZnTerp binds deeply in the major binding pocket and, in contrast to ZnBip, interacts directly with the Trp-248 VI:13/6.48 microswitch, contributing to its 8-fold higher potency. The impact of Trp-248 was further confirmed by ZnClTerp, a chloro-substituted version of ZnTerp that showed no inherent agonism but maintained positive allosteric modulation of CCL3 binding. Despite a similar overall binding mode of all three metal ion chelator complexes, the pyridine ring of ZnClTerp blocks the conformational switch of Trp-248 required for receptor activation, thereby explaining its lack of activity. Importantly, ZnClTerp becomes agonist to the same extent as ZnTerp upon Ala mutation of Ile-116 III:16/3.40 , a residue that constrains the Trp-248 microswitch in its inactive conformation. Binding studies with 125 I-CCL3 revealed an allosteric interface between the chemokine and the small molecule binding site, including residues Tyr-37 I:07/1.39 , Trp-86 II:20/2.60 , and Phe-109 III:09/3.33 The small molecules and CCL3 approach this interface from opposite directions, with some residues being mutually exploited. This study provides new insight into the molecular mechanism of CCR5 activation and paves the way for future allosteric drugs for chemokine receptors. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Novel bis-(−)-nor-meptazinol derivatives act as dual binding site AChE inhibitors with metal-complexing property

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zheng, Wei; NPFPC Key Laboratory of Contraceptives and Devices, Shanghai Institute of Planned Parenthood Research, 2140 Xietu Road, Shanghai 200032; Li, Juan

The strategy of dual binding site acetylcholinesterase (AChE) inhibition along with metal chelation may represent a promising direction for multi-targeted interventions in the pathophysiological processes of Alzheimer's disease (AD). In the present study, two derivatives (ZLA and ZLB) of a potent dual binding site AChE inhibitor bis-(−)-nor-meptazinol (bis-MEP) were designed and synthesized by introducing metal chelating pharmacophores into the middle chain of bis-MEP. They could inhibit human AChE activity with IC{sub 50} values of 9.63 μM (for ZLA) and 8.64 μM (for ZLB), and prevent AChE-induced amyloid-β (Aβ) aggregation with IC{sub 50} values of 49.1 μM (for ZLA) and 55.3more » μM (for ZLB). In parallel, molecular docking analysis showed that they are capable of interacting with both the catalytic and peripheral anionic sites of AChE. Furthermore, they exhibited abilities to complex metal ions such as Cu(II) and Zn(II), and inhibit Aβ aggregation triggered by these metals. Collectively, these results suggest that ZLA and ZLB may act as dual binding site AChEIs with metal-chelating potency, and may be potential leads of value for further study on disease-modifying treatment of AD. -- Highlights: ► Two novel bis-(−)-nor-meptazinol derivatives are designed and synthesized. ► ZLA and ZLB may act as dual binding site AChEIs with metal-chelating potency. ► They are potential leads for disease-modifying treatment of Alzheimer's disease.« less

Metal binding stoichiometry and isotherm choice in biosorption

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schiewer, S.; Wong, M.H.

1999-11-01

Seaweeds that possess a high metal binding capacity may be used as biosorbents for the removal of toxic heavy metals from wastewater. The binding of Cu and Ni by three brown algae (Sargassum, Colpomenia, Petalonia) and one green alga (Ulva) was investigated at pH 4.0 and pH 3.0. The greater binding strength of Cu is reflected in a binding constant that is about 10 times as high as that of Ni. The extent of metal binding followed the order Petalonia {approximately} Sargassum > Colpomenia > Ulva. This was caused by a decreasing number of binding sites and by much lowermore » metal binding constants for Ulva as compared to the brown algae. Three different stoichiometric assumptions are compared for describing the metal binding, which assume either that each metal ion M binds to one binding site B forming a BM complex or that a divalent metal ion M binds to two monovalent sites B forming BM{sub 0.5} or B{sub 2}M complexes, respectively. Stoichiometry plots are proposed as tools to discern the relevant binding stoichiometry. The pH effect in metal binding and the change in proton binding were well predicted for the B{sub 2}M or BM{sub 0.5} stoichiometries with the former being better for Cu and the latter preferable for Ni. Overall, the BM{sub 0.5} model is recommended because it avoids iterations.« less

Structure-function analyses of metal-binding sites of HypA reveal residues important for hydrogenase maturation in Helicobacter pylori.

PubMed

Blum, Faith C; Hu, Heidi Q; Servetas, Stephanie L; Benoit, Stéphane L; Maier, Robert J; Maroney, Michael J; Merrell, D Scott

2017-01-01

The nickel-containing enzymes of Helicobacter pylori, urease and hydrogenase, are essential for efficient colonization in the human stomach. The insertion of nickel into urease and hydrogenase is mediated by the accessory protein HypA. HypA contains an N-terminal nickel-binding site and a dynamic structural zinc-binding site. The coordination of nickel and zinc within HypA is known to be critical for urease maturation and activity. Herein, we test the hydrogenase activity of a panel of H. pylori mutant strains containing point mutations within the nickel- and zinc-binding sites. We found that the residues that are important for hydrogenase activity are those that were similarly vital for urease activity. Thus, the zinc and metal coordination sites of HypA play similar roles in urease and hydrogenase maturation. In other pathogenic bacteria, deletion of hydrogenase leads to a loss in acid resistance. Thus, the acid resistance of two strains of H. pylori containing a hydrogenase deletion was also tested. These mutant strains demonstrated wild-type levels of acid resistance, suggesting that in H. pylori, hydrogenase does not play a role in acid resistance.

Structure-function analyses of metal-binding sites of HypA reveal residues important for hydrogenase maturation in Helicobacter pylori

PubMed Central

Servetas, Stephanie L.; Benoit, Stéphane L.; Maier, Robert J.; Maroney, Michael J.

2017-01-01

The nickel-containing enzymes of Helicobacter pylori, urease and hydrogenase, are essential for efficient colonization in the human stomach. The insertion of nickel into urease and hydrogenase is mediated by the accessory protein HypA. HypA contains an N-terminal nickel-binding site and a dynamic structural zinc-binding site. The coordination of nickel and zinc within HypA is known to be critical for urease maturation and activity. Herein, we test the hydrogenase activity of a panel of H. pylori mutant strains containing point mutations within the nickel- and zinc-binding sites. We found that the residues that are important for hydrogenase activity are those that were similarly vital for urease activity. Thus, the zinc and metal coordination sites of HypA play similar roles in urease and hydrogenase maturation. In other pathogenic bacteria, deletion of hydrogenase leads to a loss in acid resistance. Thus, the acid resistance of two strains of H. pylori containing a hydrogenase deletion was also tested. These mutant strains demonstrated wild-type levels of acid resistance, suggesting that in H. pylori, hydrogenase does not play a role in acid resistance. PMID:28809946

Selection of peptides binding to metallic borides by screening M13 phage display libraries.

PubMed

Ploss, Martin; Facey, Sandra J; Bruhn, Carina; Zemel, Limor; Hofmann, Kathrin; Stark, Robert W; Albert, Barbara; Hauer, Bernhard

2014-02-10

Metal borides are a class of inorganic solids that is much less known and investigated than for example metal oxides or intermetallics. At the same time it is a highly versatile and interesting class of compounds in terms of physical and chemical properties, like semiconductivity, ferromagnetism, or catalytic activity. This makes these substances attractive for the generation of new materials. Very little is known about the interaction between organic materials and borides. To generate nanostructured and composite materials which consist of metal borides and organic modifiers it is necessary to develop new synthetic strategies. Phage peptide display libraries are commonly used to select peptides that bind specifically to metals, metal oxides, and semiconductors. Further, these binding peptides can serve as templates to control the nucleation and growth of inorganic nanoparticles. Additionally, the combination of two different binding motifs into a single bifunctional phage could be useful for the generation of new composite materials. In this study, we have identified a unique set of sequences that bind to amorphous and crystalline nickel boride (Ni3B) nanoparticles, from a random peptide library using the phage display technique. Using this technique, strong binders were identified that are selective for nickel boride. Sequence analysis of the peptides revealed that the sequences exhibit similar, yet subtle different patterns of amino acid usage. Although a predominant binding motif was not observed, certain charged amino acids emerged as essential in specific binding to both substrates. The 7-mer peptide sequence LGFREKE, isolated on amorphous Ni3B emerged as the best binder for both substrates. Fluorescence microscopy and atomic force microscopy confirmed the specific binding affinity of LGFREKE expressing phage to amorphous and crystalline Ni3B nanoparticles. This study is, to our knowledge, the first to identify peptides that bind specifically to amorphous and to crystalline Ni3B nanoparticles. We think that the identified strong binding sequences described here could potentially serve for the utilisation of M13 phage as a viable alternative to other methods to create tailor-made boride composite materials or new catalytic surfaces by a biologically driven nano-assembly synthesis and structuring.

Identification and characterization of a novel high affinity metal-binding site in the hammerhead ribozyme.

PubMed Central

Hansen, M R; Simorre, J P; Hanson, P; Mokler, V; Bellon, L; Beigelman, L; Pardi, A

1999-01-01

A novel metal-binding site has been identified in the hammerhead ribozyme by 31P NMR. The metal-binding site is associated with the A13 phosphate in the catalytic core of the hammerhead ribozyme and is distinct from any previously identified metal-binding sites. 31P NMR spectroscopy was used to measure the metal-binding affinity for this site and leads to an apparent dissociation constant of 250-570 microM at 25 degrees C for binding of a single Mg2+ ion. The NMR data also show evidence of a structural change at this site upon metal binding and these results are compared with previous data on metal-induced structural changes in the core of the hammerhead ribozyme. These NMR data were combined with the X-ray structure of the hammerhead ribozyme (Pley HW, Flaherty KM, McKay DB. 1994. Nature 372:68-74) to model RNA ligands involved in binding the metal at this A13 site. In this model, the A13 metal-binding site is structurally similar to the previously identified A(g) metal-binding site and illustrates the symmetrical nature of the tandem G x A base pairs in domain 2 of the hammerhead ribozyme. These results demonstrate that 31P NMR represents an important method for both identification and characterization of metal-binding sites in nucleic acids. PMID:10445883

The transcription factor MTF-1 is essential for basal and heavy metal-induced metallothionein gene expression.

PubMed

Heuchel, R; Radtke, F; Georgiev, O; Stark, G; Aguet, M; Schaffner, W

1994-06-15

We have described and cloned previously a factor (MTF-1) that binds specifically to heavy metal-responsive DNA sequence elements in the enhancer/promoter region of metallothionein genes. MTF-1 is a protein of 72.5 kDa that contains six zinc fingers and multiple domains for transcriptional activation. Here we report the disruption of both alleles of the MTF-1 gene in mouse embryonic stem cells by homologous recombination. The resulting null mutant cell line fails to produce detectable amounts of MTF-1. Moreover, due to the loss of MTF-1, the endogenous metallothionein I and II genes are silent, indicating that MTF-1 is required for both their basal and zinc-induced transcription. In addition to zinc, other heavy metals, including cadmium, copper, nickel and lead, also fail to activate metal-responsive promoters in null mutant cells. However, cotransfection of an MTF-1 expression vector and metal-responsive reporter genes yields strong basal transcription that can be further boosted by zinc treatment of cells. These results demonstrate that MTF-1 is essential for metallothionein gene regulation. Finally, we present evidence that MTF-1 itself is a zinc sensor, which exhibits increased DNA binding activity upon zinc treatment.

The relative influence of metal ion binding sites in the I-like domain and the interface with the hybrid domain on rolling and firm adhesion by integrin alpha4beta7.

PubMed

Chen, JianFeng; Takagi, Junichi; Xie, Can; Xiao, Tsan; Luo, Bing-Hao; Springer, Timothy A

2004-12-31

We examined the effect of conformational change at the beta(7) I-like/hybrid domain interface on regulating the transition between rolling and firm adhesion by integrin alpha(4)beta(7). An N-glycosylation site was introduced into the I-like/hybrid domain interface to act as a wedge and to stabilize the open conformation of this interface and hence the open conformation of the alpha(4) beta(7) headpiece. Wild-type alpha(4)beta(7) mediates rolling adhesion in Ca(2+) and Ca(2+)/Mg(2+) but firm adhesion in Mg(2+) and Mn(2+). Stabilizing the open headpiece resulted in firm adhesion in all divalent cations. The interaction between metal binding sites in the I-like domain and the interface with the hybrid domain was examined in double mutants. Changes at these two sites can either counterbalance one another or be additive, emphasizing mutuality and the importance of multiple interfaces in integrin regulation. A double mutant with counterbalancing deactivating ligand-induced metal ion binding site (LIMBS) and activating wedge mutations could still be activated by Mn(2+), confirming the importance of the adjacent to metal ion-dependent adhesion site (ADMIDAS) in integrin activation by Mn(2+). Overall, the results demonstrate the importance of headpiece allostery in the conversion of rolling to firm adhesion.

Relative association of Rubisco with manganese and magnesium as a regulatory mechanism in plants.

PubMed

Bloom, Arnold J; Kameritsch, Petra

2017-12-01

Rubisco, the enzyme that constitutes as much as half of the protein in a leaf, initiates either the photorespiratory pathway that supplies reductant for the assimilation of nitrate into amino acids or the C3 carbon fixation pathway that generates carbohydrates. The relative rates of these two pathways depend both on the relative extent to which O 2 and CO 2 occupies the active site of Rubisco and on whether manganese or magnesium is bound to the enzyme. This study quantified the activities of manganese and magnesium in isolated tobacco chloroplasts and the thermodynamics of binding of these metals to Rubisco purified from tobacco or a bacterium. In tobacco chloroplasts, manganese was less active than magnesium, but Rubisco purified from tobacco had a higher affinity for manganese. The activity of each metal in the chloroplast was similar in magnitude to the affinity of tobacco Rubisco for each. This indicates that, in tobacco chloroplasts, Rubisco associates almost equally with both metals and rapidly exchanges one metal for the other. Binding of magnesium was similar in Rubisco from tobacco and a bacterium, whereas binding of manganese differed greatly between the Rubisco from these species. Moreover, the ratio of leaf manganese to magnesium in C3 plants increased as atmospheric CO 2 increased. These results suggest that Rubisco has evolved to improve the energy transfers between photorespiration and nitrate assimilation and that plants regulate manganese and magnesium activities in the chloroplast to mitigate detrimental changes in their nitrogen/carbon balance as atmospheric CO 2 varies. © 2017 Scandinavian Plant Physiology Society.

An all sulfur analogue of the smallest subunit of F420-non-reducing hydrogenase from Methanococcus voltae--metal binding and structure.

PubMed

Pfeiffer, M; Klein, A; Steinert, P; Schomburg, D

The 25 amino acid long subunit VhuU of the F420-non-reducing hydrogenase from Methanococcus voltae contains selenocysteine within the consensus sequence of known [NiFe] hydrogenases DP(C or U)CxxCxxH (U = selenocysteine). The sulfur-analogue VhuUc was chemically synthesized, purified and its metal binding capability, the catalytic properties, and structural features were investigated. The polypeptide was able to bind nickel, but did not catalyse the heterolytic activation of H2. 2D-NMR spectroscopy revealed an alpha-helical secondary structure for the 15 N-terminal amino acids in 50% TFE. Nickel only binds to the C-terminus, which contains the conserved amino acid motif. Structures derived from the NMR data are compatible with the participation of both sulfur atoms from the conserved cysteine residues in a metal ion binding. Structures obtained from the data sets for Ni.VhuUc as well as Zn.VhuUc showed no further ligands. The informational value for Ni.VhuUc was low due to paramagnetism.

DISTINCT ROLES OF β1 MIDAS, ADMIDAS AND LIMBS CATION-BINDING SITES IN LIGAND RECOGNITION BY INTEGRIN α2β1*

PubMed Central

Valdramidou, Dimitra; Humphries, Martin J.; Mould, A. Paul

2012-01-01

Integrin-ligand interactions are regulated in a complex manner by divalent cations, and previous studies have identified ligand-competent, stimulatory, and inhibitory cation-binding sites. In collagen-binding integrins, such as α2β1, ligand recognition takes place exclusively at the α subunit I domain. However, activation of the αI domain depends on its interaction with a structurally similar domain in the β subunit known as the I-like or βI domain. The top face of the βI domain contains three cation-binding sites: the metal-ion dependent adhesion site (MIDAS), the ADMIDAS (adjacent to MIDAS) and LIMBS (ligand-associated metal binding site). The role of these sites in controlling ligand binding to the αI domain has yet to be elucidated. Mutation of the MIDAS or LIMBS completely blocked collagen binding to α2β1; in contrast mutation of the ADMIDAS reduced ligand recognition but this effect could be overcome by the activating mAb TS2/16. Hence, the MIDAS and LIMBS appear to be essential for the interaction between αI and βI whereas occupancy of the ADMIDAS has an allosteric effect on the conformation of βI. An activating mutation in the α2 I domain partially restored ligand binding to the MIDAS and LIMBS mutants. Analysis of the effects of Ca2+, Mg2+ and Mn2+ on ligand binding to these mutants showed that the MIDAS is a ligand-competent site through which Mn2+ stimulates ligand binding, whereas the LIMBS is a stimulatory Ca2+-binding site, occupancy of which increases the affinity of Mg2+ for the MIDAS. PMID:18820259

Equilibrium binding behavior of magnesium to wall teichoic acid.

PubMed

Thomas, Kieth J; Rice, Charles V

2015-10-01

Peptidoglycan and teichoic acids are the major cell wall components of Gram-positive bacteria that obtain and sequester metal ions required for biochemical processes. The delivery of metals to the cytoplasmic membrane is aided by anionic binding sites within the peptidoglycan and along the phosphodiester polymer of teichoic acid. The interaction with metals is a delicate balance between the need for attraction and ion diffusion to the membrane. Likewise, metal chelation from the extracellular fluid must initially have strong binding energetics that weaken within the cell wall to enable ion release. We employed atomic absorption and equilibrium dialysis to measure the metal binding capacity and metal binding affinity of wall teichoic acid and Mg2+. Data show that Mg2+ binds to WTA with a 1:2Mg2+ to phosphate ratio with a binding capacity of 1.27 μmol/mg. The affinity of Mg2+ to WTA was also found to be 41×10(3) M(-1) at low metal concentrations and 1.3×10(3) M(-1) at higher Mg2+ concentrations due to weakening electrostatic effects. These values are lower than the values describing Mg2+ interactions with peptidoglycan. However, the binding capacity of WTA is 4 times larger than peptidoglycan. External WTA initially binds metals with positive cooperativity, but metal binding switches to negative cooperativity, whereas interior WTA binds metals with only negative cooperativity. The relevance of this work is to describe changes in metal binding behavior depending on environment. When metals are sparse, chelation is strong to ensure survival yet the binding weakens when essential minerals are abundant. Copyright © 2015 Elsevier B.V. All rights reserved.

Selective class IIa histone deacetylase inhibition via a nonchelating zinc-binding group.

PubMed

Lobera, Mercedes; Madauss, Kevin P; Pohlhaus, Denise T; Wright, Quentin G; Trocha, Mark; Schmidt, Darby R; Baloglu, Erkan; Trump, Ryan P; Head, Martha S; Hofmann, Glenn A; Murray-Thompson, Monique; Schwartz, Benjamin; Chakravorty, Subhas; Wu, Zining; Mander, Palwinder K; Kruidenier, Laurens; Reid, Robert A; Burkhart, William; Turunen, Brandon J; Rong, James X; Wagner, Craig; Moyer, Mary B; Wells, Carrow; Hong, Xuan; Moore, John T; Williams, Jon D; Soler, Dulce; Ghosh, Shomir; Nolan, Michael A

2013-05-01

In contrast to studies on class I histone deacetylase (HDAC) inhibitors, the elucidation of the molecular mechanisms and therapeutic potential of class IIa HDACs (HDAC4, HDAC5, HDAC7 and HDAC9) is impaired by the lack of potent and selective chemical probes. Here we report the discovery of inhibitors that fill this void with an unprecedented metal-binding group, trifluoromethyloxadiazole (TFMO), which circumvents the selectivity and pharmacologic liabilities of hydroxamates. We confirm direct metal binding of the TFMO through crystallographic approaches and use chemoproteomics to demonstrate the superior selectivity of the TFMO series relative to a hydroxamate-substituted analog. We further apply these tool compounds to reveal gene regulation dependent on the catalytic active site of class IIa HDACs. The discovery of these inhibitors challenges the design process for targeting metalloenzymes through a chelating metal-binding group and suggests therapeutic potential for class IIa HDAC enzyme blockers distinct in mechanism and application compared to current HDAC inhibitors.

Influence of active site location on catalytic activity in de novo-designed zinc metalloenzymes.

PubMed

Zastrow, Melissa L; Pecoraro, Vincent L

2013-04-17

While metalloprotein design has now yielded a number of successful metal-bound and even catalytically active constructs, the question of where to put a metal site along a linear, repetitive sequence has not been thoroughly addressed. Often several possibilities in a given sequence may exist that would appear equivalent but may in fact differ for metal affinity, substrate access, or protein dynamics. We present a systematic variation of active site location for a hydrolytically active ZnHis3O site contained within a de novo-designed three-stranded coiled coil. We find that the maximal rate, substrate access, and metal-binding affinity are dependent on the selected position, while catalytic efficiency for p-nitrophenyl acetate hydrolysis can be retained regardless of the location of the active site. This achievement demonstrates how efficient, tailor-made enzymes which control rate, pKa, substrate and solvent access (and selectivity), and metal-binding affinity may be realized. These findings may be applied to the more advanced de novo design of constructs containing secondary interactions, such as hydrogen-bonding channels. We are now confident that changes to location for accommodating such channels can be achieved without location-dependent loss of catalytic efficiency. These findings bring us closer to our ultimate goal of incorporating the secondary interactions we believe will be necessary in order to improve both active site properties and the catalytic efficiency to be competitive with the native enzyme, carbonic anhydrase.

Locating the Binding Sites of Pb(II) Ion with Human and Bovine Serum Albumins

PubMed Central

Belatik, Ahmed; Hotchandani, Surat; Carpentier, Robert; Tajmir-Riahi, Heidar-Ali

2012-01-01

Lead is a potent environmental toxin that has accumulated above its natural level as a result of human activity. Pb cation shows major affinity towards protein complexation and it has been used as modulator of protein-membrane interactions. We located the binding sites of Pb(II) with human serum (HSA) and bovine serum albumins (BSA) at physiological conditions, using constant protein concentration and various Pb contents. FTIR, UV-visible, CD, fluorescence and X-ray photoelectron spectroscopic (XPS) methods were used to analyse Pb binding sites, the binding constant and the effect of metal ion complexation on HSA and BSA stability and conformations. Structural analysis showed that Pb binds strongly to HSA and BSA via hydrophilic contacts with overall binding constants of KPb-HSA = 8.2 (±0.8)×104 M−1 and KPb-BSA = 7.5 (±0.7)×104 M−1. The number of bound Pb cation per protein is 0.7 per HSA and BSA complexes. XPS located the binding sites of Pb cation with protein N and O atoms. Pb complexation alters protein conformation by a major reduction of α-helix from 57% (free HSA) to 48% (metal-complex) and 63% (free BSA) to 52% (metal-complex) inducing a partial protein destabilization. PMID:22574219

The Effect of Salts in Promoting Specific and Competitive Interactions between Zinc Finger Proteins and Metals

NASA Astrophysics Data System (ADS)

Li, Gongyu; Yuan, Siming; Zheng, Shihui; Chen, Yuting; Zheng, Zhen; Liu, Yangzhong; Huang, Guangming

2017-12-01

Specific protein-metal interactions (PMIs) fulfill essential functions in cells and organic bodies, and activation of these functions in vivo are mostly modulated by the complex environmental factors, including pH value, small biomolecules, and salts. Specifically, the role of salts in promoting specific PMIs and their competition among various metals has remained untapped mainly due to the difficulty to distinguish nonspecific PMIs from specific PMIs by classic spectroscopic techniques. Herein, we report Hofmeister salts differentially promote the specific PMIs by combining nanoelectrospray ionization mass spectrometry and spectroscopic techniques (fluorescence measurement and circular dichroism). Furthermore, to explore the influence of salts in competitive binding between metalloproteins and various metals, we designed a series of competitive experiments and applied to a well-defined model system, the competitive binding of zinc (II) and arsenic (III) to holo-promyelocytic leukemia protein (PML). These experiments not only provided new insights at the molecular scale as complementary to previous NMR and spectroscopic results, but also deduced the relative binding ability between zinc finger proteins and metals at the molecular scale, which avoids the mass spectrometric titration-based determination of binding constants that is frequently affected and often degraded by variable solution conditions including salt contents. [Figure not available: see fulltext.

Metal-binding proteins as metal pollution indicators

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hennig, H.F.

1986-03-01

The fact that metal-binding proteins are a consequence of elevated metal concentration in organisms is well known. What has been overlooked is that the presence of these proteins provides a unique opportunity to reformulate the criteria of metal pollution. The detoxification effect of metal-binding proteins in animals from polluted areas has been cited, but there have been only very few studies relating metal-binding proteins to pollution. This lack is due partly to the design of most experiments, which were aimed at isolation of metal-binding proteins and hence were of too short duration to allow for correlation to adverse physiological effectsmore » on the organism. In this study metal-binding proteins were isolated and characterized from five different marine animals (rock lobster, Jasus lalandii; hermit crab, Diogenes brevirostris; sandshrimp, Palaemon pacificus; black mussel, Choromytilus meridionalis; and limpet, Patella granularis). These animals were kept under identical metal-enriched conditions, hence eliminating differences in method and seasons. The study animals belonged to different phyla; varied in size, mass, age, behavior, food requirements and life stages; and accumulated metals at different rates. It is possible to link unseasonal moulting in crustacea, a known physiological effect due to a metal-enriched environment, to the production of the metal-binding protein without evidence of obvious metal body burden. Thus a new concept of pollution is defined: the presence of metal-binding proteins confirms toxic metal pollution. This concept was then tested under field conditions in the whelk Bullia digitalis and in metal-enriched grass.« less

Effects of Zinc on Particulate Methane Monooxygenase Activity and Structure*

PubMed Central

Sirajuddin, Sarah; Barupala, Dulmini; Helling, Stefan; Marcus, Katrin; Stemmler, Timothy L.; Rosenzweig, Amy C.

2014-01-01

Particulate methane monooxygenase (pMMO) is a membrane-bound metalloenzyme that oxidizes methane to methanol in methanotrophic bacteria. Zinc is a known inhibitor of pMMO, but the details of zinc binding and the mechanism of inhibition are not understood. Metal binding and activity assays on membrane-bound pMMO from Methylococcus capsulatus (Bath) reveal that zinc inhibits pMMO at two sites that are distinct from the copper active site. The 2.6 Å resolution crystal structure of Methylocystis species strain Rockwell pMMO reveals two previously undetected bound lipids, and metal soaking experiments identify likely locations for the two zinc inhibition sites. The first is the crystallographic zinc site in the pmoC subunit, and zinc binding here leads to the ordering of 10 previously unobserved residues. A second zinc site is present on the cytoplasmic side of the pmoC subunit. Parallels between these results and zinc inhibition studies of several respiratory complexes suggest that zinc might inhibit proton transfer in pMMO. PMID:24942740

The involvement of coordinative interactions in the binding of dihydrolipoamide dehydrogenase to titanium dioxide-Localization of a putative binding site.

PubMed

Dayan, Avraham; Babin, Gilad; Ganoth, Assaf; Kayouf, Nivin Samir; Nitoker Eliaz, Neta; Mukkala, Srijana; Tsfadia, Yossi; Fleminger, Gideon

2017-08-01

Titanium (Ti) and its alloys are widely used in orthodontic and orthopedic implants by virtue to their high biocompatibility, mechanical strength, and high resistance to corrosion. Biointegration of the implants with the tissue requires strong interactions, which involve biological molecules, proteins in particular, with metal oxide surfaces. An exocellular high-affinity titanium dioxide (TiO 2 )-binding protein (TiBP), purified from Rhodococcus ruber, has been previously studied in our lab. This protein was shown to be homologous with the orthologous cytoplasmic rhodococcal dihydrolipoamide dehydrogenase (rhDLDH). We have found that rhDLDH and its human homolog (hDLDH) share the TiO 2 -binding capabilities with TiBP. Intrigued by the unique TiO 2 -binding properties of hDLDH, we anticipated that it may serve as a molecular bridge between Ti-based medical structures and human tissues. The objective of the current study was to locate the region and the amino acids of the protein that mediate the protein-TiO 2 surface interaction. We demonstrated the role of acidic amino acids in the nonelectrostatic enzyme/dioxide interactions at neutral pH. The observation that the interaction of DLDH with various metal oxides is independent of their isoelectric values strengthens this notion. DLDH does not lose its enzymatic activity upon binding to TiO 2 , indicating that neither the enzyme undergoes major conformational changes nor the TiO 2 binding site is blocked. Docking predictions suggest that both rhDLDH and hDLDH bind TiO 2 through similar regions located far from the active site and the dimerization sites. The putative TiO 2 -binding regions of both the bacterial and human enzymes were found to contain a CHED (Cys, His, Glu, Asp) motif, which has been shown to participate in metal-binding sites in proteins. Copyright © 2017 John Wiley & Sons, Ltd.

Lanthanide and transition metal complexes of bioactive coumarins: molecular modeling and spectroscopic studies.

PubMed

Georgieva, I; Mihaylov, Tz; Trendafilova, N

2014-06-01

The present paper summarizes theoretical and spectroscopic investigations on a series of active coumarins and their lanthanide and transition metal complexes with application in medicine and pharmacy. Molecular modeling as well as IR, Raman, NMR and electronic spectral simulations at different levels of theory were performed to obtain important molecular descriptors: total energy, formation energy, binding energy, stability, conformations, structural parameters, electron density distribution, molecular electrostatic potential, Fukui functions, atomic charges, and reactive indexes. The computations are performed both in gas phase and in solution with consideration of the solvent effect on the molecular structural and energetic parameters. The investigations have shown that the advanced computational methods are reliable for prediction of the metal-coumarin binding mode, electron density distribution, thermodynamic properties as well as the strength and nature of the metal-coumarin interaction (not experimentally accessible) and correctly interpret the experimental spectroscopic data. Known results from biological tests for cytotoxic, antimicrobial, anti-fungal, spasmolytic and anti-HIV activities on the studied metal complexes are reported and discussed. Copyright © 2014 Elsevier Inc. All rights reserved.

Comparative serum albumin interactions and antitumor effects of Au(III) and Ga(III) ions.

PubMed

Sarioglu, Omer Faruk; Ozdemir, Ayse; Karaboduk, Kuddusi; Tekinay, Turgay

2015-01-01

In the present study, interactions of Au(III) and Ga(III) ions on human serum albumin (HSA) were studied comparatively via spectroscopic and thermal analysis methods: UV-vis absorbance spectroscopy, fluorescence spectroscopy, Fourier transform infrared (FT-IR) spectroscopy and isothermal titration calorimetry (ITC). The potential antitumor effects of these ions were studied on MCF-7 cells via Alamar blue assay. It was found that both Au(III) and Ga(III) ions can interact with HSA, however; Au(III) ions interact with HSA more favorably and with a higher affinity. FT-IR second derivative analysis results demonstrated that, high concentrations of both metal ions led to a considerable decrease in the α-helix content of HSA; while Au(III) led to around 5% of decrease in the α-helix content at 200μM, it was around 1% for Ga(III) at the same concentration. Calorimetric analysis gave the binding kinetics of metal-HSA interactions; while the binding affinity (Ka) of Au(III)-HSA binding was around 3.87×10(5)M(-1), it was around 9.68×10(3)M(-1) for Ga(III)-HSA binding. Spectroscopy studies overall suggest that both metal ions have significant effects on the chemical structure of HSA, including the secondary structure alterations. Antitumor activity studies on MCF7 tumor cell line with both metal ions revealed that, Au(III) ions have a higher antiproliferative activity compared to Ga(III) ions. Copyright © 2014 Elsevier GmbH. All rights reserved.

Influence of sulfhydryl sites on metal binding by bacteria

NASA Astrophysics Data System (ADS)

Nell, Ryan M.; Fein, Jeremy B.

2017-02-01

The role of sulfhydryl sites within bacterial cell envelopes is still unknown, but the sites may control the fate and bioavailability of metals. Organic sulfhydryl compounds are important complexing ligands in aqueous systems and they can influence metal speciation in natural waters. Though representing only approximately 5-10% of the total available binding sites on bacterial surfaces, sulfhydryl sites exhibit high binding affinities for some metals. Due to the potential importance of bacterial sulfhydryl sites in natural systems, metal-bacterial sulfhydryl site binding constants must be determined in order to construct accurate models of the fate and distribution of metals in these systems. To date, only Cd-sulfhydryl binding has been quantified. In this study, the thermodynamic stabilities of Mn-, Co-, Ni-, Zn-, Sr- and Pb-sulfhydryl bacterial cell envelope complexes were determined for the bacterial species Shewanella oneidensis MR-1. Metal adsorption experiments were conducted as a function of both pH, ranging from 5.0 to 7.0, and metal loading, from 0.5 to 40.0 μmol/g (wet weight) bacteria, in batch experiments in order to determine if metal-sulfhydryl binding occurs. Initially, the data were used to calculate the value of the stability constants for the important metal-sulfhydryl bacterial complexes for each metal-loading condition studied, assuming a single binding reaction for the dominant metal-binding site type under the pH conditions of the experiments. For most of the metals that we studied, these calculated stability constant values increased significantly with decreasing metal loading, strongly suggesting that our initial assumption was not valid and that more than one type of binding occurs at the assumed binding site. We then modeled each dataset with two distinct site types with identical acidity constants: one site with a high metal-site stability constant value, which we take to represent metal-sulfhydryl binding and which dominates under low metal loading conditions, and another more abundant site that we term non-sulfhydryl sites that becomes important at high metal loadings. The resulting calculated stability constants do not vary significantly as a function of metal loading and yield reasonable fits to the observed adsorption behaviors as a function of both pH and metal loading. We use the results to calculate the speciation of metals bound by the bacterial envelope in realistic bacteria-bearing, heavy metal contaminated systems in order to demonstrate the potential importance of metal-sulfhydryl binding in the budget of bacterially-adsorbed metals under low metal-loading conditions.

Interactions of the Metalloregulatory Protein SloR from Streptococcus mutans with Its Metal Ion Effectors and DNA Binding Site

PubMed Central

Corbett, John; Cornacchione, Louis; Daly, William; Galan, Diego; Wysota, Michael; Tivnan, Patrick; Collins, Justin; Nye, Dillon; Levitz, Talya; Breyer, Wendy A.; Glasfeld, Arthur

2015-01-01

ABSTRACT Streptococcus mutans is the causative agent of dental caries, a significant concern for human health, and therefore an attractive target for therapeutics development. Previous work in our laboratory has identified a homodimeric, manganese-dependent repressor protein, SloR, as an important regulator of cariogenesis and has used site-directed mutagenesis to map functions to specific regions of the protein. Here we extend those studies to better understand the structural interaction between SloR and its operator and its effector metal ions. The results of DNase I assays indicate that SloR protects a 42-bp region of DNA that overlaps the sloABC promoter on the S. mutans UA159 chromosome, while electrophoretic mobility shift and solution binding assays indicate that each of two SloR dimers binds to this region. Real-time semiquantitative reverse transcriptase PCR (real-time semi-qRT-PCR) experiments were used to determine the individual base pairs that contribute to SloR-DNA binding specificity. Solution studies indicate that Mn2+ is better than Zn2+ at specifically activating SloR to bind DNA, and yet the 2.8-Å resolved crystal structure of SloR bound to Zn2+ provides insight into the means by which selective activation by Mn2+ may be achieved and into how SloR may form specific interactions with its operator. Taken together, these experimental observations are significant because they can inform rational drug design aimed at alleviating and/or preventing S. mutans-induced caries formation. IMPORTANCE This report focuses on investigating the SloR protein as a regulator of essential metal ion transport and virulence gene expression in the oral pathogen Streptococcus mutans and on revealing the details of SloR binding to its metal ion effectors and binding to DNA that together facilitate this expression. We used molecular and biochemical approaches to characterize the interaction of SloR with Mn2+ and with its SloR recognition element to gain a clearer picture of the regulatory networks that optimize SloR-mediated metal ion homeostasis and virulence gene expression in S. mutans. These experiments can have a significant impact on caries treatment and/or prevention by revealing the S. mutans SloR-DNA binding interface as an appropriate target for the development of novel therapeutic interventions. PMID:26350131

Relationship between Ni(II) and Zn(II) Coordination and Nucleotide Binding by the Helicobacter pylori [NiFe]-Hydrogenase and Urease Maturation Factor HypB*

PubMed Central

Sydor, Andrew M.; Lebrette, Hugo; Ariyakumaran, Rishikesh; Cavazza, Christine; Zamble, Deborah B.

2014-01-01

The pathogen Helicobacter pylori requires two nickel-containing enzymes, urease and [NiFe]-hydrogenase, for efficient colonization of the human gastric mucosa. These enzymes possess complex metallocenters that are assembled by teams of proteins in multistep pathways. One essential accessory protein is the GTPase HypB, which is required for Ni(II) delivery to [NiFe]-hydrogenase and participates in urease maturation. Ni(II) or Zn(II) binding to a site embedded in the GTPase domain of HypB modulates the enzymatic activity, suggesting a mechanism of regulation. In this study, biochemical and structural analyses of H. pylori HypB (HpHypB) revealed an intricate link between nucleotide and metal binding. HpHypB nickel coordination, stoichiometry, and affinity were modulated by GTP and GDP, an effect not observed for zinc, and biochemical evidence suggests that His-107 coordination to nickel toggles on and off in a nucleotide-dependent manner. These results are consistent with the crystal structure of HpHypB loaded with Ni(II), GDP, and Pi, which reveals a nickel site distinct from that of zinc-loaded Methanocaldococcus jannaschii HypB as well as subtle changes to the protein structure. Furthermore, Cys-142, a metal ligand from the Switch II GTPase motif, was identified as a key component of the signal transduction between metal binding and the enzymatic activity. Finally, potassium accelerated the enzymatic activity of HpHypB but had no effect on the other biochemical properties of the protein. Altogether, this molecular level information about HpHypB provides insight into its cellular function and illuminates a possible mechanism of metal ion discrimination. PMID:24338018

Relationship between Ni(II) and Zn(II) coordination and nucleotide binding by the Helicobacter pylori [NiFe]-hydrogenase and urease maturation factor HypB.

PubMed

Sydor, Andrew M; Lebrette, Hugo; Ariyakumaran, Rishikesh; Cavazza, Christine; Zamble, Deborah B

2014-02-14

The pathogen Helicobacter pylori requires two nickel-containing enzymes, urease and [NiFe]-hydrogenase, for efficient colonization of the human gastric mucosa. These enzymes possess complex metallocenters that are assembled by teams of proteins in multistep pathways. One essential accessory protein is the GTPase HypB, which is required for Ni(II) delivery to [NiFe]-hydrogenase and participates in urease maturation. Ni(II) or Zn(II) binding to a site embedded in the GTPase domain of HypB modulates the enzymatic activity, suggesting a mechanism of regulation. In this study, biochemical and structural analyses of H. pylori HypB (HpHypB) revealed an intricate link between nucleotide and metal binding. HpHypB nickel coordination, stoichiometry, and affinity were modulated by GTP and GDP, an effect not observed for zinc, and biochemical evidence suggests that His-107 coordination to nickel toggles on and off in a nucleotide-dependent manner. These results are consistent with the crystal structure of HpHypB loaded with Ni(II), GDP, and Pi, which reveals a nickel site distinct from that of zinc-loaded Methanocaldococcus jannaschii HypB as well as subtle changes to the protein structure. Furthermore, Cys-142, a metal ligand from the Switch II GTPase motif, was identified as a key component of the signal transduction between metal binding and the enzymatic activity. Finally, potassium accelerated the enzymatic activity of HpHypB but had no effect on the other biochemical properties of the protein. Altogether, this molecular level information about HpHypB provides insight into its cellular function and illuminates a possible mechanism of metal ion discrimination.

The mechanism of Cu+ transport ATPases: interaction with CU+ chaperones and the role of transient metal-binding sites.

PubMed

Padilla-Benavides, Teresita; McCann, Courtney J; Argüello, José M

2013-01-04

Cu(+)-ATPases are membrane proteins that couple the hydrolysis of ATP to the efflux of cytoplasmic Cu(+). In cells, soluble chaperone proteins bind and distribute cytoplasmic Cu(+), delivering the ion to the transmembrane metal-binding sites in the ATPase. The structure of Legionella pneumophila Cu(+)-ATPase (Gourdon, P., Liu, X. Y., Skjørringe, T., Morth, J. P., Møller, L. B., Pedersen, B. P., and Nissen, P. (2011) Nature 475, 59-64) shows that a kinked transmembrane segment forms a "platform" exposed to the cytoplasm. In addition, neighboring invariant Met, Asp, and Glu are located at the "entrance" of the ion path. Mutations of amino acids in these regions of the Archaeoglobus fulgidus Cu(+)-ATPase CopA do not affect ATPase activity in the presence of Cu(+) free in solution. However, Cu(+) bound to the corresponding chaperone (CopZ) could not activate the mutated ATPases, and in parallel experiments, CopZ was unable to transfer Cu(+) to CopA. Furthermore, mutation of a specific electronegative patch on the CopZ surface abolishes the ATPase activation and Cu(+) transference, indicating that the region is required for the CopZ-CopA interaction. Moreover, the data suggest that the interaction is driven by the complementation of the electropositive platform in the ATPase and the electronegative Cu(+) chaperone. This docking likely places the Cu(+) proximal to the conserved carboxyl and thiol groups in the entrance site that induce metal release from the chaperone via ligand exchange. The initial interaction of Cu(+) with the pump is transient because Cu(+) is transferred from the entrance site to transmembrane metal-binding sites involved in transmembrane translocation.

Metal binding characterization and conformational studies using Raman microscopy of resin-bound poly(aspartic acid).

PubMed

Stair, Jacqueline L; Holcombe, James A

2007-03-01

The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6, 20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+. Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 mumol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., beta-sheets, alpha-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to beta-sheets and alpha-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement.

Resolving distinct molecular origins for copper effects on PAI-1.

PubMed

Bucci, Joel C; McClintock, Carlee S; Chu, Yuzhuo; Ware, Gregory L; McConnell, Kayla D; Emerson, Joseph P; Peterson, Cynthia B

2017-10-01

Components of the fibrinolytic system are subjected to stringent control to maintain proper hemostasis. Central to this regulation is the serpin plasminogen activator inhibitor-1 (PAI-1), which is responsible for specific and rapid inhibition of fibrinolytic proteases. Active PAI-1 is inherently unstable and readily converts to a latent, inactive form. The binding of vitronectin and other ligands influences stability of active PAI-1. Our laboratory recently observed reciprocal effects on the stability of active PAI-1 in the presence of transition metals, such as copper, depending on the whether vitronectin was also present (Thompson et al. Protein Sci 20:353-365, 2011). To better understand the molecular basis for these copper effects on PAI-1, we have developed a gel-based copper sensitivity assay that can be used to assess the copper concentrations that accelerate the conversion of active PAI-1 to a latent form. The copper sensitivity of wild-type PAI-1 was compared with variants lacking N-terminal histidine residues hypothesized to be involved in copper binding. In these PAI-1 variants, we observed significant differences in copper sensitivity, and these data were corroborated by latency conversion kinetics and thermodynamics of copper binding by isothermal titration calorimetry. These studies identified a copper-binding site involving histidines at positions 2 and 3 that confers a remarkable stabilization of PAI-1 beyond what is observed with vitronectin alone. A second site, independent from the two histidines, binds metal and increases the rate of the latency conversion.

A molecular catalyst for water oxidation that binds to metal oxide surfaces

PubMed Central

Sheehan, Stafford W.; Thomsen, Julianne M.; Hintermair, Ulrich; Crabtree, Robert H.; Brudvig, Gary W.; Schmuttenmaer, Charles A.

2015-01-01

Molecular catalysts are known for their high activity and tunability, but their solubility and limited stability often restrict their use in practical applications. Here we describe how a molecular iridium catalyst for water oxidation directly and robustly binds to oxide surfaces without the need for any external stimulus or additional linking groups. On conductive electrode surfaces, this heterogenized molecular catalyst oxidizes water with low overpotential, high turnover frequency and minimal degradation. Spectroscopic and electrochemical studies show that it does not decompose into iridium oxide, thus preserving its molecular identity, and that it is capable of sustaining high activity towards water oxidation with stability comparable to state-of-the-art bulk metal oxide catalysts. PMID:25757425

Mechanical work makes important contributions to surface chemistry at steps.

PubMed

Francis, M F; Curtin, W A

2015-02-13

The effect of mechanical strain on the binding energy of adsorbates to late transition metals is believed to be entirely controlled by electronic factors, with tensile stress inducing stronger binding. Here we show, via computation, that mechanical strain of late transition metals can modify binding at stepped surfaces opposite to well-established trends on flat surfaces. The mechanism driving the trend is mechanical, arising from the relaxation of stored mechanical energy. The mechanical energy change can be larger than, and of opposite sign than, the energy changes due to electronic effects and leads to a violation of trends predicted by the widely accepted electronic 'd-band' model. This trend has a direct impact on catalytic activity, which is demonstrated here for methanation, where biaxial tension is predicted to shift the activity of nickel significantly, reaching the peak of the volcano plot and comparable to cobalt and ruthenium.

DOE Office of Scientific and Technical Information (OSTI.GOV)

B Akabayov; C Richardson

Divalent metal ions are crucial as cofactors for a variety of intracellular enzymatic activities. Mg{sup 2+}, as an example, mediates binding of deoxyribonucleoside 5'-triphosphates followed by their hydrolysis in the active site of DNA polymerase. It is difficult to study the binding of Mg{sup 2+} to an active site because Mg{sup 2+} is spectroscopically silent and Mg{sup 2+} binds with low affinity to the active site of an enzyme. Therefore, we substituted Mg{sup 2+} with Mn{sup 2+}:Mn{sup 2+} that is not only visible spectroscopically but also provides full activity of the DNA polymerase of bacteriophage T7. In order to demonstratemore » that the majority of Mn{sup 2+} is bound to the enzyme, we have applied site-directed titration analysis of T7 DNA polymerase using X-ray near edge spectroscopy. Here we show how X-ray near edge spectroscopy can be used to distinguish between signal originating from Mn{sup 2+} that is free in solution and Mn{sup 2+} bound to the active site of T7 DNA polymerase. This method can be applied to other enzymes that use divalent metal ions as a cofactor.« less

Binding of the Zn2+ ion to ferric uptake regulation protein from E. coli and the competition with Fe2+ binding: a molecular modeling study of the effect on DNA binding and conformational changes of Fur

NASA Astrophysics Data System (ADS)

Jabour, Salih; Hamed, Mazen Y.

2009-04-01

The three dimensional structure of Ferric uptake regulation protein dimer from E. coli, determined by molecular modeling, was docked on a DNA fragment (iron box) and Zn2+ ions were added in two steps. The first step involved the binding of one Zn2+ ion to what is known as the zinc site which consists of the residues Cys 92, Cys 95, Asp 137, Asp141, Arg139, Glu 140, His 145 and His 143 with an average metal-Nitrogen distance of 2.5 Å and metal-oxygen distance of 3.1-3.2 Å. The second Zn2+ ion is bound to the iron activating site formed from the residues Ile 50, His 71, Asn 72, Gly 97, Asp 105 and Ala 109. The binding of the second Zn2+ ion strengthened the binding of the first ion as indicated by the shortening of the zinc-residue distances. Fe2+, when added to the complex consisting of 2Zn2+/Fur dimer/DNA, replaced the Zn2+ ion in the zinc site and when a second Fe2+ was added, it replaced the second zinc ion in the iron activating site. The binding of both zinc and iron ions induced a similar change in Fur conformations, but shifted residues closer to DNA in a different manner. This is discussed along with a possible role for the Zn2+ ion in the Fur dimer binding of DNA in its repressor activity.

Mechanistic characterization of the HDV genomic ribozyme: a mutant of the C41 motif provides insight into the positioning and thermodynamic linkage of metal ions and protons.

PubMed

Nakano, Shu-ichi; Bevilacqua, Philip C

2007-03-20

Binding of two Mg2+ and two H+ ions influences the self-cleavage activity of the genomic HDV ribozyme. The positioning of these four ligands and their thermodynamic linkage are not fully resolved. Protonated C41 engages in a base triple, whereas protonated C75 has been implicated as an acid-base catalyst in bond cleavage. Prior studies led to the identification of one structural inner-sphere ion and one catalytic outer-sphere ion. In the present study, the contributions of the C41 base triple to the metal ion- and pH-dependence of the reaction are examined. Experiments were conducted on a CG to UA double mutant (DM), which changes the base triple to one involving an unprotonated C41. Below pH 6, the DM has a steeper dependence on pH than the wild-type (WT), consistent with a single protonation misfolding the core; this conclusion is also supported by thermal denaturation studies. Between pH 6 and 8, the WT and DM display nearly identical catalytic metal ion and H+ binding profiles. In contrast, over the same pH range, the WT and DM have distinct structural ion binding profiles; for the WT, binding is favored at lower pH, whereas the DM shows no pH dependence. These data localize the structural ion to the vicinity of the C41 motif. An overall model is presented that accommodates binding affinity, coupling, and positioning of the two metal ions and the two protons within the ribozyme. The data suggest that a protonated base triple allows the WT ribozyme to maintain appreciable activity at acidic pH, which could play an important role in the life cycle of the virus.

Identification of a divalent metal cation binding site in herpes simplex virus 1 (HSV-1) ICP8 required for HSV replication.

PubMed

Bryant, Kevin F; Yan, Zhipeng; Dreyfus, David H; Knipe, David M

2012-06-01

Herpes simplex virus 1 (HSV-1) ICP8 is a single-stranded DNA-binding protein that is necessary for viral DNA replication and exhibits recombinase activity in vitro. Alignment of the HSV-1 ICP8 amino acid sequence with ICP8 homologs from other herpesviruses revealed conserved aspartic acid (D) and glutamic acid (E) residues. Amino acid residue D1087 was conserved in every ICP8 homolog analyzed, indicating that it is likely critical for ICP8 function. We took a genetic approach to investigate the functions of the conserved ICP8 D and E residues in HSV-1 replication. The E1086A D1087A mutant form of ICP8 failed to support the replication of an ICP8 mutant virus in a complementation assay. E1086A D1087A mutant ICP8 bound DNA, albeit with reduced affinity, demonstrating that the protein is not globally misfolded. This mutant form of ICP8 was also recognized by a conformation-specific antibody, further indicating that its overall structure was intact. A recombinant virus expressing E1086A D1087A mutant ICP8 was defective in viral replication, viral DNA synthesis, and late gene expression in Vero cells. A class of enzymes called DDE recombinases utilize conserved D and E residues to coordinate divalent metal cations in their active sites. We investigated whether the conserved D and E residues in ICP8 were also required for binding metal cations and found that the E1086A D1087A mutant form of ICP8 exhibited altered divalent metal binding in an in vitro iron-induced cleavage assay. These results identify a novel divalent metal cation-binding site in ICP8 that is required for ICP8 functions during viral replication.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Parkes, Marie V.; Sava Gallis, Dorina F.; Greathouse, Jeffery A.

Computational screening of metal-organic framework (MOF) materials for selective oxygen adsorption from air could lead to new sorbents for the oxyfuel combustion process feedstock streams. A comprehensive study on the effect of MOF metal chemistry on gas binding energies in two common but structurally disparate metal-organic frameworks has been undertaken. Dispersion-corrected density functional theory methods were used to calculate the oxygen and nitrogen binding energies with each of fourteen metals, respectively, substituted into two MOF series, M 2(dobdc) and M 3(btc) 2. The accuracy of DFT methods was validated by comparing trends in binding energy with experimental gas sorption measurements.more » A periodic trend in oxygen binding energies was found, with greater oxygen binding energies for early transition-metal-substituted MOFs compared to late transition metal MOFs; this was independent of MOF structural type. The larger binding energies were associated with oxygen binding in a side-on configuration to the metal, with concomitant lengthening of the O-O bond. In contrast, nitrogen binding energies were similar across the transition metal series, regardless of both MOF structural type and metal identity. Altogether, these findings suggest that early transition metal MOFs are best suited to separating oxygen from nitrogen, and that the MOF structural type is less important than the metal identity.« less

Crystallographic Location and Mutational Analysis of Zn and Cd Inhibitory Sites and Role of Lipidic Carboxylates in Rescuing Proton Path Mutants in Cytochrome c Oxidase†

PubMed Central

Qin, Ling; Mills, Denise A.; Hiser, Carrie; Murphree, Anna; Garavito, R. Michael; Ferguson-Miller, Shelagh; Hosler, Jonathan

2008-01-01

Cytochrome c oxidase (CcO) transfers protons from the inner surface of the enzyme to the buried O2 reduction site through two different pathways, termed K and D, and from the outer surface via an undefined route. These proton paths can be inhibited by metals such as zinc or cadmium, but the sites of inhibition have not been established. Anomalous difference Fourier analyses of Rhodobacter sphaeroides CcO crystals, with cadmium added, reveal metal binding sites that include the proposed initial proton donor/acceptor of the K pathway, Glu-101 of subunit II. Mutant forms of CcO that lack Glu-101II (E101A and E101A/H96A) exhibit low activity and eliminate metal binding at this site. Significant activity is restored to E101A and E101A/H96A by adding the lipophilic carboxylic compounds, arachidonic acid and cholic acid, but not by their non-carboxylic analogues. These amphipathic acids likely provide their carboxylic groups as substitute proton donors/acceptors in the absence of Glu-101II, as previously observed for arachidonic acid in mutants that alter Asp-132I of the D pathway. The activity of E101A/H96A is still inhibited by zinc, but this remaining inhibition is nearly eliminated by removal of subunit III, which is known to alter the D pathway. The results identify the Glu-101/His-96 site of subunit II as the site of metal binding that inhibits the uptake of protons into the K pathway and indicate that subunit III contributes to zinc binding and/or inhibition of the D pathway. By removing subunit III from E101A/H96A, thereby eliminating zinc inhibition of the uptake of protons from the inner surface of CcO, we confirm that an external zinc binding site is involved in inhibiting the backflow of protons to the active site. PMID:17477548

DNA binding, anti-inflammatory and analgesic evaluation of metal complexes of N/S/O donor ligands; Synthesis, spectral characterization

NASA Astrophysics Data System (ADS)

Kumar Naik, K. H.; Ashok, B.; Naik, Nagaraja; Mulla, Jameel Ahmed S.; Prakasha, Avinash

2015-04-01

Transition metal complexes containing tri-dentate NSN donor ligands i.e., 5-((1(aminomethyl)cyclohexyl)methyl)-1,3,4-thiadiazol-2-amine (AMTA) (2) and 5-(2-aminophenyl)-1,3,4-thiadiazol-2-amine (ATA) (4i-ii) have been synthesized. The newly synthesized ligands and their respective complexes were characterized by elemental analysis, molar conductance measurement and various spectral studies [infrared (IR), electronic, and NMR (for ligands only)]. Metal complexes are like [M(AMTA)2], [M(ATA)2] type, where M = Mn(II), Co(II) and Cu(II). The proposed geometries of the complexes are octahedral in nature. The synthesized ligands and their complexes were exhibits effective anti-inflammatory, analgesic and DNA binding activities. All the tested compounds exhibited significant analgesic activity, whereas the compound 4i, 4(ia) and 4(iib) is equipotent with Diclofenac sodium.

Effect of metal in M 3(btc) 2 and M 2(dobdc) MOFs for O 2/N 2 separations: A combined density functional theory and experimental study

DOE PAGES

Parkes, Marie V.; Sava Gallis, Dorina F.; Greathouse, Jeffery A.; ...

2015-03-02

Computational screening of metal-organic framework (MOF) materials for selective oxygen adsorption from air could lead to new sorbents for the oxyfuel combustion process feedstock streams. A comprehensive study on the effect of MOF metal chemistry on gas binding energies in two common but structurally disparate metal-organic frameworks has been undertaken. Dispersion-corrected density functional theory methods were used to calculate the oxygen and nitrogen binding energies with each of fourteen metals, respectively, substituted into two MOF series, M 2(dobdc) and M 3(btc) 2. The accuracy of DFT methods was validated by comparing trends in binding energy with experimental gas sorption measurements.more » A periodic trend in oxygen binding energies was found, with greater oxygen binding energies for early transition-metal-substituted MOFs compared to late transition metal MOFs; this was independent of MOF structural type. The larger binding energies were associated with oxygen binding in a side-on configuration to the metal, with concomitant lengthening of the O-O bond. In contrast, nitrogen binding energies were similar across the transition metal series, regardless of both MOF structural type and metal identity. Altogether, these findings suggest that early transition metal MOFs are best suited to separating oxygen from nitrogen, and that the MOF structural type is less important than the metal identity.« less

The Relative Influence of Metal Ion Binding Sites in the I-like Domain and the Interface with the Hybrid Domain on Rolling and Firm Adhesion by Integrin α4β7*

PubMed Central

Chen, JianFeng; Takagi, Junichi; Xie, Can; Xiao, Tsan; Luo, Bing-Hao; Springer, Timothy A.

2015-01-01

We examined the effect of conformational change at the β7 I-like/hybrid domain interface on regulating the transition between rolling and firm adhesion by integrin α4β7. An N-glycosylation site was introduced into the I-like/hybrid domain interface to act as a wedge and to stabilize the open conformation of this interface and hence the open conformation of the α4β7 headpiece. Wild-type α4β7 mediates rolling adhesion in Ca2+ and Ca2+/Mg2+ but firm adhesion in Mg2+ and Mn2+. Stabilizing the open headpiece resulted in firm adhesion in all divalent cations. The interaction between metal binding sites in the I-like domain and the interface with the hybrid domain was examined in double mutants. Changes at these two sites can either counterbalance one another or be additive, emphasizing mutuality and the importance of multiple interfaces in integrin regulation. A double mutant with counterbalancing deactivating ligand-induced metal ion binding site (LIMBS) and activating wedge mutations could still be activated by Mn2+, confirming the importance of the adjacent to metal ion-dependent adhesion site (ADMIDAS) in integrin activation by Mn2+. Overall, the results demonstrate the importance of headpiece allostery in the conversion of rolling to firm adhesion. PMID:15448154

Preorganization of molecular binding sites in designed diiron proteins.

PubMed

Maglio, Ornella; Nastri, Flavia; Pavone, Vincenzo; Lombardi, Angela; DeGrado, William F

2003-04-01

De novo protein design provides an attractive approach to critically test the features that are required for metalloprotein structure and function. Previously we designed and crystallographically characterized an idealized dimeric model for the four-helix bundle class of diiron and dimanganese proteins [Dueferri 1 (DF1)]. Although the protein bound metal ions in the expected manner, access to its active site was blocked by large bulky hydrophobic residues. Subsequently, a substrate-access channel was introduced proximal to the metal-binding center, resulting in a protein with properties more closely resembling those of natural enzymes. Here we delineate the energetic and structural consequences associated with the introduction of these binding sites. To determine the extent to which the binding site was preorganized in the absence of metal ions, the apo structure of DF1 in solution was solved by NMR and compared with the crystal structure of the di-Zn(II) derivative. The overall fold of the apo protein was highly similar to that of the di-Zn(II) derivative, although there was a rotation of one of the helices. We also examined the thermodynamic consequences associated with building a small molecule-binding site within the protein. The protein exists in an equilibrium between folded dimers and unfolded monomers. DF1 is a highly stable protein (K(diss) = 0.001 fM), but the dissociation constant increases to 0.6 nM (deltadeltaG = 5.4 kcalmol monomer) as the active-site cavity is increased to accommodate small molecules.

Structure and mode of action of cyclic lipopeptide pseudofactin II with divalent metal ions.

PubMed

Janek, Tomasz; Rodrigues, Lígia R; Gudiña, Eduardo J; Czyżnikowska, Żaneta

2016-10-01

The interaction of natural lipopeptide pseudofactin II with a series of doubly charged metal cations was examined by matrix-assisted laser-desorption ionization-time of flight (MALDI-TOF) mass spectrometry and molecular modelling. The molecular modelling for metal-pseudofactin II provides information on the metal-peptide binding sites. Overall, Mg(2+), Ca(2+) and Zn(2+) favor the association with oxygen atoms spanning the peptide backbone, whereas Cu(2+) is coordinated by three nitrogens. Circular dichroism (CD) results confirmed that Zn(2+) and Cu(2+) can disrupt the secondary structure of pseudofactin II at high concentrations, while Ca(2+) and Mg(2+) did not essentially affect the structure of the lipopeptide. Interestingly, our results showed that the addition of Zn(2+) and Cu(2+) helped smaller micelles to form larger micellar aggregates. Since pseudofactin II binds metals, we tested whether this phenomena was somehow related to its antimicrobial activity against Staphylococcus epidermidis and Proteus mirabilis. We found that the antimicrobial effect of pseudofactin II was increased by supplementation of culture media with all tested divalent metal ions. Finally, by using Gram-positive and Gram-negative bacteria we showed that the higher antimicrobial activity of metal complexes of pseudofactin II is attributed to the disruption of the cytoplasmic membrane. Copyright © 2016 Elsevier B.V. All rights reserved.

Metal ion coordination in the E. coli Nudix hydrolase dihydroneopterin triphosphate pyrophosphatase: New clues into catalytic mechanism

PubMed Central

Ukachukwu, Chiamaka U.; Freeman, Dana M.; Quirk, Stephen

2017-01-01

Dihydroneopterin triphosphate pyrophosphatase (DHNTPase), a member of the Mg2+ dependent Nudix hydrolase superfamily, is the recently-discovered enzyme that functions in the second step of the pterin branch of the folate biosynthetic pathway in E. coli. DHNTPase is of interest because inhibition of enzymes in bacterial folate biosynthetic pathways is a strategy for antibiotic development. We determined crystal structures of DHNTPase with and without activating, Mg2+-mimicking metals Co2+ and Ni2+. Four metal ions, identified by anomalous scattering, and stoichiometrically confirmed in solution by isothermal titration calorimetry, are held in place by Glu56 and Glu60 within the Nudix sequence motif, Glu117, waters, and a sulfate ion, of which the latter is further stabilized by a salt bridge with Lys7. In silico docking of the DHNTP substrate reveals a binding mode in which the pterin ring moiety is nestled in a largely hydrophobic pocket, the β-phosphate activated for nucleophilic attack overlays with the crystallographic sulfate and is in line with an activated water molecule, and remaining phosphate groups are stabilized by all four identified metal ions. The structures and binding data provide new details regarding DHNTPase metal requirements, mechanism, and suggest a strategy for efficient inhibition. PMID:28742822

Competition between Metals for Binding to Methanobactin Enables Expression of Soluble Methane Monooxygenase in the Presence of Copper

PubMed Central

Kalidass, Bhagyalakshmi; Ul-Haque, Muhammad Farhan; Baral, Bipin S.; DiSpirito, Alan A.

2014-01-01

It is well known that copper is a key factor regulating expression of the two forms of methane monooxygenase found in proteobacterial methanotrophs. Of these forms, the cytoplasmic, or soluble, methane monooxygenase (sMMO) is expressed only at low copper concentrations. The membrane-bound, or particulate, methane monooxygenase (pMMO) is constitutively expressed with respect to copper, and such expression increases with increasing copper. Recent findings have shown that copper uptake is mediated by a modified polypeptide, or chalkophore, termed methanobactin. Although methanobactin has high specificity for copper, it can bind other metals, e.g., gold. Here we show that in Methylosinus trichosporium OB3b, sMMO is expressed and active in the presence of copper if gold is also simultaneously present. Such expression appears to be due to gold binding to methanobactin produced by M. trichosporium OB3b, thereby limiting copper uptake. Such expression and activity, however, was significantly reduced if methanobactin preloaded with copper was also added. Further, quantitative reverse transcriptase PCR (RT-qPCR) of transcripts of genes encoding polypeptides of both forms of MMO and SDS-PAGE results indicate that both sMMO and pMMO can be expressed when copper and gold are present, as gold effectively competes with copper for binding to methanobactin. Such findings suggest that under certain geochemical conditions, both forms of MMO may be expressed and active in situ. Finally, these findings also suggest strategies whereby field sites can be manipulated to enhance sMMO expression, i.e., through the addition of a metal that can compete with copper for binding to methanobactin. PMID:25416758

Competition between metals for binding to methanobactin enables expression of soluble methane monooxygenase in the presence of copper.

PubMed

Kalidass, Bhagyalakshmi; Ul-Haque, Muhammad Farhan; Baral, Bipin S; DiSpirito, Alan A; Semrau, Jeremy D

2015-02-01

It is well known that copper is a key factor regulating expression of the two forms of methane monooxygenase found in proteobacterial methanotrophs. Of these forms, the cytoplasmic, or soluble, methane monooxygenase (sMMO) is expressed only at low copper concentrations. The membrane-bound, or particulate, methane monooxygenase (pMMO) is constitutively expressed with respect to copper, and such expression increases with increasing copper. Recent findings have shown that copper uptake is mediated by a modified polypeptide, or chalkophore, termed methanobactin. Although methanobactin has high specificity for copper, it can bind other metals, e.g., gold. Here we show that in Methylosinus trichosporium OB3b, sMMO is expressed and active in the presence of copper if gold is also simultaneously present. Such expression appears to be due to gold binding to methanobactin produced by M. trichosporium OB3b, thereby limiting copper uptake. Such expression and activity, however, was significantly reduced if methanobactin preloaded with copper was also added. Further, quantitative reverse transcriptase PCR (RT-qPCR) of transcripts of genes encoding polypeptides of both forms of MMO and SDS-PAGE results indicate that both sMMO and pMMO can be expressed when copper and gold are present, as gold effectively competes with copper for binding to methanobactin. Such findings suggest that under certain geochemical conditions, both forms of MMO may be expressed and active in situ. Finally, these findings also suggest strategies whereby field sites can be manipulated to enhance sMMO expression, i.e., through the addition of a metal that can compete with copper for binding to methanobactin. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Viral genome packaging terminase cleaves DNA using the canonical RuvC-like two-metal catalysis mechanism

PubMed Central

Xu, Rui-Gang; Jenkins, Huw T.; Chechik, Maria; Blagova, Elena V.; Lopatina, Anna; Klimuk, Evgeny; Minakhin, Leonid; Severinov, Konstantin

2017-01-01

Abstract Bacteriophages and large dsDNA viruses encode sophisticated machinery to translocate their DNA into a preformed empty capsid. An essential part of this machine, the large terminase protein, processes viral DNA into constituent units utilizing its nuclease activity. Crystal structures of the large terminase nuclease from the thermophilic bacteriophage G20c show that it is most similar to the RuvC family of the RNase H-like endonucleases. Like RuvC proteins, the nuclease requires either Mn2+, Mg2+ or Co2+ ions for activity, but is inactive with Zn2+ and Ca2+. High resolution crystal structures of complexes with different metals reveal that in the absence of DNA, only one catalytic metal ion is accommodated in the active site. Binding of the second metal ion may be facilitated by conformational variability, which enables the two catalytic aspartic acids to be brought closer to each other. Structural comparison indicates that in common with the RuvC family, the location of the two catalytic metals differs from other members of the RNase H family. In contrast to a recently proposed mechanism, the available data do not support binding of the two metals at an ultra-short interatomic distance. Thus we postulate that viral terminases cleave DNA by the canonical RuvC-like mechanism. PMID:28100693

Divalent Metal-Ion Complexes with Dipeptide Ligands Having Phe and His Side-Chain Anchors: Effects of Sequence, Metal Ion, and Anchor.

PubMed

Dunbar, Robert C; Berden, Giel; Martens, Jonathan K; Oomens, Jos

2015-09-24

Conformational preferences have been surveyed for divalent metal cation complexes with the dipeptide ligands AlaPhe, PheAla, GlyHis, and HisGly. Density functional theory results for a full set of complexes are presented, and previous experimental infrared spectra, supplemented by a number of newly recorded spectra obtained with infrared multiple photon dissociation spectroscopy, provide experimental verification of the preferred conformations in most cases. The overall structural features of these complexes are shown, and attention is given to comparisons involving peptide sequence, nature of the metal ion, and nature of the side-chain anchor. A regular progression is observed as a function of binding strength, whereby the weakly binding metal ions (Ba(2+) to Ca(2+)) transition from carboxylate zwitterion (ZW) binding to charge-solvated (CS) binding, while the stronger binding metal ions (Ca(2+) to Mg(2+) to Ni(2+)) transition from CS binding to metal-ion-backbone binding (Iminol) by direct metal-nitrogen bonds to the deprotonated amide nitrogens. Two new sequence-dependent reversals are found between ZW and CS binding modes, such that Ba(2+) and Ca(2+) prefer ZW binding in the GlyHis case but prefer CS binding in the HisGly case. The overall binding strength for a given metal ion is not strongly dependent on the sequence, but the histidine peptides are significantly more strongly bound (by 50-100 kJ mol(-1)) than the phenylalanine peptides.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koepf, Matthieu; Bergkamp, Jesse J.; Teillout, Anne-Lucie

The association of different metals in stable, well-defined molecular assemblies remains a great challenge of supramolecular chemistry. In such constructs, the emergence of synergism, or cooperative effects between the different metal centers is particularly intriguing. These effects can lead to uncommon reactivity or remarkable physico-chemical properties that are not otherwise achievable. For example, the association of alkaline or alkaline-earth cations and transition metals is pivotal for the activity of several biomolecules and human-made catalysts that carry out fundamental redox transformations (water oxidation, nitrogen reduction, water–gas shift reaction, etc.). In many cases the precise nature of the interactions between the alkaline-earthmore » cations and the redox-active transition metals remains elusive due to the difficulty of building stable molecular heterometallic assemblies that associate transition metals and alkaline or alkaline-earth cations in a controlled way. In this work we present the rational design of porphyrin-based ligands possessing a second binding site for alkaline-earth cations above the porphyrin macrocycle primary complexation site. We demonstrate that by using a combination of crown ether and carboxylic acid substituents suitably positioned on the periphery of the porphyrin, bitopic ligands can be obtained. The binding of calcium, a typical alkaline-earth cation, by the newly prepared ligands has been studied in detail and we show that a moderately large binding constant can be achieved in protic media using ligands that possess some degree of structural flexibility. The formation of Zn–Ca assemblies discussed in this work is viewed as a stepping stone towards the assembly of well defined molecular transition metal-alkaline earth bimetallic centers using a versatile organic scaffold.« less

Identification and partial characterization of a low affinity metal-binding site in the light chain of tetanus toxin.

PubMed

Wright, J F; Pernollet, M; Reboul, A; Aude, C; Colomb, M G

1992-05-05

Tetanus toxin was shown to contain a metal-binding site for zinc and copper. Equilibrium dialysis binding experiments using 65Zn indicated an association constant of 9-15 microM, with one zinc-binding site/toxin molecule. The zinc-binding site was localized to the toxin light chain as determined by binding of 65Zn to the light chain but not to the heavy chain after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transfer to Immobilon membranes. Copper was an efficient inhibitor of 65Zn binding to tetanus toxin and caused two peptide bond cleavages in the toxin light chain in the presence of ascorbate. These metal-catalyzed oxidative cleavages were inhibited by the presence of zinc. Partial characterization of metal-catalyzed oxidative modifications of a peptide based on a putative metal-binding site (HELIH) in the toxin light chain was used to map the metal-binding site in the protein.

Models of metal binding structures in fulvic acid from the Suwannee River, Georgia

USGS Publications Warehouse

Leenheer, J.A.; Brown, G.K.; MacCarthy, P.; Cabaniss, S.E.

1998-01-01

Fulvic acid, isolated from the Suwannee River, Georgia, was assessed for its ability to bind Ca2+, Cd2+, Cu2+, Ni2+, and Zn2+ ions at pH 6 before and after extensive fractionation that was designed to reveal the nature of metal binding functional groups. The binding constant for Ca2+ ion had the greatest increase of all the ions in a metal binding fraction that was selected for intensive characterization for the purpose of building quantitative average model structures. The 'metal binding' fraction was characterized by quantitative 13C NMR, 1H NMR, and FT-1R spectrometry and elemental, titrimetric, and molecular weight determinations. The characterization data revealed that carboxyl groups were clustered in short- chain aliphatic dibasic acid structures. The Ca2+ binding data suggested that ether-substituted oxysuccinic acid structures are good models for the metal binding sites at pH 6. Structural models were derived based upon oxidation and photolytic rearrangements of cutin, lignin, and tannin precursors. These structural models rich in substituted dibasic acid structures revealed polydentate binding sites with the potential for both inner-sphere and outer-sphere type binding. The majority of the fulvic acid molecule was involved with metal binding rather than a small substructural unit.Fulvic acid, isolated from the Suwannee River, Georgia, was assessed for its ability to bind Ca2+, Cd2+, Cu2+, Ni2+, and Zn2+ ions at pH 6 before and after extensive fractionation that was designed to reveal the nature of metal binding functional groups. The binding constant for Ca2+ ion had the greatest increase of all the ions in a metal binding fraction that was selected for intensive characterization for the purpose of building quantitative average model structures. The `metal binding' fraction was characterized by quantitative 13C NMR, 1H NMR, and FT-IR spectrometry and elemental, titrimetric, and molecular weight determinations. The characterization data revealed that carboxyl groups were clustered in short-chain aliphatic dibasic acid structures. The Ca2+ binding data suggested that ether-substituted oxysuccinic acid structures are good models for the metal binding sites at pH 6. Structural models were derived based upon oxidation and photolytic rearrangements of cutin, lignin, and tannin precursors. These structural models rich in substituted dibasic acid structures revealed polydentate binding sites with the potential for both inner-sphere and outer-sphere type binding. The majority of the fulvic acid molecule was involved with metal binding rather than a small substructural unit.

High Affinity Binding of Indium and Ruthenium Ions by Gastrins

PubMed Central

Baldwin, Graham S.; George, Graham N.; Pushie, M. Jake

2015-01-01

The peptide hormone gastrin binds two ferric ions with high affinity, and iron binding is essential for the biological activity of non-amidated forms of the hormone. Since gastrins act as growth factors in gastrointestinal cancers, and as peptides labelled with Ga and In isotopes are increasingly used for cancer diagnosis, the ability of gastrins to bind other metal ions was investigated systematically by absorption spectroscopy. The coordination structures of the complexes were characterized by extended X-ray absorption fine structure (EXAFS) spectroscopy. Changes in the absorption of gastrin in the presence of increasing concentrations of Ga3+ were fitted by a 2 site model with dissociation constants (Kd) of 3.3 x 10−7 and 1.1 x 10−6 M. Although the absorption of gastrin did not change upon the addition of In3+ ions, the changes in absorbance on Fe3+ ion binding in the presence of indium ions were fitted by a 2 site model with Kd values for In3+ of 6.5 x 10−15 and 1.7 x 10−7 M. Similar results were obtained with Ru3+ ions, although the Kd values for Ru3+ of 2.6 x 10−13 and 1.2 x 10−5 M were slightly larger than observed for In3+. The structures determined by EXAFS all had metal:gastrin stoichiometries of 2:1 but, while the metal ions in the Fe, Ga and In complexes were bridged by a carboxylate and an oxygen with a metal-metal separation of 3.0–3.3 Å, the Ru complex clearly demonstrated a short range Ru—Ru separation, which was significantly shorter, at 2.4 Å, indicative of a metal-metal bond. We conclude that gastrin selectively binds two In3+ or Ru3+ ions, and that the affinity of the first site for In3+ or Ru3+ ions is higher than for ferric ions. Some of the metal ion-gastrin complexes may be useful for cancer diagnosis and therapy. PMID:26457677

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hill, Shannon E.; Nguyen, Elaine; Ukachukwu, Chiamaka U.

Dihydroneopterin triphosphate pyrophosphatase (DHNTPase), a member of the Mg2+ dependent Nudix hydrolase superfamily, is the recently-discovered enzyme that functions in the second step of the pterin branch of the folate biosynthetic pathway in E. coli. DHNTPase is of interest because inhibition of enzymes in bacterial folate biosynthetic pathways is a strategy for antibiotic development. We determined crystal structures of DHNTPase with and without activating, Mg2+-mimicking metals Co2+ and Ni2+. Four metal ions, identified by anomalous scattering, and stoichiometrically confirmed in solution by isothermal titration calorimetry, are held in place by Glu56 and Glu60 within the Nudix sequence motif, Glu117, waters,more » and a sulfate ion, of which the latter is further stabilized by a salt bridge with Lys7. In silico docking of the DHNTP substrate reveals a binding mode in which the pterin ring moiety is nestled in a largely hydrophobic pocket, the β-phosphate activated for nucleophilic attack overlays with the crystallographic sulfate and is in line with an activated water molecule, and remaining phosphate groups are stabilized by all four identified metal ions. The structures and binding data provide new details regarding DHNTPase metal requirements, mechanism, and suggest a strategy for efficient inhibition.« less

CheckMyMetal: a macromolecular metal-binding validation tool

PubMed Central

Porebski, Przemyslaw J.

2017-01-01

Metals are essential in many biological processes, and metal ions are modeled in roughly 40% of the macromolecular structures in the Protein Data Bank (PDB). However, a significant fraction of these structures contain poorly modeled metal-binding sites. CheckMyMetal (CMM) is an easy-to-use metal-binding site validation server for macromolecules that is freely available at http://csgid.org/csgid/metal_sites. The CMM server can detect incorrect metal assignments as well as geometrical and other irregularities in the metal-binding sites. Guidelines for metal-site modeling and validation in macromolecules are illustrated by several practical examples grouped by the type of metal. These examples show CMM users (and crystallographers in general) problems they may encounter during the modeling of a specific metal ion. PMID:28291757

Structures of minute virus of mice replication initiator protein N-terminal domain: Insights into DNA nicking and origin binding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tewary, Sunil K.; Liang, Lingfei; Lin, Zihan

Members of the Parvoviridae family all encode a non-structural protein 1 (NS1) that directs replication of single-stranded viral DNA, packages viral DNA into capsid, and serves as a potent transcriptional activator. Here we report the X-ray structure of the minute virus of mice (MVM) NS1 N-terminal domain at 1.45 Å resolution, showing that sites for dsDNA binding, ssDNA binding and cleavage, nuclear localization, and other functions are integrated on a canonical fold of the histidine-hydrophobic-histidine superfamily of nucleases, including elements specific for this Protoparvovirus but distinct from its Bocaparvovirus or Dependoparvovirus orthologs. High resolution structural analysis reveals a nickase activemore » site with an architecture that allows highly versatile metal ligand binding. The structures support a unified mechanism of replication origin recognition for homotelomeric and heterotelomeric parvoviruses, mediated by a basic-residue-rich hairpin and an adjacent helix in the initiator proteins and by tandem tetranucleotide motifs in the replication origins. - Highlights: • The structure of a parvovirus replication initiator protein has been determined; • The structure sheds light on mechanisms of ssDNA binding and cleavage; • The nickase active site is preconfigured for versatile metal ligand binding; • The binding site for the double-stranded replication origin DNA is identified; • A single domain integrates multiple functions in virus replication.« less

Interaction of external alkali metal ions with the Na-K pump of human erythrocytes: a comparison of their effects on activation of the pump and on the rate of ouabain binding

PubMed Central

1978-01-01

The effects of external alkali metal ions on the rate of ouabain binding and on the rate of the Na-K pump were examined in human red blood cells. In Na-containing solutions, K, Cs, and Li decreased the rate of ouabain binding. For K and Cs, the kinetics of this effect were similar to those for their activation of the pump. In Na-free (choline- substituted) solutions the rate of ouabain binding was decreased by K whereas it was promoted by Cs and Li. External Na increased the rate of ouabain binding whether or not external K was present, and the kinetics of this effect were not the same as those for inhibition of the pump by Na. These findings are interpreted to mean that not only do the cations affect ouabain binding at the external loading sites on the pump from which ions are translocated inward, but that there are additional sites on the external aspect of the pump at which cations can promote ouabain binding, and that these sites can be occupied by Li, Na, and Cs. It is postulated that these latter sites are those from which Na is discharged after outward translocation by the pump. PMID:702113

N- vs. C-Domain Selectivity of Catalytic Inactivation of Human Angiotensin Converting Enzyme by Lisinopril-Coupled Transition Metal Chelates

PubMed Central

Hocharoen, Lalintip; Joyner, Jeff C.; Cowan, J. A.

2014-01-01

The N- and C-terminal domains of human somatic Angiotensin I Converting Enzyme (sACE-1) demonstrate distinct physiological functions, with resulting interest in the development of domain-selective inhibitors for specific therapeutic applications. Herein, the activity of lisinopril-coupled transition metal chelates were tested for both reversible binding and irreversible catalytic inactivation of sACE-1. C/N domain binding selectivity ratios ranged from 1 to 350, while rates of irreversible catalytic inactivation of the N- and C-domains were found to be significantly greater for the N-domain, suggesting a more optimal orientation of the M-chelate-lisinopril complexes within the active site of the N-domain of sACE-1. Finally, the combined effect of binding selectivity and inactivation selectivity was assessed for each catalyst (double-filter selectivity factors), and several catalysts were found to cause domain-selective catalytic inactivation. The results of this study demonstrate the ability to optimize the target selectivity of catalytic metallopeptides through both binding and orientation factors (double-filter effect). PMID:24228790

N- versus C-domain selectivity of catalytic inactivation of human angiotensin converting enzyme by lisinopril-coupled transition metal chelates.

PubMed

Hocharoen, Lalintip; Joyner, Jeff C; Cowan, J A

2013-12-27

The N- and C-terminal domains of human somatic angiotensin I converting enzyme (sACE-1) demonstrate distinct physiological functions, with resulting interest in the development of domain-selective inhibitors for specific therapeutic applications. Herein, the activity of lisinopril-coupled transition metal chelates was tested for both reversible binding and irreversible catalytic inactivation of each domain of sACE-1. C/N domain binding selectivity ratios ranged from 1 to 350, while rates of irreversible catalytic inactivation of the N- and C-domains were found to be significantly greater for the N-domain, suggesting a more optimal orientation of M-chelate-lisinopril complexes within the active site of the N-domain of sACE-1. Finally, the combined effect of binding selectivity and inactivation selectivity was assessed for each catalyst (double-filter selectivity factors), and several catalysts were found to cause domain-selective catalytic inactivation. The results of this study demonstrate the ability to optimize the target selectivity of catalytic metallopeptides through both binding and catalytic factors (double-filter effect).

Low concentrations of copper in drinking water increase AP-1 binding in the brain.

PubMed

Lung, Shyang; Li, Huihui; Bondy, Stephen C; Campbell, Arezoo

2015-12-01

Copper (Cu) in trace amounts is essential for biological organisms. However, dysregulation of the redox-active metal has been implicated in different neurological disorders such as Wilson's, Menkes', Alzheimer's, and Parkinson's diseases. Since many households use Cu tubing in the plumbing system, and corrosion causes the metal to leach into the drinking water, there may be adverse effects on the central nervous system connected with low-level chronic exposure. The present study demonstrates that treatment with a biologically relevant concentration of Cu for 3 months significantly increases activation of the redox-modulated transcription factor AP-1 in mouse brains. This was independent of an upstream kinase indicated in AP-1 activation. Another redox-active transcription factor, NF-κB, was not significantly modified by the Cu exposure. These results indicate that the effect of Cu on AP-1 is unique and may involve direct modulation of DNA binding. © The Author(s) 2012.

Structural basis for the metal-selective activation of the manganese transport regulator of Bacillus subtilis.

PubMed

Kliegman, Joseph I; Griner, Sarah L; Helmann, John D; Brennan, Richard G; Glasfeld, Arthur

2006-03-21

The manganese transport regulator (MntR) of Bacillus subtilis is activated by Mn(2+) to repress transcription of genes encoding transporters involved in the uptake of manganese. MntR is also strongly activated by cadmium, both in vivo and in vitro, but it is poorly activated by other metal cations, including calcium and zinc. The previously published MntR.Mn(2+) structure revealed a binuclear complex of manganese ions with a metal-metal separation of 3.3 A (herein designated the AB conformer). Analysis of four additional crystal forms of MntR.Mn(2+) reveals that the AB conformer is only observed in monoclinic crystals at 100 K, suggesting that this conformation may be stabilized by crystal packing forces. In contrast, monoclinic crystals analyzed at room temperature (at either pH 6.5 or pH 8.5), and a second hexagonal crystal form (analyzed at 100 K), all reveal the shift of one manganese ion by 2.5 A, thereby leading to a newly identified conformation (the AC conformer) with an internuclear distance of 4.4 A. Significantly, the cadmium and calcium complexes of MntR also contain binuclear complexes with a 4.4 A internuclear separation. In contrast, the zinc complex of MntR contains only one metal ion per subunit, in the A site. Isothermal titration calorimetry confirms the stoichiometry of Mn(2+), Cd(2+), and Zn(2+) binding to MntR. We propose that the specificity of MntR activation is tied to productive binding of metal ions at two sites; the A site appears to act as a selectivity filter, determining whether the B or C site will be occupied and thereby fully activate MntR.

Membrane interaction and functional plasticity of inositol polyphosphate 5-phosphatases.

PubMed

Braun, Werner; Schein, Catherine H

2014-05-06

In this issue of Structure, Trésaugues and colleagues determined the interaction of membrane-bound phosphoinositides with three clinically significant human inositol polyphosphate 5-phosphatases (I5Ps). A comparison to the structures determined with soluble substrates revealed differences in the binding mode and suggested how the I5Ps and apurinic endonuclease (APE1) activities evolved from the same metal-binding active center. Copyright © 2014 Elsevier Ltd. All rights reserved.

ETMB-RBF: discrimination of metal-binding sites in electron transporters based on RBF networks with PSSM profiles and significant amino acid pairs.

PubMed

Ou, Yu-Yen; Chen, Shu-An; Wu, Sheng-Cheng

2013-01-01

Cellular respiration is the process by which cells obtain energy from glucose and is a very important biological process in living cell. As cells do cellular respiration, they need a pathway to store and transport electrons, the electron transport chain. The function of the electron transport chain is to produce a trans-membrane proton electrochemical gradient as a result of oxidation-reduction reactions. In these oxidation-reduction reactions in electron transport chains, metal ions play very important role as electron donor and acceptor. For example, Fe ions are in complex I and complex II, and Cu ions are in complex IV. Therefore, to identify metal-binding sites in electron transporters is an important issue in helping biologists better understand the workings of the electron transport chain. We propose a method based on Position Specific Scoring Matrix (PSSM) profiles and significant amino acid pairs to identify metal-binding residues in electron transport proteins. We have selected a non-redundant set of 55 metal-binding electron transport proteins as our dataset. The proposed method can predict metal-binding sites in electron transport proteins with an average 10-fold cross-validation accuracy of 93.2% and 93.1% for metal-binding cysteine and histidine, respectively. Compared with the general metal-binding predictor from A. Passerini et al., the proposed method can improve over 9% of sensitivity, and 14% specificity on the independent dataset in identifying metal-binding cysteines. The proposed method can also improve almost 76% sensitivity with same specificity in metal-binding histidine, and MCC is also improved from 0.28 to 0.88. We have developed a novel approach based on PSSM profiles and significant amino acid pairs for identifying metal-binding sites from electron transport proteins. The proposed approach achieved a significant improvement with independent test set of metal-binding electron transport proteins.

ETMB-RBF: Discrimination of Metal-Binding Sites in Electron Transporters Based on RBF Networks with PSSM Profiles and Significant Amino Acid Pairs

PubMed Central

Ou, Yu-Yen; Chen, Shu-An; Wu, Sheng-Cheng

2013-01-01

Background Cellular respiration is the process by which cells obtain energy from glucose and is a very important biological process in living cell. As cells do cellular respiration, they need a pathway to store and transport electrons, the electron transport chain. The function of the electron transport chain is to produce a trans-membrane proton electrochemical gradient as a result of oxidation–reduction reactions. In these oxidation–reduction reactions in electron transport chains, metal ions play very important role as electron donor and acceptor. For example, Fe ions are in complex I and complex II, and Cu ions are in complex IV. Therefore, to identify metal-binding sites in electron transporters is an important issue in helping biologists better understand the workings of the electron transport chain. Methods We propose a method based on Position Specific Scoring Matrix (PSSM) profiles and significant amino acid pairs to identify metal-binding residues in electron transport proteins. Results We have selected a non-redundant set of 55 metal-binding electron transport proteins as our dataset. The proposed method can predict metal-binding sites in electron transport proteins with an average 10-fold cross-validation accuracy of 93.2% and 93.1% for metal-binding cysteine and histidine, respectively. Compared with the general metal-binding predictor from A. Passerini et al., the proposed method can improve over 9% of sensitivity, and 14% specificity on the independent dataset in identifying metal-binding cysteines. The proposed method can also improve almost 76% sensitivity with same specificity in metal-binding histidine, and MCC is also improved from 0.28 to 0.88. Conclusions We have developed a novel approach based on PSSM profiles and significant amino acid pairs for identifying metal-binding sites from electron transport proteins. The proposed approach achieved a significant improvement with independent test set of metal-binding electron transport proteins. PMID:23405059

Further Insights into Metal-DOM Interaction: Consideration of Both Fluorescent and Non-Fluorescent Substances

PubMed Central

Xu, Huacheng; Zhong, Jicheng; Yu, Guanghui; Wu, Jun; Jiang, Helong; Yang, Liuyan

2014-01-01

Information on metal binding with fluorescent substances has been widely studied. By contrast, information on metal binding with non-fluorescent substances remains lacking despite the dominance of these substances in aquatic systems. In this study, the metal binding properties of both fluorescent and non-fluorescent substances were investigated by using metal titration combined with two-dimensional correlation spectroscopy (2D–COS) analysis. The organic matters in the eutrophic algae-rich lake, including natural organic matters (NOM) and algae-induced extracellular polymeric substances (EPS), both contained fluorescent and non-fluorescent substances. The peaks in the one-dimensional spectra strongly overlapped, while 2D–COS can decompose the overlapped peaks and thus enhanced the spectral resolution. Moreover, 2D FTIR COS demonstrated that the binding susceptibility of organic ligands in both NOM and algal EPS matrices followed the order: 3400>1380>1650 cm−1, indicative the significant contribution of non-fluorescent ligands in metal binding. The modified Stern-Volmer equation also revealed a substantial metal binding potential for the non-fluorescent substances (logKM: 3.57∼4.92). As for the effects of organic ligands on metal binding, EPS was characterized with higher binding ability than NOM for both fluorescent and non-fluorescent ligands. Algae-induced EPS and the non-fluorescent substances in eutrophic algae-rich lakes should not be overlooked because of their high metal binding potential. PMID:25380246

Modeling metal binding to soils: the role of natural organic matter.

PubMed

Gustafsson, Jon Petter; Pechová, Pavlina; Berggren, Dan

2003-06-15

The use of mechanistically based models to simulate the solution concentrations of heavy metals in soils is complicated by the presence of different sorbents that may bind metals. In this study, the binding of Zn, Pb, Cu, and Cd by 14 different Swedish soil samples was investigated. For 10 of the soils, it was found that the Stockholm Humic Model (SHM) was able to describe the acid-base characteristics, when using the concentrations of "active" humic substances and Al as fitting parameters. Two additional soils could be modeled when ion exchange to clay was also considered, using a component additivity approach. For dissolved Zn, Cd, Ca, and Mg reasonable model fits were produced when the metal-humic complexation parameters were identical for the 12 soils modeled. However, poor fits were obtained for Pb and Cu in Aquept B horizons. In two of the soil suspensions, the Lund A and Romfartuna Bhs, the calculated speciation agreed well with results obtained by using cation-exchange membranes. The results suggest that organic matter is an important sorbent for metals in many surface horizons of soils in temperate and boreal climates, and the necessity of properly accounting for the competition from Al in simulations of dissolved metal concentrations is stressed.

Calcium-independent metal-ion catalytic mechanism of anthrax edema factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shen, Yuequan; Zhukovskaya, Natalia L.; Guo, Qing

2009-11-18

Edema factor (EF), a key anthrax exotoxin, has an anthrax protective antigen-binding domain (PABD) and a calmodulin (CaM)-activated adenylyl cyclase domain. Here, we report the crystal structures of CaM-bound EF, revealing the architecture of EF PABD. CaM has N- and C-terminal domains and each domain can bind two calcium ions. Calcium binding induces the conformational change of CaM from closed to open. Structures of the EF-CaM complex show how EF locks the N-terminal domain of CaM into a closed conformation regardless of its calcium-loading state. This represents a mechanism of how CaM effector alters the calcium affinity of CaM andmore » uncouples the conformational change of CaM from calcium loading. Furthermore, structures of EF-CaM complexed with nucleotides show that EF uses two-metal-ion catalysis, a prevalent mechanism in DNA and RNA polymerases. A histidine (H351) further facilitates the catalysis of EF by activating a water to deprotonate 3'OH of ATP. Mammalian adenylyl cyclases share no structural similarity with EF and they also use two-metal-ion catalysis, suggesting the catalytic mechanism-driven convergent evolution of two structurally diverse adenylyl cyclases.« less

DNA incision evaluation, binding investigation and biocidal screening of Cu(II), Ni(II) and Co(II) complexes with isoxazole Schiff bases.

PubMed

Ganji, Nirmala; Chityala, Vijay Kumar; Marri, Pradeep Kumar; Aveli, Rambabu; Narendrula, Vamsikrishna; Daravath, Sreenu; Shivaraj

2017-10-01

Two new series of binary metal complexes [M(L 1 ) 2 ] and [M(L 2 ) 2 ] where, M=Cu(II), Ni(II) & Co(II) and L 1 =4-((3,4-dimethylisoxazol-5-ylimino)methyl)benzene-1,3-diol; L 2 =2-((3,4-dimethylisoxazol-5-ylimino)methyl)-5-methoxyphenol were synthesized and characterized by elemental analysis, 1 H NMR, 13 C NMR, FT-IR, ESI mass, UV-Visible, magnetic moment, ESR, SEM and powder XRD studies. Based on these results, a square planar geometry is assigned for all the metal complexes where the Schiff base acts as uninegatively charged bidentate chelating agent via the hydroxyl oxygen and azomethine nitrogen atoms. DNA binding studies of all the complexes with calf thymus DNA have been comprehensively investigated using electronic absorption spectroscopy, fluorescence quenching and viscosity studies. The oxidative and photo cleavage affinity of metal complexes towards supercoiled pBR322 DNA has been ascertained by agarose gel electrophoresis assay. From the results, it is observed that all the metal complexes bind effectively to CT-DNA via an intercalative mode of binding and also cleave pBR322 DNA in a promising manner. Further the Cu(II) complexes have shown better binding and cleavage properties towards DNA. The antimicrobial activities of the Schiff bases and their metal complexes were studied on bacterial and fungal strains and the results denoted that the complexes are more potent than their Schiff base ligands. Copyright © 2017 Elsevier B.V. All rights reserved.

Zinc as Allosteric Ion Channel Modulator: Ionotropic Receptors as Metalloproteins.

PubMed

Peralta, Francisco Andrés; Huidobro-Toro, Juan Pablo

2016-07-02

Zinc is an essential metal to life. This transition metal is a structural component of many proteins and is actively involved in the catalytic activity of cell enzymes. In either case, these zinc-containing proteins are metalloproteins. However, the amino acid residues that serve as ligands for metal coordination are not necessarily the same in structural proteins compared to enzymes. While crystals of structural proteins that bind zinc reveal a higher preference for cysteine sulfhydryls rather than histidine imidazole rings, catalytic enzymes reveal the opposite, i.e., a greater preference for the histidines over cysteines for catalysis, plus the influence of carboxylic acids. Based on this paradigm, we reviewed the putative ligands of zinc in ionotropic receptors, where zinc has been described as an allosteric modulator of channel receptors. Although these receptors do not strictly qualify as metalloproteins since they do not normally bind zinc in structural domains, they do transitorily bind zinc at allosteric sites, modifying transiently the receptor channel's ion permeability. The present contribution summarizes current information showing that zinc allosteric modulation of receptor channels occurs by the preferential metal coordination to imidazole rings as well as to the sulfhydryl groups of cysteine in addition to the carboxyl group of acid residues, as with enzymes and catalysis. It is remarkable that most channels, either voltage-sensitive or transmitter-gated receptor channels, are susceptible to zinc modulation either as positive or negative regulators.

Zinc as Allosteric Ion Channel Modulator: Ionotropic Receptors as Metalloproteins

PubMed Central

Peralta, Francisco Andrés; Huidobro-Toro, Juan Pablo

2016-01-01

Zinc is an essential metal to life. This transition metal is a structural component of many proteins and is actively involved in the catalytic activity of cell enzymes. In either case, these zinc-containing proteins are metalloproteins. However, the amino acid residues that serve as ligands for metal coordination are not necessarily the same in structural proteins compared to enzymes. While crystals of structural proteins that bind zinc reveal a higher preference for cysteine sulfhydryls rather than histidine imidazole rings, catalytic enzymes reveal the opposite, i.e., a greater preference for the histidines over cysteines for catalysis, plus the influence of carboxylic acids. Based on this paradigm, we reviewed the putative ligands of zinc in ionotropic receptors, where zinc has been described as an allosteric modulator of channel receptors. Although these receptors do not strictly qualify as metalloproteins since they do not normally bind zinc in structural domains, they do transitorily bind zinc at allosteric sites, modifying transiently the receptor channel’s ion permeability. The present contribution summarizes current information showing that zinc allosteric modulation of receptor channels occurs by the preferential metal coordination to imidazole rings as well as to the sulfhydryl groups of cysteine in addition to the carboxyl group of acid residues, as with enzymes and catalysis. It is remarkable that most channels, either voltage-sensitive or transmitter-gated receptor channels, are susceptible to zinc modulation either as positive or negative regulators. PMID:27384555

Using metal-ligand binding characteristics to predict metal toxicity: quantitative ion character-activity relationships (QICARs).

PubMed Central

Newman, M C; McCloskey, J T; Tatara, C P

1998-01-01

Ecological risk assessment can be enhanced with predictive models for metal toxicity. Modelings of published data were done under the simplifying assumption that intermetal trends in toxicity reflect relative metal-ligand complex stabilities. This idea has been invoked successfully since 1904 but has yet to be applied widely in quantitative ecotoxicology. Intermetal trends in toxicity were successfully modeled with ion characteristics reflecting metal binding to ligands for a wide range of effects. Most models were useful for predictive purposes based on an F-ratio criterion and cross-validation, but anomalous predictions did occur if speciation was ignored. In general, models for metals with the same valence (i.e., divalent metals) were better than those combining mono-, di-, and trivalent metals. The softness parameter (sigma p) and the absolute value of the log of the first hydrolysis constant ([symbol: see text] log KOH [symbol: see text]) were especially useful in model construction. Also, delta E0 contributed substantially to several of the two-variable models. In contrast, quantitative attempts to predict metal interactions in binary mixtures based on metal-ligand complex stabilities were not successful. PMID:9860900

FINDSITE-metal: Integrating evolutionary information and machine learning for structure-based metal binding site prediction at the proteome level

PubMed Central

Brylinski, Michal; Skolnick, Jeffrey

2010-01-01

The rapid accumulation of gene sequences, many of which are hypothetical proteins with unknown function, has stimulated the development of accurate computational tools for protein function prediction with evolution/structure-based approaches showing considerable promise. In this paper, we present FINDSITE-metal, a new threading-based method designed specifically to detect metal binding sites in modeled protein structures. Comprehensive benchmarks using different quality protein structures show that weakly homologous protein models provide sufficient structural information for quite accurate annotation by FINDSITE-metal. Combining structure/evolutionary information with machine learning results in highly accurate metal binding annotations; for protein models constructed by TASSER, whose average Cα RMSD from the native structure is 8.9 Å, 59.5% (71.9%) of the best of top five predicted metal locations are within 4 Å (8 Å) from a bound metal in the crystal structure. For most of the targets, multiple metal binding sites are detected with the best predicted binding site at rank 1 and within the top 2 ranks in 65.6% and 83.1% of the cases, respectively. Furthermore, for iron, copper, zinc, calcium and magnesium ions, the binding metal can be predicted with high, typically 70-90%, accuracy. FINDSITE-metal also provides a set of confidence indexes that help assess the reliability of predictions. Finally, we describe the proteome-wide application of FINDSITE-metal that quantifies the metal binding complement of the human proteome. FINDSITE-metal is freely available to the academic community at http://cssb.biology.gatech.edu/findsite-metal/. PMID:21287609

Non-thiolate ligation of nickel by nucleotide-free UreG of Klebsiella aerogenes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Martin-Diaconescu, Vlad; Joseph, Crisjoe A.; Boer, Jodi L.

Nickel-dependent ureases are activated by a multiprotein complex that includes the GTPase UreG. Prior studies showed that nucleotide-free UreG from Klebsiella aerogenes is monomeric and binds one nickel or zinc ion with near-equivalent affinity using an undefined binding site, whereas nucleotide-free UreG from Helicobacter pylori selectively binds one zinc ion per dimer via a universally conserved Cys-Pro-His motif in each protomer. Iodoacetamide-treated K. aerogenes UreG was nearly unaffected in nickel binding compared to non-treated sample, suggesting the absence of thiolate ligands to the metal. X-ray absorption spectroscopy of nickel-bound UreG showed the metal possessed four-coordinate geometry with all O/N donormore » ligands including one imidazole, thus confirming the absence of thiolate ligation. The nickel site in Strep-tag II-modified protein possessed six-coordinate geometry, again with all O/N donor ligands, but now including two or three imidazoles. An identical site was noted for the Strep-tag II-modified H74A variant, substituted in the Cys-Pro-His motif, ruling out coordination by this His residue. These results are consistent with metal binding to both His6 and a His residue of the fusion peptide in Strep-tagged K. aerogenes UreG. We conclude that the nickel- and zinc-binding site in nucleotide-free K. aerogenes UreG is distinct from that of nucleotide-free H. pylori UreG and does not involve the Cys-Pro-His motif. Further, we show the Strep-tag II can perturb metal coordination of this protein.« less

The role of chloride in the mechanism of O(2) activation at the mononuclear nonheme Fe(II) center of the halogenase HctB.

PubMed

Pratter, Sarah M; Light, Kenneth M; Solomon, Edward I; Straganz, Grit D

2014-07-02

Mononuclear nonheme Fe(II) (MNH) and α-ketoglutarate (α-KG) dependent halogenases activate O2 to perform oxidative halogenations of activated and nonactivated carbon centers. While the mechanism of halide incorporation into a substrate has been investigated, the mechanism by which halogenases prevent oxidations in the absence of chloride is still obscure. Here, we characterize the impact of chloride on the metal center coordination and reactivity of the fatty acyl-halogenase HctB. Stopped-flow kinetic studies show that the oxidative transformation of the Fe(II)-α-KG-enzyme complex is >200-fold accelerated by saturating concentrations of chloride in both the absence and presence of a covalently bound substrate. By contrast, the presence of substrate, which generally brings about O2 activation at enzymatic MNH centers, only has an ∼10-fold effect in the absence of chloride. Circular dichroism (CD) and magnetic CD (MCD) studies demonstrate that chloride binding triggers changes in the metal center ligation: chloride binding induces the proper binding of the substrate as shown by variable-temperature, variable-field (VTVH) MCD studies of non-α-KG-containing forms and the conversion from six-coordinate (6C) to 5C/6C mixtures when α-KG is bound. In the presence of substrate, a site with square pyramidal five-coordinate (5C) geometry is observed, which is required for O2 activation at enzymatic MNH centers. In the absence of substrate an unusual trigonal bipyramidal site is formed, which accounts for the observed slow, uncoupled reactivity. Molecular dynamics simulations suggest that the binding of chloride to the metal center of HctB leads to a conformational change in the enzyme that makes the active site more accessible to the substrate and thus facilitates the formation of the catalytically competent enzyme-substrate complex. Results are discussed in relation to other MNH dependent halogenases.

Capture of Pb2+ and Cu2+ Metal Cations by Neisseria meningitidis-type Capsular Polysaccharides.

PubMed

Ghimire, Sujan; McCarthy, Pumtiwitt C

2018-05-05

Heavy metal pollution of water is a significant environmental and public health concern. Current biological strategies for heavy metal removal from water are performed using microbial biopolymers, including polysaccharides, that are already fully formed. This creates limitations in adapting polysaccharides to increase binding affinity for specific metals. We propose that altering the specificity of polysaccharide-producing enzymes could be beneficial to improving metal capture by modified polysaccharides. We assess binding of Cu 2+ and Pb 2+ metal cations to Neisseria meningitidis -type polysaccharides. All concentrations of metal cations tested were able to completely bind to colominic acid. This polymer is equivalent to the capsular polysaccharide of N. meningitidis serogroup B comprised of a homopolymer of negatively charged sialic acid. There was slightly less binding observed with N. meningitidis serogroup W, which contains repeating units of the neutral sugar galactose and sialic acid. Our work represents the first assessment of the metal-binding properties of these capsular polysaccharides. Future work will seek to optimize metal-binding with Neisseria meningitidis serogroup W polysaccharide.

Evolution of Metal(Loid) Binding Sites in Transcriptional Regulators

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ordonez, E.; Thiyagarajan, S.; Cook, J.D.

2009-05-22

Expression of the genes for resistance to heavy metals and metalloids is transcriptionally regulated by the toxic ions themselves. Members of the ArsR/SmtB family of small metalloregulatory proteins respond to transition metals, heavy metals, and metalloids, including As(III), Sb(III), Cd(II), Pb(II), Zn(II), Co(II), and Ni(II). These homodimeric repressors bind to DNA in the absence of inducing metal(loid) ion and dissociate from the DNA when inducer is bound. The regulatory sites are often three- or four-coordinate metal binding sites composed of cysteine thiolates. Surprisingly, in two different As(III)-responsive regulators, the metalloid binding sites were in different locations in the repressor, andmore » the Cd(II) binding sites were in two different locations in two Cd(II)-responsive regulators. We hypothesize that ArsR/SmtB repressors have a common backbone structure, that of a winged helix DNA-binding protein, but have considerable plasticity in the location of inducer binding sites. Here we show that an As(III)-responsive member of the family, CgArsR1 from Corynebacterium glutamicum, binds As(III) to a cysteine triad composed of Cys{sup 15}, Cys{sup 16}, and Cys{sup 55}. This binding site is clearly unrelated to the binding sites of other characterized ArsR/SmtB family members. This is consistent with our hypothesis that metal(loid) binding sites in DNA binding proteins evolve convergently in response to persistent environmental pressures.« less

Isolation of copper-binding proteins from activated sludge culture.

PubMed

Fukushi, K; Kato, S; Antsuki, T; Omura, T

2001-01-01

Six copper-binding microbial proteins were isolated from activated sludge cultures grown on media containing copper at various concentrations. Molecular weights among isolated proteins were ranged from 1.3k to 1 74k dalton. Isolated proteins were compared for their copper binding capabilities. Proteins isolated from cultures grown in the presence of copper in the growth media exhibited higher copper binding capabilities than those isolated from the culture grown in the absence of copper. The highest metal uptake of 61.23 (mol copper/mol protein) was observed by a protein isolated from a culture grown with copper at a concentration of 0.25 mM. This isolated protein (CBP2) had a molecular weight of 24k dalton. Other protein exhibited copper binding capability of 4.8-32.5 (mol copper/mol protein).

Synthesis, spectroscopic, molecular orbital calculation, cytotoxic, molecular docking of DNA binding and DNA cleavage studies of transition metal complexes with N-benzylidene-N'-salicylidene-1,1-diaminopropane

NASA Astrophysics Data System (ADS)

Al-Mogren, Muneerah M.; Alaghaz, Abdel-Nasser M. A.; Elbohy, Salwa A. H.

2013-10-01

Eight mononuclear chromium(III), manganese(II), iron(III), cobalt(II), nickel(II), copper(II), zinc(II) and cadmium(II) complexes of Schiff's base ligand were synthesized and determined by different physical techniques. The complexes are insoluble in common organic solvents but soluble in DMF and DMSO. The measured molar conductance values in DMSO indicate that the complexes are non-electrolytic in nature. All the eight metal complexes have been fully characterized with the help of elemental analyses, molecular weights, molar conductance values, magnetic moments and spectroscopic data. The analytical data helped to elucidate the structure of the metal complexes. The Schiff base is found to act as tridentate ligand using N2O donor set of atoms leading to an octahedral geometry for the complexes around all the metal ions. Quantum chemical calculations were performed with semi-empirical method to find the optimum geometry of the ligand and its complexes. Additionally in silico, the docking studies and the calculated pharmacokinetic parameters show promising futures for application of the ligand and complexes as high potency agents for DNA binding activity. The interaction of the complexes with calf thymus DNA (CT-DNA) has been investigated by UV absorption method, and the mode of CT-DNA binding to the complexes has been explored. Furthermore, the DNA cleavage activity by the complexes was performed. The Schiff base and their complexes have been screened for their antibacterial activity against bacterial strains [Staphylococcus aureus (RCMB010027), Staphylococcus epidermidis (RCMB010024), Bacillis subtilis (RCMB010063), Proteous vulgaris (RCMB 010085), Klebsiella pneumonia (RCMB 010093) and Shigella flexneri (RCMB 0100542)] and fungi [(Aspergillus fumigates (RCMB 02564), Aspergillus clavatus (RCMB 02593) and Candida albicans (RCMB05035)] by disk diffusion method. All the metal complexes have potent biocidal activity than the free ligand.

The Molybdenum Cofactor Biosynthetic Protein Cnx1 Complements Molybdate-Repairable Mutants, Transfers Molybdenum to the Metal Binding Pterin, and Is Associated with the Cytoskeleton

PubMed Central

Schwarz, Günter; Schulze, Jutta; Bittner, Florian; Eilers, Thomas; Kuper, Jochen; Bollmann, Gabriele; Nerlich, Andrea; Brinkmann, Henner; Mendel, Ralf R.

2000-01-01

Molybdenum (Mo) plays an essential role in the active site of all eukaryotic Mo-containing enzymes. In plants, Mo enzymes are important for nitrate assimilation, phytohormone synthesis, and purine catabolism. Mo is bound to a unique metal binding pterin (molybdopterin [MPT]), thereby forming the active Mo cofactor (Moco), which is highly conserved in eukaryotes, eubacteria, and archaebacteria. Here, we describe the function of the two-domain protein Cnx1 from Arabidopsis in the final step of Moco biosynthesis. Cnx1 is constitutively expressed in all organs and in plants grown on different nitrogen sources. Mo-repairable cnxA mutants from Nicotiana plumbaginifolia accumulate MPT and show altered Cnx1 expression. Transformation of cnxA mutants and the corresponding Arabidopsis chl-6 mutant with cnx1 cDNA resulted in functional reconstitution of their Moco deficiency. We also identified a point mutation in the Cnx1 E domain of Arabidopsis chl-6 that causes the molybdate-repairable phenotype. Recombinant Cnx1 protein is capable of synthesizing Moco. The G domain binds and activates MPT, whereas the E domain is essential for activating Mo. In addition, Cnx1 binds to the cytoskeleton in the same way that its mammalian homolog gephyrin does in neuronal cells, which suggests a hypothetical model for anchoring the Moco-synthetic machinery by Cnx1 in plant cells. PMID:11148290

Inhibition of cyclin-dependent kinase CDK1 by oxindolimine ligands and corresponding copper and zinc complexes.

PubMed

Miguel, Rodrigo Bernardi; Petersen, Philippe Alexandre Divina; Gonzales-Zubiate, Fernando A; Oliveira, Carla Columbano; Kumar, Naresh; do Nascimento, Rafael Rodrigues; Petrilli, Helena Maria; da Costa Ferreira, Ana Maria

2015-10-01

Oxindolimine-copper(II) and zinc(II) complexes that previously have shown to induce apoptosis, with DNA and mitochondria as main targets, exhibit here significant inhibition of kinase CDK1/cyclin B protein. Copper species are more active than the corresponding zinc, and the free ligand shows to be less active, indicating a major influence of coordination in the process, and a further modulation by the coordinated ligand. Molecular docking and classical molecular dynamics provide a better understanding of the effectiveness and kinase inhibition mechanism by these compounds, showing that the metal complex provides a stronger interaction than the free ligand with the ATP-binding site. The metal ion introduces charge in the oxindole species, giving it a more rigid conformation that then becomes more effective in its interactions with the protein active site. Analogous experiments resulted in no significant effect regarding phosphatase inhibition. These results can explain the cytotoxicity of these metal complexes towards different tumor cells, in addition to its capability of binding to DNA, and decreasing membrane potential of mitochondria.

Introducing a 2-His-1-Glu Nonheme Iron Center into Myoglobin Confers Nitric Oxide Reductase Activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Y.W.; Robinson, H.; Yeung, N.

2010-07-28

A conserved 2-His-1-Glu metal center, as found in natural nonheme iron-containing enzymes, was engineered into sperm whale myoglobin by replacing Leu29 and Phe43 with Glu and His, respectively (swMb L29E, F43H, H64, called Fe{sub B}Mb(-His)). A high resolution (1.65 {angstrom}) crystal structure of Cu(II)-CN?-Fe{sub B}Mb(-His) was determined, demonstrating that the unique 2-His-1-Glu metal center was successfully created within swMb. The Fe{sub B}Mb(-His) can bind Cu, Fe, or Zn ions, with both Cu(I)-Fe{sub B}Mb(-His) and Fe(II)-Fe{sub B}Mb(-His) exhibiting nitric oxide reductase (NOR) activities. Cu dependent NOR activity was significantly higher than that of Fe in the same metal binding site. EPRmore » studies showed that the reduction of NO to N{sub 2}O catalyzed by these two enzymes resulted in different intermediates; a five-coordinate heme-NO species was observed for Cu(I)-Fe{sub B}Mb(-His) due to the cleavage of the proximal heme Fe-His bond, while Fe(II)-Fe{sub B}Mb(-His) remained six-coordinate. Therefore, both the metal ligand, Glu29, and the metal itself, Cu or Fe, play crucial roles in NOR activity. This study presents a novel protein model of NOR and provides insights into a newly discovered member of the NOR family, gNOR.« less

Toxic metals (Ni2+, Pb2+, Hg2+) binding affinity of dissolved organic matter (DOM) derived from different ages municipal landfill leachate

NASA Astrophysics Data System (ADS)

Rikta, S. Y.; Tareq, Shafi M.; Uddin, M. Khabir

2018-03-01

Solid waste production is rapidly increasing in Bangladesh and landfill leachate is the consequence of the decomposition of this waste. These leachates contain heavy metals and significant amount of dissolved organic matter (DOM). DOM is known to have considerable role in heavy metals speciation. Hence, it is important to characterize DOM/leachate and evaluate toxic metals binding affinity of DOM. The objectives of this study were to characterize the DOM in landfill leachate through physico-chemical and optical analyses and to investigate the toxic metals (Ni2+, Pb2+ and Hg2+) binding affinity of three different ages (fresh sample L-1, young sample L-2 and mature sample L-3) DOM samples. Results suggested that leachate is a potential pollutant which contained very high organic pollutant load. Conditional stability constant (Log K) and percentages of fluorophores that correspond to metal binding (% f) values indicated that young DOM sample (L-2) had the highest binding affinity to all the three metals ions. In general, DOM samples showed the following order affinity to the metal ions; Ni2+ binding affinity: L-2 > L-3 > L-1, Pb2+ binding affinity: L-2 > L-3 > L-1 and Hg2+ binding affinity: L-2 > L-1 > L-3.

Validating metal binding sites in macromolecule structures using the CheckMyMetal web server

PubMed Central

Zheng, Heping; Chordia, Mahendra D.; Cooper, David R.; Chruszcz, Maksymilian; Müller, Peter; Sheldrick, George M.

2015-01-01

Metals play vital roles in both the mechanism and architecture of biological macromolecules. Yet structures of metal-containing macromolecules where metals are misidentified and/or suboptimally modeled are abundant in the Protein Data Bank (PDB). This shows the need for a diagnostic tool to identify and correct such modeling problems with metal binding environments. The "CheckMyMetal" (CMM) web server (http://csgid.org/csgid/metal_sites/) is a sophisticated, user-friendly web-based method to evaluate metal binding sites in macromolecular structures in respect to 7350 metal binding sites observed in a benchmark dataset of 2304 high resolution crystal structures. The protocol outlines how the CMM server can be used to detect geometric and other irregularities in the structures of metal binding sites and alert researchers to potential errors in metal assignment. The protocol also gives practical guidelines for correcting problematic sites by modifying the metal binding environment and/or redefining metal identity in the PDB file. Several examples where this has led to meaningful results are described in the anticipated results section. CMM was designed for a broad audience—biomedical researchers studying metal-containing proteins and nucleic acids—but is equally well suited for structural biologists to validate new structures during modeling or refinement. The CMM server takes the coordinates of a metal-containing macromolecule structure in the PDB format as input and responds within a few seconds for a typical protein structure modeled with a few hundred amino acids. PMID:24356774

Modulation of individual steps in group I intron catalysis by a peripheral metal ion.

PubMed

Forconi, Marcello; Piccirilli, Joseph A; Herschlag, Daniel

2007-10-01

Enzymes are complex macromolecules that catalyze chemical reactions at their active sites. Important information about catalytic interactions is commonly gathered by perturbation or mutation of active site residues that directly contact substrates. However, active sites are engaged in intricate networks of interactions within the overall structure of the macromolecule, and there is a growing body of evidence about the importance of peripheral interactions in the precise structural organization of the active site. Here, we use functional studies, in conjunction with published structural information, to determine the effect of perturbation of a peripheral metal ion binding site on catalysis in a well-characterized catalytic RNA, the Tetrahymena thermophila group I ribozyme. We perturbed the metal ion binding site by site-specifically introducing a phosphorothioate substitution in the ribozyme's backbone, replacing the native ligands (the pro-R (P) oxygen atoms at positions 307 and 308) with sulfur atoms. Our data reveal that these perturbations affect several reaction steps, including the chemical step, despite the absence of direct contacts of this metal ion with the atoms involved in the chemical transformation. As structural probing with hydroxyl radicals did not reveal significant change in the three-dimensional structure upon phosphorothioate substitution, the effects are likely transmitted through local, rather subtle conformational rearrangements. Addition of Cd(2+), a thiophilic metal ion, rescues some reaction steps but has deleterious effects on other steps. These results suggest that native interactions in the active site may have been aligned by the naturally occurring peripheral residues and interactions to optimize the overall catalytic cycle.

Novel calcium recognition constructions in proteins: Calcium blade and EF-hand zone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Denesyuk, Alexander I., E-mail: adenesyu@abo.fi; Institute for Biological Instrumentation of the Russian Academy of Sciences, Pushchino 142290; Permyakov, Sergei E.

Metal ions can regulate various cell processes being first, second or third messengers, and some of them, especially transition metal ions, take part in catalysis in many enzymes. As an intracellular ion, Ca{sup 2+} is involved in many cellular functions from fertilization and contraction, cell differentiation and proliferation, to apoptosis and cancer. Here, we have identified and described two novel calcium recognition environments in proteins: the calcium blade zone and the EF-hand zone, common to 12 and 8 different protein families, respectively. Each of the two environments contains three distinct structural elements: (a) the well-known characteristic Dx[DN]xDG motif; (b) anmore » adjacent structurally identical segment, which binds metal ion in the same way between the calcium blade zone and the EF-hand zone; and (c) the following structurally variable segment, which distinguishes the calcium blade zone from the EF-hand zone. Both zones have sequence insertions between the last residue of the zone and calcium-binding residues in positions V or VI. The long insertion often connects the active and the calcium-binding sites in proteins. Using the structurally identical segments as an anchor, we were able to construct the classical calmodulin type EF-hand calcium-binding site out of two different calcium-binding motifs from two unrelated proteins.« less

Development of metal-chelating inhibitors for the Class II fructose 1,6-bisphosphate (FBP) aldolase.

PubMed

Labbé, Geneviève; Krismanich, Anthony P; de Groot, Sarah; Rasmusson, Timothy; Shang, Muhong; Brown, Matthew D R; Dmitrienko, Gary I; Guillemette, J Guy

2012-07-01

It has long been suggested that the essential and ubiquitous enzyme fructose 1,6-bisphosphate (FBP) aldolase could be a good drug target against bacteria and fungi, since lower organisms possess a metal-dependant (Class II) FBP aldolase, as opposed to higher organisms which possess a Schiff-base forming (Class I) FBP aldolase. We have tested the capacity of derivatives of the metal-chelating compound dipicolinic acid (DPA), as well a thiol-containing compound, to inhibit purified recombinant Class II FBP aldolases from Mycobacterium tuberculosis, Pseudomonas aeruginosa, Bacillus cereus, Bacillus anthracis, and from the Rice Blast causative agent Magnaporthe grisea. The aldolase from M. tuberculosis was the most sensitive to the metal-chelating inhibitors, with an IC(50) of 5.2 μM with 2,3-dimercaptopropanesulfonate (DMPS) and 28 μM with DPA. DMPS and the synthesized inhibitor 6-(phosphonomethyl)picolinic acid inhibited the enzyme in a time-dependent, competitive fashion, with second order rate constants of 273 and 270 M(-1) s(-1) respectively for the binding of these compounds to the M. tuberculosis aldolase's active site in the presence of the substrate FBP (K(M) 27.9 μM). The most potent first generation inhibitors were modeled into the active site of the M. tuberculosis aldolase structure, with results indicating that the metal chelators tested cannot bind the catalytic zinc in a bidentate fashion while it remains in its catalytic location, and that most enzyme-ligand interactions involve the phosphate binding pocket residues. Copyright © 2012 Elsevier Inc. All rights reserved.

Current Understanding of the Binding Sites, Capacity, Affinity, and Biological Significance of Metals in Melanin

PubMed Central

Hong, Lian; Simon, John D.

2008-01-01

Metal chelation is often invoked as one of the main biological functions of melanin. In order to understand the interaction between metals and melanin, extensive studies have been carried out to determine the nature of the metal binding sites, binding capacity and affinity. These data are central to efforts aimed at elucidating the role metal binding plays in determining the physical, structural, biological, and photochemical properties of melanin. This article examines the current state of understanding of this field. PMID:17580858

Metal complexes as DNA intercalators.

PubMed

Liu, Hong-Ke; Sadler, Peter J

2011-05-17

DNA has a strong affinity for many heterocyclic aromatic dyes, such as acridine and its derivatives. Lerman in 1961 first proposed intercalation as the source of this affinity, and this mode of DNA binding has since attracted considerable research scrutiny. Organic intercalators can inhibit nucleic acid synthesis in vivo, and they are now common anticancer drugs in clinical therapy. The covalent attachment of organic intercalators to transition metal coordination complexes, yielding metallointercalators, can lead to novel DNA interactions that influence biological activity. Metal complexes with σ-bonded aromatic side arms can act as dual-function complexes: they bind to DNA both by metal coordination and through intercalation of the attached aromatic ligand. These aromatic side arms introduce new modes of DNA binding, involving mutual interactions of functional groups held in close proximity. The biological activity of both cis- and trans-diamine Pt(II) complexes is dramatically enhanced by the addition of σ-bonded intercalators. We have explored a new class of organometallic "piano-stool" Ru(II) and Os(II) arene anticancer complexes of the type [(η(6)-arene)Ru/Os(XY)Cl](+). Here XY is, for example, ethylenediamine (en), and the arene ligand can take many forms, including tetrahydroanthracene, biphenyl, or p-cymene. Arene-nucleobase stacking interactions can have a significant influence on both the kinetics and thermodynamics of DNA binding. In particular, the cytotoxic activity, conformational distortions, recognition by DNA-binding proteins, and repair mechanisms are dependent on the arene. A major difficulty in developing anticancer drugs is cross-resistance, a phenomenon whereby a cell that is resistant to one drug is also resistant to another drug in the same class. These new complexes are non-cross-resistant with cisplatin towards cancer cells: they constitute a new class of anticancer agents, with a mechanism of action that differs from the anticancer drug cisplatin and its analogs. The Ru-arene complexes with dual functions are more potent towards cancer cells than their nonintercalating analogs. In this Account, we focus on recent studies of dual-function organometallic Ru(II)- and Os(II)-arene complexes and the methods used to detect arene-DNA intercalation. We relate these interactions to the mechanism of anticancer activity and to structure-activity relationships. The interactions between these complexes and DNA show close similarities to those of covalent polycyclic aromatic carcinogens, especially to N7-alkylating intercalation compounds. However, Ru-arene complexes exhibit some new features. Classical intercalation and base extrusion next to the metallated base is observed for {(η(6)-biphenyl)Ru(ethylenediamine)}(2+) adducts of a 14-mer duplex, while penetrating arene intercalation occurs for adducts of the nonaromatic bulky intercalator {(η(6)-tetrahydroanthracene)Ru(ethylenediamine)}(2+) with a 6-mer duplex. The introduction of dual-function Ru-arene complexes introduces new mechanisms of antitumor activity, novel mechanisms for attack on DNA, and new concepts for developing structure- activity relationships. We hope this discussion will stimulate thoughtful and focused research on the design of anticancer chemotherapeutic agents using these unique approaches.

The relationship between metal toxicity and biotic ligand binding affinities in aquatic and soil organisms: a review.

PubMed

Ardestani, Masoud M; van Straalen, Nico M; van Gestel, Cornelis A M

2014-12-01

The biotic ligand model (BLM) is a theoretical, potentially mechanistic approach to assess metal bioavailability in soil and aquatic systems. In a BLM, toxicity is linked to the fraction of biotic ligand occupied, which in turn, depends on the various components of the solution, including activity of the metal. Bioavailability is a key factor in determining toxicity and uptake of metals in organisms. In this study, the present status of BLM development for soil and aquatic organisms is summarized. For all species and all metals, toxicity was correlated with the conditional biotic ligand binding constants. For almost all organisms, values for Ag, Cu, and Cd were higher than those for Zn and Ni. The constants derived for aquatic systems seem to be equally valid for soil organisms, but in the case of soils, bioavailability from the soil solution is greatly influenced by the presence of the soil solid phase. Copyright © 2014 Elsevier Ltd. All rights reserved.

Metal Binding Studies and EPR Spectroscopy of the Manganese Transport Regulator MntR†

PubMed Central

Golynskiy, Misha V.; Gunderson, William A.; Hendrich, Michael P.; Cohen, Seth M.

2007-01-01

Manganese transport regulator (MntR) is a member of the diphtheria toxin repressor (DtxR) family of transcription factors that is responsible for manganese homeostasis in Bacillus subtilis. Prior biophysical studies have focused on the metal-mediated DNA binding of MntR [Lieser, S. A., Davis, T. C., Helmann, J. D., and Cohen, S. M. (2003) Biochemistry 42, 12634-12642], as well as metal stabilization of the MntR structure [Golynskiy, M. V., Davis, T. C., Helmann, J. D., and Cohen, S. M. (2005) Biochemistry 44, 3380-3389], but only limited data on the metal-binding affinities for MntR are available. Herein, the metal-binding affinities of MntR were determined by using electron paramagnetic resonance (EPR) spectroscopy, as well as competition experiments with the fluorimetric dyes Fura-2 and Mag-fura-2. MntR was not capable of competing with Fura-2 for the binding of transition metal ions. Therefore, the metal-binding affinities and stoichiometries of Mag-fura-2 for Mn2+, Co2+, Ni2+, Zn2+, and Cd2+ were determined and utilized in MntR/Mag-fura-2 competition experiments. The measured Kd values for MntR metal binding are comparable to those reported for DtxR metal binding [Kd from 10-7 to 10-4 M; D’Aquino, J. A., et al. (2005) Proc. Natl. Acad. Sci. U.S.A. 102, 18408-18413], AntR [a homologue from Bacillus anthracis; Sen, K. I. et al. (2006) Biochemistry 45, 4295-4303], and generally follow the Irving-Williams series. Direct detection of the dinuclear Mn2+ site in MntR with EPR spectroscopy is presented, and the exchange interaction was determined, J = -0.2 cm-1. This value is lower in magnitude than most known dinuclear Mn2+ sites in proteins and synthetic complexes and is consistent with a dinuclear Mn2+ site with a longer Mn···Mn distance (4.4 Å) observed in some of the available crystal structures. MntR is found to have a surprisingly low binding affinity (∼160 μM) for its cognate metal ion Mn2+. Moreover, the results of DNA binding studies in the presence of limiting metal ion concentrations were found to be consistent with the measured metal-binding constants. The metal-binding affinities of MntR reported here help to elucidate the regulatory mechanism of this metal-dependent transcription factor. PMID:17176058

Staphylococcal surface display of metal-binding polyhistidyl peptides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Samuelson, P.; Wernerus, H.; Svedberg, M.

2000-03-01

Recombinant Staphylococcus xylosus and Staphylococcus carnosus strains were generated with surface-exposed chimeric proteins containing polyhistidyl peptides designed for binding to divalent metal ions. Surface accessibility of the chimeric surface proteins was demonstrated and the chimeric surface proteins were found to be functional in terms of metal binding, since the recombinant staphylococcal cells were shown to have gained Ni{sup 2+}- and Cd{sup 2+}-binding capacity, suggesting that such bacteria could find use in bioremediation of heavy metals. This is, to their knowledge, the first time that recombinant, surface-exposed metal-binding peptides have been expressed on gram-positive bacteria. Potential environmental or biosensor applications formore » such recombinant staphylococci as biosorbents are discussed.« less

Stereoselective hydrogenation of olefins using rhodium-substituted carbonic anhydrase--a new reductase.

PubMed

Jing, Qing; Okrasa, Krzysztof; Kazlauskas, Romas J

2009-01-01

One useful synthetic reaction missing from nature's toolbox is the direct hydrogenation of substrates using hydrogen. Instead nature uses cofactors like NADH to reduce organic substrates, which adds complexity and cost to these reductions. To create an enzyme that can directly reduce organic substrates with hydrogen, researchers have combined metal hydrogenation catalysts with proteins. One approach is an indirect link where a ligand is linked to a protein and the metal binds to the ligand. Another approach is direct linking of the metal to protein, but nonspecific binding of the metal limits this approach. Herein, we report a direct hydrogenation of olefins catalyzed by rhodium(I) bound to carbonic anhydrase (CA-[Rh]). We minimized nonspecific binding of rhodium by replacing histidine residues on the protein surface using site-directed mutagenesis or by chemically modifying the histidine residues. Hydrogenation catalyzed by CA-[Rh] is slightly slower than for uncomplexed rhodium(I), but the protein environment induces stereoselectivity favoring cis- over trans-stilbene by about 20:1. This enzyme is the first cofactor-independent reductase that reduces organic molecules using hydrogen. This catalyst is a good starting point to create variants with tailored reactivity and selectivity. This strategy to insert transition metals in the active site of metalloenzymes opens opportunities to a wider range of enzyme-catalyzed reactions.

Metal mixture modeling evaluation project: 2. Comparison of four modeling approaches.

PubMed

Farley, Kevin J; Meyer, Joseph S; Balistrieri, Laurie S; De Schamphelaere, Karel A C; Iwasaki, Yuichi; Janssen, Colin R; Kamo, Masashi; Lofts, Stephen; Mebane, Christopher A; Naito, Wataru; Ryan, Adam C; Santore, Robert C; Tipping, Edward

2015-04-01

As part of the Metal Mixture Modeling Evaluation (MMME) project, models were developed by the National Institute of Advanced Industrial Science and Technology (Japan), the US Geological Survey (USA), HDR|HydroQual (USA), and the Centre for Ecology and Hydrology (United Kingdom) to address the effects of metal mixtures on biological responses of aquatic organisms. A comparison of the 4 models, as they were presented at the MMME workshop in Brussels, Belgium (May 2012), is provided in the present study. Overall, the models were found to be similar in structure (free ion activities computed by the Windermere humic aqueous model [WHAM]; specific or nonspecific binding of metals/cations in or on the organism; specification of metal potency factors or toxicity response functions to relate metal accumulation to biological response). Major differences in modeling approaches are attributed to various modeling assumptions (e.g., single vs multiple types of binding sites on the organism) and specific calibration strategies that affected the selection of model parameters. The models provided a reasonable description of additive (or nearly additive) toxicity for a number of individual toxicity test results. Less-than-additive toxicity was more difficult to describe with the available models. Because of limitations in the available datasets and the strong interrelationships among the model parameters (binding constants, potency factors, toxicity response parameters), further evaluation of specific model assumptions and calibration strategies is needed. © 2014 SETAC.

An active site rearrangement within the Tetrahymena group I ribozyme releases nonproductive interactions and allows formation of catalytic interactions

PubMed Central

Sengupta, Raghuvir N.; Van Schie, Sabine N.S.; Giambaşu, George; Dai, Qing; Yesselman, Joseph D.; York, Darrin; Piccirilli, Joseph A.; Herschlag, Daniel

2016-01-01

Biological catalysis hinges on the precise structural integrity of an active site that binds and transforms its substrates and meeting this requirement presents a unique challenge for RNA enzymes. Functional RNAs, including ribozymes, fold into their active conformations within rugged energy landscapes that often contain misfolded conformers. Here we uncover and characterize one such “off-pathway” species within an active site after overall folding of the ribozyme is complete. The Tetrahymena group I ribozyme (E) catalyzes cleavage of an oligonucleotide substrate (S) by an exogenous guanosine (G) cofactor. We tested whether specific catalytic interactions with G are present in the preceding E•S•G and E•G ground-state complexes. We monitored interactions with G via the effects of 2′- and 3′-deoxy (–H) and −amino (–NH2) substitutions on G binding. These and prior results reveal that G is bound in an inactive configuration within E•G, with the nucleophilic 3′-OH making a nonproductive interaction with an active site metal ion termed MA and with the adjacent 2′-OH making no interaction. Upon S binding, a rearrangement occurs that allows both –OH groups to contact a different active site metal ion, termed MC, to make what are likely to be their catalytic interactions. The reactive phosphoryl group on S promotes this change, presumably by repositioning the metal ions with respect to G. This conformational transition demonstrates local rearrangements within an otherwise folded RNA, underscoring RNA's difficulty in specifying a unique conformation and highlighting Nature's potential to use local transitions of RNA in complex function. PMID:26567314

An active site rearrangement within the Tetrahymena group I ribozyme releases nonproductive interactions and allows formation of catalytic interactions.

PubMed

Sengupta, Raghuvir N; Van Schie, Sabine N S; Giambaşu, George; Dai, Qing; Yesselman, Joseph D; York, Darrin; Piccirilli, Joseph A; Herschlag, Daniel

2016-01-01

Biological catalysis hinges on the precise structural integrity of an active site that binds and transforms its substrates and meeting this requirement presents a unique challenge for RNA enzymes. Functional RNAs, including ribozymes, fold into their active conformations within rugged energy landscapes that often contain misfolded conformers. Here we uncover and characterize one such "off-pathway" species within an active site after overall folding of the ribozyme is complete. The Tetrahymena group I ribozyme (E) catalyzes cleavage of an oligonucleotide substrate (S) by an exogenous guanosine (G) cofactor. We tested whether specific catalytic interactions with G are present in the preceding E•S•G and E•G ground-state complexes. We monitored interactions with G via the effects of 2'- and 3'-deoxy (-H) and -amino (-NH(2)) substitutions on G binding. These and prior results reveal that G is bound in an inactive configuration within E•G, with the nucleophilic 3'-OH making a nonproductive interaction with an active site metal ion termed MA and with the adjacent 2'-OH making no interaction. Upon S binding, a rearrangement occurs that allows both -OH groups to contact a different active site metal ion, termed M(C), to make what are likely to be their catalytic interactions. The reactive phosphoryl group on S promotes this change, presumably by repositioning the metal ions with respect to G. This conformational transition demonstrates local rearrangements within an otherwise folded RNA, underscoring RNA's difficulty in specifying a unique conformation and highlighting Nature's potential to use local transitions of RNA in complex function. © 2015 Sengupta et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

XAFS studies of metal-ligand interactions at organic surfaces and in solution

NASA Astrophysics Data System (ADS)

Boyanov, Maxim I.

X-ray absorption fine structure spectroscopy (XAFS) was used as a structural probe to determine the mechanism of metal adsorption to organic surfaces. Two specific systems were investigated, Pb adsorption to heneicosanoic acid Langmuir monolayers (CH3(CH2)19COOH), and Cd adsorption to isolated cell walls of the Bacillus subtilis bacterium. Although the study of these systems is important for quite different reasons, the goal in both is metal binding site speciation and structural characterization of the surface complex. The adsorption of aqueous Cd to B. subtilis was studied as a function of pH by fluorescence mode bulk XAFS. Samples were prepared at six pH values in the range 3.4 to 7.8, and the bacterial functional groups responsible for the adsorption were identified under each condition. Under the experimental Cd and bacterial concentrations, the spectroscopy results indicate that Cd binds predominantly to protonated phosphoryl ligands below pH 4.4, while at higher pH adsorption to carboxyl groups becomes increasingly important. At pH 7.8 we observe the activation of an additional binding site, which we tentatively ascribe to deprotonated phosphoryl ligands. A quantitative Cd speciation diagram for the pH range is presented. Grazing-incidence Pb L3 edge XAFS was used in situ to determine the adsorption complex structure in the Pb-Langmuir monolayer study. The results indicate covalent binding of the Pb cations to the carboxyl headgroups, and the observed Pb-Pb coordination suggests that the metal is adsorbed as a hydrolysis polymer, rather than as individual Pb 2+ ions. The data suggest a bidentate binding mechanism and a one Pb atom to one carboxyl headgroup binding stoichiometry. We discuss how this adsorption model can explain the peculiarities observed with Pb in previous metal-Langmuir monolayer studies. A systematic study of the metal local environment in aqueous solutions was conducted and used in the above analyses. Perchlorate and acetate salt solutions of Cd, Pb, Mn, Cr, and Cu were characterized as standards of hydrated ions and metal-carboxyl complexes. The utility of XAFS in differentiating between the ionic, monodentate, bridging-bidentate, and bidentate metal-carboxyl complexes through C-C multiple scattering effects and XANES features is demonstrated.

Modulating supramolecular binding of carbon dioxide in a redox-active porous metal-organic framework

PubMed Central

Lu, Zhenzhong; Godfrey, Harry G. W.; da Silva, Ivan; Cheng, Yongqiang; Savage, Mathew; Tuna, Floriana; McInnes, Eric J. L.; Teat, Simon J.; Gagnon, Kevin J.; Frogley, Mark D.; Manuel, Pascal; Rudić, Svemir; Ramirez-Cuesta, Anibal J.; Easun, Timothy L.; Yang, Sihai; Schröder, Martin

2017-01-01

Hydrogen bonds dominate many chemical and biological processes, and chemical modification enables control and modulation of host–guest systems. Here we report a targeted modification of hydrogen bonding and its effect on guest binding in redox-active materials. MFM-300(VIII) {[VIII2(OH)2(L)], LH4=biphenyl-3,3′,5,5′-tetracarboxylic acid} can be oxidized to isostructural MFM-300(VIV), [VIV2O2(L)], in which deprotonation of the bridging hydroxyl groups occurs. MFM-300(VIII) shows the second highest CO2 uptake capacity in metal-organic framework materials at 298 K and 1 bar (6.0 mmol g−1) and involves hydrogen bonding between the OH group of the host and the O-donor of CO2, which binds in an end-on manner, =1.863(1) Å. In contrast, CO2-loaded MFM-300(VIV) shows CO2 bound side-on to the oxy group and sandwiched between two phenyl groups involving a unique ···c.g.phenyl interaction [3.069(2), 3.146(3) Å]. The macroscopic packing of CO2 in the pores is directly influenced by these primary binding sites. PMID:28194014

Role of Conserved Glycine in Zinc-dependent Medium Chain Dehydrogenase/Reductase Superfamily*

PubMed Central

Tiwari, Manish Kumar; Singh, Raushan Kumar; Singh, Ranjitha; Jeya, Marimuthu; Zhao, Huimin; Lee, Jung-Kul

2012-01-01

The medium-chain dehydrogenase/reductase (MDR) superfamily consists of a large group of enzymes with a broad range of activities. Members of this superfamily are currently the subject of intensive investigation, but many aspects, including the zinc dependence of MDR superfamily proteins, have not yet have been adequately investigated. Using a density functional theory-based screening strategy, we have identified a strictly conserved glycine residue (Gly) in the zinc-dependent MDR superfamily. To elucidate the role of this conserved Gly in MDR, we carried out a comprehensive structural, functional, and computational analysis of four MDR enzymes through a series of studies including site-directed mutagenesis, isothermal titration calorimetry, electron paramagnetic resonance (EPR), quantum mechanics, and molecular mechanics analysis. Gly substitution by other amino acids posed a significant threat to the metal binding affinity and activity of MDR superfamily enzymes. Mutagenesis at the conserved Gly resulted in alterations in the coordination of the catalytic zinc ion, with concomitant changes in metal-ligand bond length, bond angle, and the affinity (Kd) toward the zinc ion. The Gly mutants also showed different spectroscopic properties in EPR compared with those of the wild type, indicating that the binding geometries of the zinc to the zinc binding ligands were changed by the mutation. The present results demonstrate that the conserved Gly in the GHE motif plays a role in maintaining the metal binding affinity and the electronic state of the catalytic zinc ion during catalysis of the MDR superfamily enzymes. PMID:22500022

Pd n Ag (4-n) and Pd n Pt (4-n) clusters on MgO (100): a density functional surface genetic algorithm investigation

DOE PAGES

Heard, Christopher J.; Heiles, Sven; Vajda, Stefan; ...

2014-08-07

We employed the novel surface mode of the Birmingham Cluster Genetic Algorithm (S-BCGA) for the global optimisation of noble metal tetramers upon an MgO(100) substrate at the GGA-DFT level of theory. The effect of element identity and alloying in surface-bound neutral subnanometre clusters is determined by energetic comparison between all compositions of Pd nAg (4-n) and Pd nPt (4-n). And while the binding strengths to the surface increase in the order Pt > Pd > Ag, the excess energy profiles suggest a preference for mixed clusters for both cases. The binding of CO is also modelled, showing that the adsorptionmore » site can be predicted solely by electrophilicity. Comparison to CO binding on a single metal atom shows a reversal of the 5s-d activation process for clusters, weakening the cluster surface interaction on CO adsorption. Charge localisation determines homotop, CO binding and surface site preferences. Furthermore, the electronic behaviour, which is intermediate between molecular and metallic particles allows for tunable features in the subnanometre size range.« less

Metal centre effects on HNO binding in porphyrins and the electronic origin: metal's electronic configuration, position in the periodic table, and oxidation state.

PubMed

Yang, Liu; Fang, Weihai; Zhang, Yong

2012-04-21

HNO binds to many different metals in organometallic and bioinorganic chemistry. To help understand experimentally observed metal centre effects, a quantum chemical investigation was performed, revealing clear general binding trends with respect to metal centre characteristics and the electronic origin for the first time. This journal is © The Royal Society of Chemistry 2012

Decreased Sensitivity to Changes in the Concentration of Metal Ions as the Basis for the Hyperactivity of DtxR(E175K)

DOE Office of Scientific and Technical Information (OSTI.GOV)

D’Aquino, J. Alejandro; Denninger, Andrew R.; Moulin, Aaron G.

2010-01-12

The metal-ion-activated diphtheria toxin repressor (DtxR) is responsible for the regulation of virulence and other genes in Corynebacterium diphtheriae. A single point mutation in DtxR, DtxR(E175K), causes this mutant repressor to have a hyperactive phenotype. Mice infected with Mycobacterium tuberculosis transformed with plasmids carrying this mutant gene show reduced signs of the tuberculosis infection. Corynebacterial DtxR is able to complement mycobacterial IdeR and vice versa. To date, an explanation for the hyperactivity of DtxR(E175K) has remained elusive. In an attempt to address this issue, we have solved the first crystal structure of DtxR(E175K) and characterized this mutant using circular dichroism,more » isothermal titration calorimetry, and other biochemical techniques. The results show that although DtxR(E175K) and the wild type have similar secondary structures, DtxR(E175K) gains additional thermostability upon activation with metal ions, which may lead to this mutant requiring a lower concentration of metal ions to reach the same levels of thermostability as the wild-type protein. The E175K mutation causes binding site 1 to retain metal ion bound at all times, which can only be removed by incubation with an ion chelator. The crystal structure of DtxR(E175K) shows an empty binding site 2 without evidence of oxidation of Cys102. The association constant for this low-affinity binding site of DtxR(E175K) obtained from calorimetric titration with Ni(II) is K{sub a} = 7.6 {+-} 0.5 x 10{sup 4}, which is very similar to the reported value for the wild-type repressor, K{sub a} = 6.3 x 10{sup 4}. Both the wild type and DtxR(E175K) require the same amount of metal ion to produce a shift in the electrophoretic mobility shift assay, but unlike the wild type, DtxR(E175K) binding to its cognate DNA [tox promoter-operator (toxPO)] does not require metal-ion supplementation in the running buffer. In the timescale of these experiments, the Mn(II)-DtxR(E175K)-toxPO complex is insensitive to changes in the environmental cation concentrations. In addition to Mn(II), Ni(II), Co(II), Cd(II), and Zn(II) are able to sustain the hyperactive phenotype. These results demonstrate a prominent role of binding site 1 in the activation of DtxR and support the hypothesis that DtxR(E175K) attenuates the expression of virulence due to the decreased ability of the Me(II)-DtxR(E175K)-toxPO complex to dissociate at low concentrations of metal ions.« less

Effect of heavy metals ions on enzyme activity in the Mediterranean mussel, Donax trunculus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mizrahi, L.; Achituv, Y.

Heavy metal ions strongly are bound by sulfhydryl groups of proteins. Sulfhydryl binding changes the structure and enzymatic activities of proteins and causes toxic effects evident at the whole organism level. Heavy metal ions like Cd, Cu, Hg, Zn, and Pb in sufficiently high concentrations might kill organisms or cause other adverse effects that changing aquatic community structures. Bivalves are known to be heavy metal accumulators. The aim of the present study was to examine the effects of different concentrations of each of five heavy metal ions on the activity of four enzymes in D. trunculus. As it is knownmore » that heavy metals inhibit the activity of a wide range of enzymes, the authors chose representative examples of dehydrogenases (lactate and malate dehydrogenases), respiratory enzyme (cytochrome oxidase) and digestive enzyme ({alpha}-amylase). The acute effects of different concentrations of selected metals were examined. These concentrations were higher than those found usually in the locality where the animals occur, but might be encountered during a given event of pollution.« less

Divergent assembly mechanisms of the manganese/iron cofactors in R2lox and R2c proteins.

PubMed

Kutin, Yuri; Srinivas, Vivek; Fritz, Matthieu; Kositzki, Ramona; Shafaat, Hannah S; Birrell, James; Bill, Eckhard; Haumann, Michael; Lubitz, Wolfgang; Högbom, Martin; Griese, Julia J; Cox, Nicholas

2016-09-01

A manganese/iron cofactor which performs multi-electron oxidative chemistry is found in two classes of ferritin-like proteins, the small subunit (R2) of class Ic ribonucleotide reductase (R2c) and the R2-like ligand-binding oxidase (R2lox). It is unclear how a heterodimeric Mn/Fe metallocofactor is assembled in these two related proteins as opposed to a homodimeric Fe/Fe cofactor, especially considering the structural similarity and proximity of the two metal-binding sites in both protein scaffolds and the similar first coordination sphere ligand preferences of Mn II and Fe II . Using EPR and Mössbauer spectroscopies as well as X-ray anomalous dispersion, we examined metal loading and cofactor activation of both proteins in vitro (in solution). We find divergent cofactor assembly mechanisms for the two systems. In both cases, excess Mn II promotes heterobimetallic cofactor assembly. In the absence of Fe II , R2c cooperatively binds Mn II at both metal sites, whereas R2lox does not readily bind Mn II at either site. Heterometallic cofactor assembly is favored at substoichiometric Fe II concentrations in R2lox. Fe II and Mn II likely bind to the protein in a stepwise fashion, with Fe II binding to site 2 initiating cofactor assembly. In R2c, however, heterometallic assembly is presumably achieved by the displacement of Mn II by Fe II at site 2. The divergent metal loading mechanisms are correlated with the putative in vivo functions of R2c and R2lox, and most likely with the intracellular Mn II /Fe II concentrations in the host organisms from which they were isolated. Copyright © 2016 Elsevier Inc. All rights reserved.

Deconvolution of the role of metal and pH in metal coordinating polymers

NASA Astrophysics Data System (ADS)

Cazzell, Seth; Holten-Andersen, Niels

Nature uses metal binding amino acids to engineer both mechanical properties and structural functionality. Some examples of this metal binding behavior can be found in both mussel foot protein and DNA binding protein. The mussel byssal thread contains reversible intermolecular protein-metal bonds, allowing it to withstand harsh intertidal environments. Zinc fingers form intramolecular protein-metal bonds to stabilize the tertiary structure of DNA binding proteins, allowing specific structural functionality. Inspired by both these metal-binding materials, we present mechanical and spectroscopic characterization of a model polymer system, designed to mimic this bonding. Through these studies, we are able to answer fundamental polymer physics questions, such as the role of pH and metal to ligand ratio, illuminating both the macroscopic and microscopic material behavior. These understandings further bio-inspired engineering techniques that are used to design viscoelastic soft materials. I was supported by the Department of Defense (DoD) through the National Defense Science & Engineering Graduate Fellowship (NDSEG) Program.

A simple model for metal cation-phosphate interactions in nucleic acids in the gas phase: alkali metal cations and trimethyl phosphate.

PubMed

Ruan, Chunhai; Huang, Hai; Rodgers, M T

2008-02-01

Threshold collision-induced dissociation techniques are employed to determine the bond dissociation energies (BDEs) of complexes of alkali metal cations to trimethyl phosphate, TMP. Endothermic loss of the intact TMP ligand is the only dissociation pathway observed for all complexes. Theoretical calculations at the B3LYP/6-31G* level of theory are used to determine the structures, vibrational frequencies, and rotational constants of neutral TMP and the M+(TMP) complexes. Theoretical BDEs are determined from single point energy calculations at the B3LYP/6-311+G(2d,2p) level using the B3LYP/6-31G* optimized geometries. The agreement between theory and experiment is reasonably good for all complexes except Li+(TMP). The absolute M+-(TMP) BDEs are found to decrease monotonically as the size of the alkali metal cation increases. No activated dissociation was observed for alkali metal cation binding to TMP. The binding of alkali metal cations to TMP is compared with that to acetone and methanol.

Targeting Mycobacterium tuberculosis Topoisomerase I by Small-Molecule Inhibitors

PubMed Central

Godbole, Adwait Anand; Ahmed, Wareed; Bhat, Rajeshwari Subray; Bradley, Erin K.; Ekins, Sean

2014-01-01

We describe inhibition of Mycobacterium tuberculosis topoisomerase I (MttopoI), an essential mycobacterial enzyme, by two related compounds, imipramine and norclomipramine, of which imipramine is clinically used as an antidepressant. These molecules showed growth inhibition of both Mycobacterium smegmatis and M. tuberculosis cells. The mechanism of action of these two molecules was investigated by analyzing the individual steps of the topoisomerase I (topoI) reaction cycle. The compounds stimulated cleavage, thereby perturbing the cleavage-religation equilibrium. Consequently, these molecules inhibited the growth of the cells overexpressing topoI at a low MIC. Docking of the molecules on the MttopoI model suggested that they bind near the metal binding site of the enzyme. The DNA relaxation activity of the metal binding mutants harboring mutations in the DxDxE motif was differentially affected by the molecules, suggesting that the metal coordinating residues contribute to the interaction of the enzyme with the drug. Taken together, the results highlight the potential of these small molecules, which poison the M. tuberculosis and M. smegmatis topoisomerase I, as leads for the development of improved molecules to combat mycobacterial infections. Moreover, targeting metal coordination in topoisomerases might be a general strategy to develop new lead molecules. PMID:25534741

Mechanistic study of manganese-substituted glycerol dehydrogenase using a kinetic and thermodynamic analysis.

PubMed

Fang, Baishan; Niu, Jin; Ren, Hong; Guo, Yingxia; Wang, Shizhen

2014-01-01

Mechanistic insights regarding the activity enhancement of dehydrogenase by metal ion substitution were investigated by a simple method using a kinetic and thermodynamic analysis. By profiling the binding energy of both the substrate and product, the metal ion's role in catalysis enhancement was revealed. Glycerol dehydrogenase (GDH) from Klebsiella pneumoniae sp., which demonstrated an improvement in activity by the substitution of a zinc ion with a manganese ion, was used as a model for the mechanistic study of metal ion substitution. A kinetic model based on an ordered Bi-Bi mechanism was proposed considering the noncompetitive product inhibition of dihydroxyacetone (DHA) and the competitive product inhibition of NADH. By obtaining preliminary kinetic parameters of substrate and product inhibition, the number of estimated parameters was reduced from 10 to 4 for a nonlinear regression-based kinetic parameter estimation. The simulated values of time-concentration curves fit the experimental values well, with an average relative error of 11.5% and 12.7% for Mn-GDH and GDH, respectively. A comparison of the binding energy of enzyme ternary complex for Mn-GDH and GDH derived from kinetic parameters indicated that metal ion substitution accelerated the release of dioxyacetone. The metal ion's role in catalysis enhancement was explicated.

Role of Metal Ions on the Activity of Mycobacterium tuberculosis Pyrazinamidase

PubMed Central

Sheen, Patricia; Ferrer, Patricia; Gilman, Robert H.; Christiansen, Gina; Moreno-Román, Paola; Gutiérrez, Andrés H.; Sotelo, Jun; Evangelista, Wilfredo; Fuentes, Patricia; Rueda, Daniel; Flores, Myra; Olivera, Paula; Solis, José; Pesaresi, Alessandro; Lamba, Doriano; Zimic, Mirko

2012-01-01

Pyrazinamidase of Mycobacterium tuberculosis catalyzes the conversion of pyrazinamide to the active molecule pyrazinoic acid. Reduction of pyrazinamidase activity results in a level of pyrazinamide resistance. Previous studies have suggested that pyrazinamidase has a metal-binding site and that a divalent metal cofactor is required for activity. To determine the effect of divalent metals on the pyrazinamidase, the recombinant wild-type pyrazinamidase corresponding to the H37Rv pyrazinamide-susceptible reference strain was expressed in Escherichia coli with and without a carboxy terminal. His-tagged pyrazinamidase was inactivated by metal depletion and reactivated by titration with divalent metals. Although Co2+, Mn2+, and Zn2+ restored pyrazinamidase activity, only Co2+ enhanced the enzymatic activity to levels higher than the wild-type pyrazinamidase. Cu2+, Fe2+, Fe3+, and Mg2+ did not restore the activity under the conditions tested. Various recombinant mutated pyrazinamidases with appropriate folding but different enzymatic activities showed a differential pattern of recovered activity. X-ray fluorescence and atomic absorbance spectroscopy showed that recombinant wild-type pyrazinamidase expressed in E. coli most likely contained Zn. In conclusion, this study suggests that M. tuberculosis pyrazinamidase is a metalloenzyme that is able to coordinate several ions, but in vivo, it is more likely to coordinate Zn2+. However, in vitro, the metal-depleted enzyme could be reactivated by several divalent metals with higher efficiency than Zn. PMID:22764307

Synthesis, structures, nuclease activity, cytotoxicity, DFT and molecular docking studies of two nitrato bridged homodinuclear (Cu-Cu, Zn-Zn) complexes containing 2,2'-bipyridine and a chalcone derivative.

PubMed

Gaur, Ruchi; Choubey, Diksha Kumari; Usman, Mohammad; Ward, Benzamin D; Roy, Jagat Kumar; Mishra, Lallan

2017-08-01

Nitrato briged dinuclear complexes of type [Cu 2 (L) 2 (bpy) 2 (NO 3 )](NO 3 )·4H 2 O, 1 and [Zn 2 (L) 2 (bpy) 2 (NO 3 )](NO 3 )·4H 2 O, 2 (L=deprotonated form of free ligand LH, [1-(2-hydroxyphenyl)-3-(9-anthracenyl) propenone; bpy=2,2'bipyridine] are synthesized and characterized using a battery of physicochemical techniques and X-ray crystallography. A distorted square pyramidal geometry is assigned to them with N 2 O 3 coordination core around the metal ion. The co-ligand L binds the metal ions through its O,O' atoms in anti-syn mode. The metal centers in complexes 1 and 2 are separated via bridging nitrato group at a distance of 6.073Å and 5.635Å respectively. Their structures and absorption spectra are supported by the computational studies using density functional theory (DFT) and TD-DFT. Both complexes exhibit nuclease activity and cleave supercoiled (form I) DNA. The complex 1 preferentially binds major groove of DNA and follows an oxidative pathway whereas complex 2 binds with minor groove of DNA via hydrolytic pathway. Both complexes inhibit topoisomerase I relaxation activity with IC 50 values of 7 and 35μM. Molecular docking studies support the groove binding and topoisomerase I binding of the complexes. The complex 1 showed a significant cytotoxicity against HeLa cell lines (a cervical cancer cell lines) in vitro with IC 50 value calculated as 2.9±0.021μM as compared to 28.2±0. 044μΜ for complex 2. Complex 2 induces the cell apoptosis at a later-stage as compared to complex 1. The cell apoptosis and topoisomerase inhibition by complexes enable them to be potential candidates as future anticancer drugs. Copyright © 2017 Elsevier B.V. All rights reserved.

Structural characterization of tartrate dehydrogenase: a versatile enzyme catalyzing multiple reactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Malik, Radhika; Viola, Ronald E.

2010-10-28

The first structure of an NAD-dependent tartrate dehydrogenase (TDH) has been solved to 2 {angstrom} resolution by single anomalous diffraction (SAD) phasing as a complex with the intermediate analog oxalate, Mg{sup 2+} and NADH. This TDH structure from Pseudomonas putida has a similar overall fold and domain organization to other structurally characterized members of the hydroxy-acid dehydrogenase family. However, there are considerable differences between TDH and these functionally related enzymes in the regions connecting the core secondary structure and in the relative positioning of important loops and helices. The active site in these complexes is highly ordered, allowing the identificationmore » of the substrate-binding and cofactor-binding groups and the ligands to the metal ions. Residues from the adjacent subunit are involved in both the substrate and divalent metal ion binding sites, establishing a dimer as the functional unit and providing structural support for an alternating-site reaction mechanism. The divalent metal ion plays a prominent role in substrate binding and orientation, together with several active-site arginines. Functional groups from both subunits form the cofactor-binding site and the ammonium ion aids in the orientation of the nicotinamide ring of the cofactor. A lysyl amino group (Lys192) is the base responsible for the water-mediated proton abstraction from the C2 hydroxyl group of the substrate that begins the catalytic reaction, followed by hydride transfer to NAD. A tyrosyl hydroxyl group (Tyr141) functions as a general acid to protonate the enolate intermediate. Each substrate undergoes the initial hydride transfer, but differences in substrate orientation are proposed to account for the different reactions catalyzed by TDH.« less

Molecular dissection of a putative iron reductase from Desulfotomaculum reducens MI-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Zhi; Kim, David D.; Nelson, Ornella D.

2015-10-08

Desulfotomaculum reducens MI-1 is a Firmicute strain capable of reducing a variety of heavy metal ions and has a great potential in heavy metal bioremediation.We recently identified Dred_2421 as a potential iron reductase through proteomic study of D. reducens. The current study examines its iron-reduction mechanism. Dred_2421, like its close homolog from Escherichia coli (2, 4-dienoyl-CoA reductase), has an FMN-binding N-terminal domain (NTD), an FAD-binding C-terminal domain (CTD), and a 4Fee4S cluster between the two domains. To understand the mechanism of the iron-reduction activity and the role of each domain, we generated a series of variants for each domain andmore » investigated their iron reduction activity. Our results suggest that CTD is the main contributor of the iron-reduction activity, and that NTD and the 4Fee4S cluster are not directly involved in such activity. This study provides a mechanistic understanding of the ironereductase activity of Dred_2421 and may also help to elucidate other physiological activities this enzyme may have.« less

Revised Model of Calcium and Magnesium Binding to the Bacterial Cell Wall

PubMed Central

Thomas, Kieth J.; Rice, Charles V.

2014-01-01

Metals bind to the bacterial cell wall yet the binding mechanisms and affinity constants are not fully understood. The cell wall of gram positive bacteria is characterized by a thick layer of peptidoglycan and anionic teichoic acids anchored in the cytoplasmic membrane (lipoteichoic acid) or covalently bound to the cell wall (wall teichoic acid). The polyphosphate groups of teichoic acid provide one-half of the metal binding sites for calcium and magnesium, contradicting previous reports that calcium binding is 100% dependent on teichoic acid. The remaining binding sites are formed with the carboxyl units of peptidoglycan. In this work we report equilibrium association constants and total metal binding capacities for the interaction of calcium and magnesium ions with the bacterial cell wall. Metal binding is much stronger and previously reported. Curvature of Scatchard plots from the binding data and the resulting two regions of binding affinity suggest the presence of negative cooperative binding, meaning that the binding affinity decreases as more ions become bound to the sample. For Ca2+, Region I has a KA = (1.0 ± 0.2) × 106 M−1 and Region II has a KA = (0.075 ± 0.058) × 106 M−1. For Mg2+, KA1 = (1.5 ± 0.1) × 106 and KA2 = (0.17 ± 0.10) × 106. A binding capacity (η) is reported for both regions. However, since binding is still occurring in Region II, the total binding capacity is denoted by η2, which are 0.70 ± 0.04 µmol/mg and 0.67 ± 0.03 µmol/mg for Ca2+ and Mg2+ respectively. These data contradict the current paradigm of there being a single metal affinity value that is constant over a range of concentrations. We also find that measurement of equilibrium binding constants is highly sample dependent, suggesting a role for diffusion of metals through heterogeneous cell wall fragments. As a result, we are able to reconcile many contradictory theories that describe binding affinity and the binding mode of divalent metal cations. PMID:25315444

Introducing a 2-His-1-Glu Nonheme Iron Center into Myoglobin Confers Nitric Oxide Reductase Activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Y Lin; N Yeung; Y Gao

2011-12-31

A conserved 2-His-1-Glu metal center, as found in natural nonheme iron-containing enzymes, was engineered into sperm whale myoglobin by replacing Leu29 and Phe43 with Glu and His, respectively (swMb L29E, F43H, H64, called Fe{sub B}Mb(-His)). A high resolution (1.65 {angstrom}) crystal structure of Cu(II)-CN{sup -}-Fe{sub B}Mb(-His) was determined, demonstrating that the unique 2-His-1-Glu metal center was successfully created within swMb. The Fe{sub B}Mb(-His) can bind Cu, Fe, or Zn ions, with both Cu(I)-Fe{sub B}Mb(-His) and Fe(II)-Fe{sub B}Mb(-His) exhibiting nitric oxide reductase (NOR) activities. Cu dependent NOR activity was significantly higher than that of Fe in the same metal binding site.more » EPR studies showed that the reduction of NO to N{sub 2}O catalyzed by these two enzymes resulted in different intermediates; a five-coordinate heme-NO species was observed for Cu(I)-Fe{sub B}Mb(-His) due to the cleavage of the proximal heme Fe-His bond, while Fe(II)-Fe{sub B}Mb(-His) remained six-coordinate. Therefore, both the metal ligand, Glu29, and the metal itself, Cu or Fe, play crucial roles in NOR activity. This study presents a novel protein model of NOR and provides insights into a newly discovered member of the NOR family, gNOR.« less

Kinetics of heavy metal adsorption and desorption in soil: Developing a unified model based on chemical speciation

NASA Astrophysics Data System (ADS)

Peng, Lanfang; Liu, Paiyu; Feng, Xionghan; Wang, Zimeng; Cheng, Tao; Liang, Yuzhen; Lin, Zhang; Shi, Zhenqing

2018-03-01

Predicting the kinetics of heavy metal adsorption and desorption in soil requires consideration of multiple heterogeneous soil binding sites and variations of reaction chemistry conditions. Although chemical speciation models have been developed for predicting the equilibrium of metal adsorption on soil organic matter (SOM) and important mineral phases (e.g. Fe and Al (hydr)oxides), there is still a lack of modeling tools for predicting the kinetics of metal adsorption and desorption reactions in soil. In this study, we developed a unified model for the kinetics of heavy metal adsorption and desorption in soil based on the equilibrium models WHAM 7 and CD-MUSIC, which specifically consider metal kinetic reactions with multiple binding sites of SOM and soil minerals simultaneously. For each specific binding site, metal adsorption and desorption rate coefficients were constrained by the local equilibrium partition coefficients predicted by WHAM 7 or CD-MUSIC, and, for each metal, the desorption rate coefficients of various binding sites were constrained by their metal binding constants with those sites. The model had only one fitting parameter for each soil binding phase, and all other parameters were derived from WHAM 7 and CD-MUSIC. A stirred-flow method was used to study the kinetics of Cd, Cu, Ni, Pb, and Zn adsorption and desorption in multiple soils under various pH and metal concentrations, and the model successfully reproduced most of the kinetic data. We quantitatively elucidated the significance of different soil components and important soil binding sites during the adsorption and desorption kinetic processes. Our model has provided a theoretical framework to predict metal adsorption and desorption kinetics, which can be further used to predict the dynamic behavior of heavy metals in soil under various natural conditions by coupling other important soil processes.

The Role of zinc in the disulphide stress-regulated anti-sigma factor RsrA from Streptomyces coelicolor.

PubMed

Li, Wei; Bottrill, Andrew R; Bibb, Maureen J; Buttner, Mark J; Paget, Mark S B; Kleanthous, Colin

2003-10-17

The regulation of disulphide stress in actinomycetes such as Streptomyces coelicolor is known to involve the zinc-containing anti-sigma factor RsrA that binds and inactivates the redox-regulated sigma factor sigmaR. However, it is not known how RsrA senses disulphide stress nor what role the metal ion plays. Using in vitro assays, we show that while zinc is not required for sigmaR binding it is required for functional anti-sigma factor activity, and that it plays a critical role in modulating the reactivity of RsrA cysteine thiol groups towards oxidation. Apo-RsrA is easily oxidised and, while the Zn-bound form is relatively resistant, the metal ion is readily expelled when the protein is treated with strong oxidants such as diamide. We also show, using a combination of proteolysis and mass spectrometry, that the first critical disulphide to form in RsrA involves Cys11 and one of either Cys41 or Cys44, all previously implicated in metal binding. Circular dichroism spectroscopy was used to follow structural changes during oxidation of RsrA, which indicated that concomitant with formation of this critical disulphide bond is a major restructuring of the protein where its alpha-helical content increases. Our data demonstrate that RsrA can only bind sigmaR in the reduced state and that this state is stabilised by zinc. Redox stress induces disulphide bond formation amongst zinc-ligating residues, expelling the metal ion and stabilising a structure incapable of binding the sigma factor.

How water molecules affect the catalytic activity of hydrolases--a XANES study of the local structures of peptide deformylase.

PubMed

Cui, Peixin; Wang, Yu; Chu, Wangsheng; Guo, Xiaoyun; Yang, Feifei; Yu, Meijuan; Zhao, Haifeng; Dong, Yuhui; Xie, Yaning; Gong, Weimin; Wu, Ziyu

2014-12-12

Peptide deformylase (PDF) is a prokaryotic enzyme that catalyzes the deformylation of nascent peptides generated during protein synthesis and water molecules play a key role in these hydrolases. Using X-ray absorption near edge spectroscopy (XANES) and ab initio calculations we accurately probe the local atomic environment of the metal ion binding in the active site of PDF at different pH values and with different metal ions. This new approach is an effective way to monitor existing correlations among functions and structural changes. We show for the first time that the enzymatic activity depends on pH values and metal ions via the bond length of the nearest coordinating water (Wat1) to the metal ion. Combining experimental and theoretical data we may claim that PDF exhibits an enhanced enzymatic activity only when the distance of the Wat1 molecule with the metal ion falls in the limited range from 2.15 to 2.55 Å.

Metal-Mediated Modulation of Streptococcal Cysteine Protease Activity and Its Biological Implications

PubMed Central

Chella Krishnan, Karthickeyan; Mukundan, Santhosh; Landero Figueroa, Julio A.; Caruso, Joseph A.

2014-01-01

Streptococcal cysteine protease (SpeB), the major secreted protease produced by group A streptococcus (GAS), cleaves both host and bacterial proteins and contributes importantly to the pathogenesis of invasive GAS infections. Modulation of SpeB expression and/or its activity during invasive GAS infections has been shown to affect bacterial virulence and infection severity. Expression of SpeB is regulated by the GAS CovR-CovS two-component regulatory system, and we demonstrated that bacteria with mutations in the CovR-CovS two-component regulatory system are selected for during localized GAS infections and that these bacteria lack SpeB expression and exhibit a hypervirulent phenotype. Additionally, in a separate study, we showed that expression of SpeB can also be modulated by human transferrin- and/or lactoferrin-mediated iron chelation. Accordingly, the goal of this study was to investigate the possible roles of iron and other metals in modulating SpeB expression and/or activity in a manner that would potentiate bacterial virulence. Here, we report that the divalent metals zinc and copper inhibit SpeB activity at the posttranslational level. Utilizing online metal-binding site prediction servers, we identified two putative metal-binding sites in SpeB, one of which involves the catalytic-dyad residues 47Cys and 195His. Based on our findings, we propose that zinc and/or copper availability in the bacterial microenvironment can modulate the proteolytic activity of SpeB in a manner that preserves the integrity of several other virulence factors essential for bacterial survival and dissemination within the host and thereby may exacerbate the severity of invasive GAS infections. PMID:24799625

Metallation and mismetallation of iron and manganese proteins in vitro and in vivo: the class I ribonucleotide reductases as a case study.

PubMed

Cotruvo, Joseph A; Stubbe, Joanne

2012-10-01

How cells ensure correct metallation of a given protein and whether a degree of promiscuity in metal binding has evolved are largely unanswered questions. In a classic case, iron- and manganese-dependent superoxide dismutases (SODs) catalyze the disproportionation of superoxide using highly similar protein scaffolds and nearly identical active sites. However, most of these enzymes are active with only one metal, although both metals can bind in vitro and in vivo. Iron(ii) and manganese(ii) bind weakly to most proteins and possess similar coordination preferences. Their distinct redox properties suggest that they are unlikely to be interchangeable in biological systems except when they function in Lewis acid catalytic roles, yet recent work suggests this is not always the case. This review summarizes the diversity of ways in which iron and manganese are substituted in similar or identical protein frameworks. As models, we discuss (1) enzymes, such as epimerases, thought to use Fe(II) as a Lewis acid under normal growth conditions but which switch to Mn(II) under oxidative stress; (2) extradiol dioxygenases, which have been found to use both Fe(II) and Mn(II), the redox role of which in catalysis remains to be elucidated; (3) SODs, which use redox chemistry and are generally metal-specific; and (4) the class I ribonucleotide reductases (RNRs), which have evolved unique biosynthetic pathways to control metallation. The primary focus is the class Ib RNRs, which can catalyze formation of a stable radical on a tyrosine residue in their β2 subunits using either a di-iron or a recently characterized dimanganese cofactor. The physiological roles of enzymes that can switch between iron and manganese cofactors are discussed, as are insights obtained from the studies of many groups regarding iron and manganese homeostasis and the divergent and convergent strategies organisms use for control of protein metallation. We propose that, in many of the systems discussed, "discrimination" between metals is not performed by the protein itself, but it is instead determined by the environment in which the protein is expressed.

Parkinson Disease Protein DJ-1 Binds Metals and Protects against Metal-induced Cytotoxicity*

PubMed Central

Björkblom, Benny; Adilbayeva, Altynai; Maple-Grødem, Jodi; Piston, Dominik; Ökvist, Mats; Xu, Xiang Ming; Brede, Cato; Larsen, Jan Petter; Møller, Simon Geir

2013-01-01

The progressive loss of motor control due to reduction of dopamine-producing neurons in the substantia nigra pars compacta and decreased striatal dopamine levels are the classically described features of Parkinson disease (PD). Neuronal damage also progresses to other regions of the brain, and additional non-motor dysfunctions are common. Accumulation of environmental toxins, such as pesticides and metals, are suggested risk factors for the development of typical late onset PD, although genetic factors seem to be substantial in early onset cases. Mutations of DJ-1 are known to cause a form of recessive early onset Parkinson disease, highlighting an important functional role for DJ-1 in early disease prevention. This study identifies human DJ-1 as a metal-binding protein able to evidently bind copper as well as toxic mercury ions in vitro. The study further characterizes the cytoprotective function of DJ-1 and PD-mutated variants of DJ-1 with respect to induced metal cytotoxicity. The results show that expression of DJ-1 enhances the cells' protective mechanisms against induced metal toxicity and that this protection is lost for DJ-1 PD mutations A104T and D149A. The study also shows that oxidation site-mutated DJ-1 C106A retains its ability to protect cells. We also show that concomitant addition of dopamine exposure sensitizes cells to metal-induced cytotoxicity. We also confirm that redox-active dopamine adducts enhance metal-catalyzed oxidation of intracellular proteins in vivo by use of live cell imaging of redox-sensitive S3roGFP. The study indicates that even a small genetic alteration can sensitize cells to metal-induced cell death, a finding that may revive the interest in exogenous factors in the etiology of PD. PMID:23792957

Metal Toxicity at the Synapse: Presynaptic, Postsynaptic, and Long-Term Effects

PubMed Central

Sadiq, Sanah; Ghazala, Zena; Chowdhury, Arnab; Büsselberg, Dietrich

2012-01-01

Metal neurotoxicity is a global health concern. This paper summarizes the evidence for metal interactions with synaptic transmission and synaptic plasticity. Presynaptically metal ions modulate neurotransmitter release through their interaction with synaptic vesicles, ion channels, and the metabolism of neurotransmitters (NT). Many metals (e.g., Pb 2+, Cd 2+, and Hg +) also interact with intracellular signaling pathways. Postsynaptically, processes associated with the binding of NT to their receptors, activation of channels, and degradation of NT are altered by metals. Zn 2+, Pb 2+, Cu 2+, Cd 2+, Ni 2+, Co 2+, Li 3+, Hg +, and methylmercury modulate NMDA, AMPA/kainate, and/or GABA receptors activity. Al 3+, Pb 2+, Cd 2+, and As 2 O 3 also impair synaptic plasticity by targeting molecules such as CaM, PKC, and NOS as well as the transcription machinery involved in the maintenance of synaptic plasticity. The multiple effects of metals might occur simultaneously and are based on the specific metal species, metal concentrations, and the types of neurons involved. PMID:22287959

Spectroscopic and Thermodynamic Characterization of the Metal-Binding Sites in the LH1-RC Complex from Thermophilic Photosynthetic Bacterium Thermochromatium tepidum.

PubMed

Kimura, Yukihiro; Yura, Yuki; Hayashi, Yusuke; Li, Yong; Onoda, Moe; Yu, Long-Jiang; Wang-Otomo, Zheng-Yu; Ohno, Takashi

2016-12-15

The light-harvesting 1 reaction center (LH1-RC) complex from thermophilic photosynthetic bacterium Thermochromatium (Tch.) tepidum exhibits enhanced thermostability and an unusual LH1 Q y transition, both induced by Ca 2+ binding. In this study, metal-binding sites and metal-protein interactions in the LH1-RC complexes from wild-type (B915) and biosynthetically Sr 2+ -substituted (B888) Tch. tepidum were investigated by isothermal titration calorimetry (ITC), atomic absorption (AA), and attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopies. The ITC measurements revealed stoichiometric ratios of approximately 1:1 for binding of Ca 2+ , Sr 2+ , or Ba 2+ to the LH1 αβ-subunit, indicating the presence of 16 binding sites in both B915 and B888. The AA analysis provided direct evidence for Ca 2+ and Sr 2+ binding to B915 and B888, respectively, in their purified states. Metal-binding experiments supported that Ca 2+ and Sr 2+ (or Ba 2+ ) competitively associate with the binding sites in both species. The ATR-FTIR difference spectra upon Ca 2+ depletion and Sr 2+ substitution demonstrated that dissociation and binding of Ca 2+ are predominantly responsible for metal-dependent conformational changes of B915 and B888. The present results are largely compatible with the recent structural evidence that another binding site for Sr 2+ (or Ba 2+ ) exists in the vicinity of the Ca 2+ -binding site, a part of which is shared in both metal-binding sites.

Structure-guided engineering of Anticalins with improved binding behavior and biochemical characteristics for application in radio-immuno imaging and/or therapy.

PubMed

Eggenstein, E; Eichinger, A; Kim, H-J; Skerra, A

2014-02-01

Modern strategies in radio-immuno therapy and in vivo imaging require robust, small, and specific ligand-binding proteins. In this context we have previously developed artificial lipocalins, so-called Anticalins, with high binding activity toward rare-earth metal-chelate complexes using combinatorial protein design. Here we describe further improvement of the Anticalin C26 via in vitro affinity maturation to yield CL31, which has a fourfold slower dissociation half-life above 2h. Also, we present the crystallographic analyses of both the initial and the improved Anticalin, providing insight into the molecular mechanism of chelated metal binding and the role of amino acid substitutions during the step-wise affinity maturation. Notably, one of the four structurally variable loops that form the ligand pocket in the lipocalin scaffold undergoes a significant conformational change from C26 to CL31, acting as a lid that closes over the accommodated metal-chelate ligand. A systematic mutational study indicated that further improvement of ligand affinity is difficult to achieve while providing clues on the contribution of relevant side chains in the engineered binding pocket. Unexpectedly, some of the amino acid replacements led to strong increases - more then 10-fold - in the yield of soluble protein from periplasmic secretion in Escherichia coli. Copyright © 2013 Elsevier Inc. All rights reserved.

Anthrax Toxin Receptor 1 / Tumor Endothelial Marker 8: Mutation of Conserved Inserted Domain Residues Overrides Cytosolic Control of Protective Antigen Binding†

PubMed Central

Ramey, Jordan D.; Villareal, Valerie A.; Ng, Charles; Ward, Sabrina; Xiong, Jian-Ping; Clubb, Robert T.; Bradley, Kenneth A.

2010-01-01

Anthrax toxin receptor 1 (ANTXR1) / tumor endothelial marker 8 (TEM8) is one of two known proteinaceous cell surface anthrax toxin receptors. A metal ion dependent adhesion site (MIDAS) present in the integrin-like inserted (I) domain of ANTXR1 mediates the binding of the anthrax toxin subunit, protective antigen (PA). Here we provide evidence that single point mutations in the I domain can override regulation of ANTXR1 ligand-binding activity mediated by intracellular signals. A previously reported MIDAS-mutant of ANTXR1 (T118A) was found to retain normal metal ion binding and secondary structure but failed to bind PA, consistent with a locked inactive state. Conversely, mutation of a conserved I domain phenylalanine residue to a tryptophan (F205W) increased the proportion of cell-surface ANTXR1 that bound PA, consistent with a locked active state. Interestingly, the KD and total amount of PA bound by the isolated ANTXR1 I domain was not affected by the F205W mutation, indicating that ANTXR1 is preferentially found in the active state in the absence of inside-out signaling. Circular dichroism (CD) spectroscopy and 1H-15N heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) revealed that structural changes between T118A, F205W and WT I domains were minor despite a greater than 103-fold difference in their abilities to bind toxin. Regulation of toxin binding has important implications for the design of toxin inhibitors and for the targeting of ANTXR1 for anti-tumor therapies. PMID:20690680

Synthesis, spectroscopic, biological activity and thermal characterization of ceftazidime with transition metals

NASA Astrophysics Data System (ADS)

Masoud, Mamdouh S.; Ali, Alaa E.; Elasala, Gehan S.; Kolkaila, Sherif A.

2018-03-01

Synthesis, physicochemical characterization and thermal analysis of ceftazidime complexes with transition metals (Cr(III), Mn(II), Fe(III), Co(II), Ni(II), Cu(II), Zn(II), Cd(II) and Hg(II)) were discussed. It's obtained that ceftazidime act as bidentate ligand. From magnetic measurement and spectral data, octahedral structures were proposed for all complexes except for cobalt, nickel and mercury had tetrahedral structural. Hyper chemistry program confirmed binding sites of ceftazidime. Ceftazidime complexes show higher activity than ceftazidime for some strains. From TG and DTA curves the thermal decomposition mechanisms of ceftazidime and their metal complexes were suggested. The thermal decomposition of the complexes ended with the formation of metal oxides as a final product except in case of Hg complex.

Urban Biomining: Biological Extraction of Metals and Materials from Electronics Waste Using a Synthetic Biology Approach

NASA Astrophysics Data System (ADS)

Urbina-Navarrete, J.; Rothschild, L.

2016-12-01

End-of-life electronics waste (e-waste) containing toxic and valuable materials is a rapidly progressing human health and environmental issue. Using synthetic biology tools, we have developed a recycling method for e-waste. Our innovation is to use a recombinant version of a naturally-occurring silica-degrading enzyme to depolymerize the silica in metal- and glass- containing e-waste components, and subsequently, to use engineered bacterial surfaces to bind and separate metals from a solution. The bacteria with bound metals can then be used as "bio-ink" to print new circuits using a novel plasma jet electronics printing technology. Here, we present the results from our initial studies that focus on the specificity of metal-binding motifs for a cognate metal. The candidate motifs that show high affinity and specificity will be engineered into bacterial surfaces for downstream applications in biologically-mediated metal recycling. Since the chemistry and role of Cu in metalloproteins is relatively well-characterized, we are using Cu as a proxy to elucidate metal and biological ligand interactions with various metals in e-waste. We assess the binding parameters of 3 representative classes of Cu-binding motifs using isothermal titration calorimetry; 1) natural motifs found in metalloproteins, 2) consensus motifs, and 3) rationally designed peptides that are predicted, in silico, to bind Cu. Our results indicate that naturally-occurring motifs have relative high affinity and specificity for Cu (association constant for Cu Ka 104 M-1, Zn Ka 103 M-1) when competing ions are present in the aqueous milieu. However, motifs developed through rational design by applying quantum mechanical methods that take into account complexation energies of the elemental binding partners and molecular geometry of the cognate metal, not only show high affinity for the cognate metal (Cu Ka 106 M-1), but they show specificity and discrimination against other metal ions that would be competitors for the same binding sites. This is an initial proof-of-concept study that focuses on Cu-binding; however the overall objective of this research is to have peptides that selectively bind many metals from e-waste and this would allow for the separation of the metals from a solution, at ambient temperatures and under non-toxic conditions.

The mystery of gold's chemical activity: local bonding, morphology and reactivity of atomic oxygen.

PubMed

Baker, Thomas A; Liu, Xiaoying; Friend, Cynthia M

2011-01-07

Recently, gold has been intensely studied as a catalyst for key synthetic reactions. Gold is an attractive catalyst because, surprisingly, it is highly active and very selective for partial oxidation processes suggesting promise for energy-efficient "green" chemistry. The underlying origin of the high activity of Au is a controversial subject since metallic gold is commonly thought to be inert. Herein, we establish that one origin of the high activity for gold catalysis is the extremely reactive nature of atomic oxygen bound in 3-fold coordination sites on metallic gold. This is the predominant form of O at low concentrations on the surface, which is a strong indication that it is most relevant to catalytic conditions. Atomic oxygen bound to metallic Au in 3-fold sites has high activity for CO oxidation, oxidation of olefins, and oxidative transformations of alcohols and amines. Among the factors identified as important in Au-O interaction are the morphology of the surface, the local binding site of oxygen, and the degree of order of the oxygen overlayer. In this Perspective, we present an overview of both theory and experiments that identify the reactive forms of O and their associated charge density distributions and bond strengths. We also analyze and model the release of Au atoms induced by O binding to the surface. This rough surface also has the potential for O(2) dissociation, which is a critical step if Au is to be activated catalytically. We further show the strong parallels between product distributions and reactivity for O-covered Au at low pressure (ultrahigh vacuum) and for nanoporous Au catalysts operating at atmospheric pressure as evidence that atomic O is the active species under working catalytic conditions when metallic Au is present. We briefly discuss the possible contributions of oxidants that may contain intact O-O bonds and of the Au-metal oxide support interface in Au catalysis. Finally, the challenges and future directions for fully understanding the activity of gold are considered.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Leenheer, J.A.; Brown, G.K.; Cabaniss, S.E.

Fulvic acid, isolated from the Suwannee River, Georgia, was assessed for its ability to bind Ca{sup 2+}, Cd{sup 2+}, Cu{sup 2+}, Ni{sup 2+}, and Zn{sup 2+} ions at pH 6 before and after extensive fractionation that was designed to reveal the nature of metal binding functional groups. The binding constant for Ca{sup 2+} ion had the greatest increase of all the ions in a metal binding fraction that was selected for intensive characterization for the purpose of building quantitative average model structures. The metal binding fraction was characterized by quantitative {sup 13}C NMR, {sup 1}H NMR, and FT-IR spectrometry andmore » elemental, titrimetric, and molecular weight determinations. The characterization data revealed that carboxyl groups were clustered in short-chain aliphatic dibasic acid structures. The Ca{sup 2+} binding data suggested that ether-substituted oxysuccinic acid structures are good models for the metal binding sites at pH 6. Structural models were derived based upon oxidation and photolytic rearrangements of cutin, lignin, and tannin precursors. These structural models rich in substituted dibasic acid structures revealed polydentate binding sites with the potential for both inner-sphere and outer-sphere type binding. The majority of the fulvic acid molecule was involved with metal binding rather than a small substructural unit.« less

Sulfur and selenium antioxidants: challenging radical scavenging mechanisms and developing structure-activity relationships based on metal binding.

PubMed

Zimmerman, Matthew T; Bayse, Craig A; Ramoutar, Ria R; Brumaghim, Julia L

2015-04-01

Because sulfur and selenium antioxidants can prevent oxidative damage, numerous animal and clinical trials have investigated the ability of these compounds to prevent the oxidative stress that is an underlying cause of cardiovascular disease, Alzheimer's disease, and cancer, among others. One of the most common sources of oxidative damage is metal-generated hydroxyl radical; however, very little research has focused on determining the metal-binding abilities and structural attributes that affect oxidative damage prevention by sulfur and selenium compounds. In this review, we describe our ongoing investigations into sulfur and selenium antioxidant prevention of iron- and copper-mediated oxidative DNA damage. We determined that many sulfur and selenium compounds inhibit Cu(I)-mediated DNA damage and that DNA damage prevention varies dramatically when Fe(II) is used in place of Cu(I) to generate hydroxyl radical. Oxidation potentials of the sulfur or selenium compounds do not correlate with their ability to prevent DNA damage, highlighting the importance of metal coordination rather than reactive oxygen species scavenging as an antioxidant mechanism. Additional gel electrophoresis, mass spectrometry, and UV-visible studies confirmed sulfur and selenium antioxidant binding to Cu(I) and Fe(II). Ultimately, our studies established that both the hydroxyl-radical-generating metal ion and the chemical environment of the sulfur or selenium significantly affect DNA damage prevention and that metal coordination is an essential mechanism for these antioxidants. Copyright © 2015 Elsevier Inc. All rights reserved.

Impact of cadmium, cobalt and nickel on sequence-specific DNA binding of p63 and p73 in vitro and in cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adámik, Matej; Bažantová, Pavla; Department of Biology and Ecology, Faculty of Science, University of Ostrava, Chittussiho 10, 701 03 Ostrava

Highlights: • DNA binding of p53 family core domains is inhibited by cadmium, cobalt and nickel. • Binding to DNA protects p53 family core domains from metal induced inhibition. • Cadmium, cobalt and nickel induced inhibition was reverted by EDTA in vitro. - Abstract: Site-specific DNA recognition and binding activity belong to common attributes of all three members of tumor suppressor p53 family proteins: p53, p63 and p73. It was previously shown that heavy metals can affect p53 conformation, sequence-specific binding and suppress p53 response to DNA damage. Here we report for the first time that cadmium, nickel and cobalt,more » which have already been shown to disturb various DNA repair mechanisms, can also influence p63 and p73 sequence-specific DNA binding activity and transactivation of p53 family target genes. Based on results of electrophoretic mobility shift assay and luciferase reporter assay, we conclude that cadmium inhibits sequence-specific binding of all three core domains to p53 consensus sequences and abolishes transactivation of several promoters (e.g. BAX and MDM2) by 50 μM concentrations. In the presence of specific DNA, all p53 family core domains were partially protected against loss of DNA binding activity due to cadmium treatment. Effective cadmium concentration to abolish DNA–protein interactions was about two times higher for p63 and p73 proteins than for p53. Furthermore, we detected partial reversibility of cadmium inhibition for all p53 family members by EDTA. DTT was able to reverse cadmium inhibition only for p53 and p73. Nickel and cobalt abolished DNA–p53 interaction at sub-millimolar concentrations while inhibition of p63 and p73 DNA binding was observed at millimolar concentrations. In summary, cadmium strongly inhibits p53, p63 and p73 DNA binding in vitro and in cells in comparison to nickel and cobalt. The role of cadmium inhibition of p53 tumor suppressor family in carcinogenesis is discussed.« less

Two conformations of the integrin A-domain (I-domain): a pathway for activation?

PubMed

Lee, J O; Bankston, L A; Arnaout, M A; Liddington, R C

1995-12-15

Integrins are plasma membrane proteins that mediate adhesion to other cells and to components of the extracellular matrix. Most integrins are constitutively inactive in resting cells, but are rapidly and reversibly activated in response to agonists, leading to highly regulated cell adhesion. This activation is associated with conformational changes in their extracellular portions, but the nature of the structural changes that lead to a change in adhesiveness is not understood. The interactions of several integrins with their extracellular ligands are mediated by an A-type domain (generally called the I-domain in integrins). Binding of the I-domain to protein ligands is dependent on divalent cations. We have described previously the structure of the I-domain from complement receptor 3 with bound Mg2+, in which the glutamate side chain from a second I-domain completes the octahedral coordination sphere of the metal, acting as a ligand mimetic. We now describe a new crystal form of the I-domain with bound Mn2+, in which water completes the metal coordination sphere and there is no equivalent of the glutamate ligand. Comparison of the two crystal forms reveals a change in metal coordination which is linked to a large (10 A) shift of the C-terminal helix and the burial of two phenylalanine residues into the hydrophobic core of the Mn2+ form. These structural changes, analogous to those seen in the signal-transducing G-proteins, alter the electrophilicity of the metal, reducing its ability to bind ligand-associated acidic residues, and dramatically alter the surface of the protein implicated in binding ligand. Our observations provide the first atomic resolution view of conformational changes in an integrin domain, and suggest how these changes are linked to a change in integrin adhesiveness. We propose that the Mg2+ form represents the conformation of the domain in the active state and the Mn2+ form the conformation in the inactive state of the integrin.

Role of metal ions in catalysis by HIV integrase analyzed using a quantitative PCR disintegration assay.

PubMed

Diamond, Tracy L; Bushman, Frederic D

2006-01-01

Paired metal ions have been proposed to be central to the catalytic mechanisms of RNase H nucleases, bacterial transposases, Holliday junction resolvases, retroviral integrases and many other enzymes. Here we present a sensitive assay for DNA transesterification in which catalysis by human immunodeficiency virus-type 1 (HIV-1) integrase (IN) connects two DNA strands (disintegration reaction), allowing detection using quantitative PCR (qPCR). We present evidence suggesting that the three acidic residues of the IN active site function through metal binding using metal rescue. In this method, the catalytic acidic residues were each substituted with cysteines. Mn2+ binds tightly to the sulfur atoms of the cysteine residues, but Mg2+ does not. We found that Mn2+, but not Mg2+, could rescue catalysis of each cysteine-substituted enzyme, providing evidence for functionally important metal binding by all three residues. We also used the PCR-boosted assay to show that HIV-1 IN could carry out transesterification reactions involving DNA 5' hydroxyl groups as well as 3' hydroxyls as nucleophiles. Lastly, we show that Mn2+ by itself (i.e. without enzyme) can catalyze formation of a low level of PCR-amplifiable product under extreme conditions, allowing us to estimate the rate enhancement due to the IN-protein scaffold as at least 60 million-fold.

Role of metal ions in catalysis by HIV integrase analyzed using a quantitative PCR disintegration assay

PubMed Central

Diamond, Tracy L.; Bushman, Frederic D.

2006-01-01

Paired metal ions have been proposed to be central to the catalytic mechanisms of RNase H nucleases, bacterial transposases, Holliday junction resolvases, retroviral integrases and many other enzymes. Here we present a sensitive assay for DNA transesterification in which catalysis by human immunodeficiency virus-type 1 (HIV-1) integrase (IN) connects two DNA strands (disintegration reaction), allowing detection using quantitative PCR (qPCR). We present evidence suggesting that the three acidic residues of the IN active site function through metal binding using metal rescue. In this method, the catalytic acidic residues were each substituted with cysteines. Mn2+ binds tightly to the sulfur atoms of the cysteine residues, but Mg2+ does not. We found that Mn2+, but not Mg2+, could rescue catalysis of each cysteine-substituted enzyme, providing evidence for functionally important metal binding by all three residues. We also used the PCR-boosted assay to show that HIV-1 IN could carry out transesterification reactions involving DNA 5′ hydroxyl groups as well as 3′ hydroxyls as nucleophiles. Lastly, we show that Mn2+ by itself (i.e. without enzyme) can catalyze formation of a low level of PCR-amplifiable product under extreme conditions, allowing us to estimate the rate enhancement due to the IN-protein scaffold as at least 60 million-fold. PMID:17085478

The amyloid architecture provides a scaffold for enzyme-like catalysts.

PubMed

Al-Garawi, Z S; McIntosh, B A; Neill-Hall, D; Hatimy, A A; Sweet, S M; Bagley, M C; Serpell, L C

2017-08-03

Natural biological enzymes possess catalytic sites that are generally surrounded by a large three-dimensional scaffold. However, the proportion of the protein molecule that participates in the catalytic reaction is relatively small. The generation of artificial or miniature enzymes has long been a focus of research because enzyme mimetics can be produced with high activity at low cost. These enzymes aim to mimic the active sites without the additional architecture contributed by the protein chain. Previous work has shown that amyloidogenic peptides are able to self-assemble to create an active site that is capable of binding zinc and catalysing an esterase reaction. Here, we describe the structural characterisation of a set of designed peptides that form an amyloid-like architecture and reveal that their capability to mimic carbonic anhydrase and serve as enzyme-like catalysts is related to their ability to self-assemble. These amyloid fibril structures can bind the metal ion Zn 2+ via a three-dimensional arrangement of His residues created by the amyloid architecture. Our results suggest that the catalytic efficiency of amyloid-like assembly is not only zinc-dependent but also depends on an active centre created by the peptides which is, in turn, dependent on the ordered architecture. These fibrils have good esterase activity, and they may serve as good models for the evolution of modern-day enzymes. Furthermore, they may be useful in designing self-assembling fibrils for applications as metal ion catalysts. This study also demonstrates that the ligands surrounding the catalytic site affect the affinity of the zinc-binding site to bind the substrate contributing to the enzymatic activity of the assembled peptides.

The metal chaperone Atox1 regulates the activity of the human copper transporter ATP7B by modulating domain dynamics.

PubMed

Yu, Corey H; Yang, Nan; Bothe, Jameson; Tonelli, Marco; Nokhrin, Sergiy; Dolgova, Natalia V; Braiterman, Lelita; Lutsenko, Svetlana; Dmitriev, Oleg Y

2017-11-03

The human transporter ATP7B delivers copper to the biosynthetic pathways and maintains copper homeostasis in the liver. Mutations in ATP7B cause the potentially fatal hepatoneurological disorder Wilson disease. The activity and intracellular localization of ATP7B are regulated by copper, but the molecular mechanism of this regulation is largely unknown. We show that the copper chaperone Atox1, which delivers copper to ATP7B, and the group of the first three metal-binding domains (MBD1-3) are central to the activity regulation of ATP7B. Atox1-Cu binding to ATP7B changes domain dynamics and interactions within the MBD1-3 group and activates ATP hydrolysis. To understand the mechanism linking Atox1-MBD interactions and enzyme activity, we have determined the MBD1-3 conformational space using small angle X-ray scattering and identified changes in MBD dynamics caused by apo -Atox1 and Atox1-Cu by solution NMR. The results show that copper transfer from Atox1 decreases domain interactions within the MBD1-3 group and increases the mobility of the individual domains. The N-terminal segment of MBD1-3 was found to interact with the nucleotide-binding domain of ATP7B, thus physically coupling the domains involved in copper binding and those involved in ATP hydrolysis. Taken together, the data suggest a regulatory mechanism in which Atox1-mediated copper transfer activates ATP7B by releasing inhibitory constraints through increased freedom of MBD1-3 motions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The rhizotoxicity of metal cations is related to their strength of binding to hard ligands.

PubMed

Kopittke, Peter M; Menzies, Neal W; Wang, Peng; McKenna, Brigid A; Wehr, J Bernhard; Lombi, Enzo; Kinraide, Thomas B; Blamey, F Pax C

2014-02-01

Mechanisms whereby metal cations are toxic to plant roots remain largely unknown. Aluminum, for example, has been recognized as rhizotoxic for approximately 100 yr, but there is no consensus on its mode of action. The authors contend that the primary mechanism of rhizotoxicity of many metal cations is nonspecific and that the magnitude of toxic effects is positively related to the strength with which they bind to hard ligands, especially carboxylate ligands of the cell-wall pectic matrix. Specifically, the authors propose that metal cations have a common toxic mechanism through inhibiting the controlled relaxation of the cell wall as required for elongation. Metal cations such as Al(3+) and Hg(2+), which bind strongly to hard ligands, are toxic at relatively low concentrations because they bind strongly to the walls of cells in the rhizodermis and outer cortex of the root elongation zone with little movement into the inner tissues. In contrast, metal cations such as Ca(2+), Na(+), Mn(2+), and Zn(2+) , which bind weakly to hard ligands, bind only weakly to the cell wall and move farther into the root cylinder. Only at high concentrations is their weak binding sufficient to inhibit the relaxation of the cell wall. Finally, different mechanisms would explain why certain metal cations (for example, Tl(+), Ag(+), Cs(+), and Cu(2+)) are sometimes more toxic than expected through binding to hard ligands. The data presented in the present study demonstrate the importance of strength of binding to hard ligands in influencing a range of important physiological processes within roots through nonspecific mechanisms. © 2013 SETAC.

External protons destabilize the activated voltage sensor in hERG channels.

PubMed

Shi, Yu Patrick; Cheng, Yen May; Van Slyke, Aaron C; Claydon, Tom W

2014-03-01

Extracellular acidosis shifts hERG channel activation to more depolarized potentials and accelerates channel deactivation; however, the mechanisms underlying these effects are unclear. External divalent cations, e.g., Ca(2+) and Cd(2+), mimic these effects and coordinate within a metal ion binding pocket composed of three acidic residues in hERG: D456 and D460 in S2 and D509 in S3. A common mechanism may underlie divalent cation and proton effects on hERG gating. Using two-electrode voltage clamp, we show proton sensitivity of hERG channel activation (pKa = 5.6), but not deactivation, was greatly reduced in the presence of Cd(2+) (0.1 mM), suggesting a common binding site for the Cd(2+) and proton effect on activation and separable effects of protons on activation and deactivation. Mutational analysis confirmed that D509 plays a critical role in the pH dependence of activation, as shown previously, and that cooperative actions involving D456 and D460 are also required. Importantly, neutralization of all three acidic residues abolished the proton-induced shift of activation, suggesting that the metal ion binding pocket alone accounts for the effects of protons on hERG channel activation. Voltage-clamp fluorimetry measurements demonstrated that protons shifted the voltage dependence of S4 movement to more depolarized potentials. The data indicate a site and mechanism of action for protons on hERG activation gating; protonation of D456, D460 and D509 disrupts interactions between these residues and S4 gating charges to destabilize the activated configuration of S4.

Does Variation of the Inter-Domain Linker Sequence Modulate the Metal Binding Behaviour of Helix pomatia Cd-Metallothionein?

PubMed Central

Gil-Moreno, Selene; Jiménez-Martí, Elena; Palacios, Òscar; Zerbe, Oliver; Dallinger, Reinhard; Capdevila, Mercè; Atrian, Sílvia

2015-01-01

Snail metallothioneins (MTs) constitute an ideal model to study structure/function relationships in these metal-binding polypeptides. Helix pomatia harbours three MT isoforms: the highly specific CdMT and CuMT, and an unspecific Cd/CuMT, which represent paralogous proteins with extremely different metal binding preferences while sharing high sequence similarity. Preceding work allowed assessing that, although, the Cys residues are responsible for metal ion coordination, metal specificity or preference is achieved by diversification of the amino acids interspersed between them. The metal-specific MT polypeptides fold into unique, energetically-optimized complexes of defined metal content, when binding their cognate metal ions, while they produce a mixture of complexes, none of them representing a clear energy minimum, with non-cognate metal ions. Another critical, and so far mostly unexplored, region is the stretch linking the individual MT domains, each of which represents an independent metal cluster. In this work, we have designed and analyzed two HpCdMT constructs with substituted linker segments, and determined their coordination behavior when exposed to both cognate and non-cognate metal ions. Results unequivocally show that neither length nor composition of the inter-domain linker alter the features of the Zn(II)- and Cd(II)-complexes, but surprisingly that they influence their ability to bind Cu(I), the non-cognate metal ion. PMID:26703589

Metal stabilization of collagen and de novo designed mimetic peptides

PubMed Central

Parmar, Avanish S.; Xu, Fei; Pike, Douglas H.; Belure, Sandeep V.; Hasan, Nida F.; Drzewiecki, Kathryn E.; Shreiber, David I.; Nanda, Vikas

2017-01-01

We explore the design of metal binding sites to modulate triple-helix stability of collagen and collagen-mimetic peptides. Globular proteins commonly utilize metals to connect tertiary structural elements that are well separated in sequence, constraining structure and enhancing stability. It is more challenging to engineer structural metals into fibrous protein scaffolds, which lack the extensive tertiary contacts seen in globular proteins. In the collagen triple helix, the structural adjacency of the carboxy-termini of the three chains makes this region an attractive target for introducing metal binding sites. We engineered His3 sites based on structural modeling constraints into a series of designed homotrimeric and heterotrimeric peptides, assessing the capacity of metal binding to improve stability and in the case of heterotrimers, affect specificity of assembly. Notable enhancements in stability for both homo and heteromeric systems were observed upon addition of zinc(II) and several other metal ions only when all three histidine ligands were present. Metal binding affinities were consistent with the expected Irving-Williams series for imidazole. Unlike other metals tested, copper(II) also bound to peptides lacking histidine ligands. Acetylation of the peptide N-termini prevented copper binding, indicating proline backbone amide metal-coordination at this site. Copper similarly stabilized animal extracted Type I collagen in a metal specific fashion, highlighting the potential importance of metal homeostasis within the extracellular matrix. PMID:26225466

Metal Stabilization of Collagen and de Novo Designed Mimetic Peptides.

PubMed

Parmar, Avanish S; Xu, Fei; Pike, Douglas H; Belure, Sandeep V; Hasan, Nida F; Drzewiecki, Kathryn E; Shreiber, David I; Nanda, Vikas

2015-08-18

We explore the design of metal binding sites to modulate triple-helix stability of collagen and collagen-mimetic peptides. Globular proteins commonly utilize metals to connect tertiary structural elements that are well separated in sequence, constraining structure and enhancing stability. It is more challenging to engineer structural metals into fibrous protein scaffolds, which lack the extensive tertiary contacts seen in globular proteins. In the collagen triple helix, the structural adjacency of the carboxy-termini of the three chains makes this region an attractive target for introducing metal binding sites. We engineered His3 sites based on structural modeling constraints into a series of designed homotrimeric and heterotrimeric peptides, assessing the capacity of metal binding to improve stability and in the case of heterotrimers, affect specificity of assembly. Notable enhancements in stability for both homo- and heteromeric systems were observed upon addition of zinc(II) and several other metal ions only when all three histidine ligands were present. Metal binding affinities were consistent with the expected Irving-Williams series for imidazole. Unlike other metals tested, copper(II) also bound to peptides lacking histidine ligands. Acetylation of the peptide N-termini prevented copper binding, indicating proline backbone amide metal-coordination at this site. Copper similarly stabilized animal extracted Type I collagen in a metal-specific fashion, highlighting the potential importance of metal homeostasis within the extracellular matrix.

Exploration of cellular DNA lesion, DNA-binding and biocidal ordeal of novel curcumin based Knoevenagel Schiff base complexes incorporating tryptophan: Synthesis and structural validation

NASA Astrophysics Data System (ADS)

Chandrasekar, Thiravidamani; Raman, Natarajan

2016-07-01

A few novel Schiff base transition metal complexes of general formula [MLCl] (where, L = Schiff base, obtained by the condensation reaction of Knoevenagel condensate of curcumin, L-tryptophan and M = Cu(II), Ni(II), Co(II), and Zn(II)), were prepared by stencil synthesis. They were typified using UV-vis, IR, EPR spectral techniques, micro analytical techniques, magnetic susceptibility and molar conductivity. Geometry of the metal complexes was examined and recognized as square planar. DNA binding and viscosity studies revealed that the metal(II) complexes powerfully bound via an intercalation mechanism with the calf thymus DNA. Gel-electrophoresis technique was used to investigate the DNA cleavage competence of the complexes and they establish to approve the cleavage of pBR322 DNA in presence of oxidant H2O2. This outcome inferred that the synthesized complexes showed better nuclease activity. Moreover, the complexes were monitored for antimicrobial activities. The results exposed that the synthesized compounds were forceful against all the microbes under exploration.

β-Lactam antibiotics. Spectroscopy and molecular orbital (MO) calculations . Part I: IR studies of complexation in penicillin-transition metal ion systems and semi-empirical PM3 calculations on simple model compounds

NASA Astrophysics Data System (ADS)

Kupka, Teobald

1997-12-01

IR studies were preformed to determine possible transition metal ion binding sites of penicillin. the observed changes in spectral position and shape of characteristic IR bands of cloxacillin in the presence of transition metal ions (both in solutions and in the solid state) indicate formation of M-L complexes with engagement of -COO - and/or -CONH- functional groups. The small shift of νCO towards higher frequencies rules out direct M-L interaction via β-lactam carbonyl. PM3 calculations on simple model compounds (substituted formamide, cyclic ketones, lactams and substituted monocyclic β-lactams) have been performed. All structures were fully optimized and the calculated bond lengths, angles, heats of formation and CO stretching frequencies were discussed to determine the β-lactam binding sites and to explain its susceptibility towards nucleophilic attack (hydrolysis in vitro) and biological activity. The relative changes of calculated values were critically compared with available experimental data and same correlation between structural parameters and in vivo activity was shown.

SOD activity and DNA binding properties of a new symmetric porphyrin Schiff base ligand and its metal complexes.

PubMed

Çay, Sevim; Köse, Muhammet; Tümer, Ferhan; Gölcü, Ayşegül; Tümer, Mehmet

2015-12-05

4-Methoxy-2,6-bis(hydroxymethyl)phenol (1) was prepared from the reaction of 4-methoxyphenol and formaldehyde. The compound (1) was then oxidized to the 4-methoxy-2,6-diformylphenol (2) compound. Molecular structure of compound (2) was determined by X-ray diffraction method. A new symmetric porphyrin Schiff base ligand 4-methoxy-2,6-bis[5-(4-iminophenyl)-10,15,20-triphenylporphyrin]phenol (L) was prepared from the reaction of the 5-(4-aminophenyl)-10,15,20-triphenylporphyrin (TTP-NH2) and the compound (2) in the toluene solution. The metal complexes (Cu(II), Fe(III), Mn(III), Pt(II) and Zn(II)) of the ligand (L) were synthesized and characterized by the spectroscopic and analytical methods. The DNA (fish sperm FSdsDNA) binding studies of the ligand and its complexes were performed using UV-vis spectroscopy. Additionally, superoxide dismutase activities of the porphyrin Schiff base metal complexes were investigated. Additionally, electrochemical, photoluminescence and thermal properties of the compounds were investigated. Copyright © 2015 Elsevier B.V. All rights reserved.

SPECT/CT localization of oral radioiodine activity: a retrospective study and in-vitro assessment.

PubMed

Burlison, Jared S; Hartshorne, Michael F; Voda, Alan M; Cocks, Franklin H; Fair, Joanna R

2013-12-01

We sought to further localize radioiodine activity in the mouth on post-thyroid cancer therapy imaging using single-photon emission computed tomography/computed tomography (SPECT/CT). We retrospectively reviewed all patients (58) who underwent thyroid cancer therapy with iodine-131 (131I) at our institution from August 2009 to March 2011 whose post-therapy radioiodine imaging included neck SPECT/CT. A small group (six) of diagnostic 131I scans including SPECT/CT was also reviewed. Separately, we performed in-vitro 131I (sodium iodide) binding assays with amalgam and Argenco HP 77 (77% dental gold alloy) as proof of principle for these interactions. Of the 58 post-therapy patients, 45 (78%) had undergone metallic dental restorations, and of them 41 (91%) demonstrated oral 131I activity localizing preferentially to those restorations. It was observed that radioiodine also localized to other dental restorations and to orthodontic hardware. Gum-line activity in edentulous patients suggests radioiodine interaction with denture adhesive. In vitro, dental amalgam and Argenco HP 77 bound 131I in a time-dependent manner over 1-16 days of exposure. Despite subsequent washings with normal saline, significant 131I activity (maximally 12% for amalgam and 68% for Argenco HP 77) was retained by these metals. Subsequent soaking in a saturated solution of potassium iodide partially displaced 131I from amalgam, with near-total displacement of I from Argenco HP 77. SPECT/CT shows that radioiodine in the oral cavity localizes to metallic dental restorations. Furthermore, in-vitro studies demonstrate partially reversible binding of 131I to common dental metals.

Pro-inflammatory effects of metals in persons and animals exposed to tobacco smoke.

PubMed

Milnerowicz, Halina; Ściskalska, Milena; Dul, Magdalena

2015-01-01

Metals present in tobacco smoke have the ability to cause a pro-oxidant/antioxidant imbalance through the direct generation of free radicals in accordance with the Fenton or Haber-Weiss reaction and redox properties. Metals can also interact with antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase) and small molecular antioxidants (glutathione) through binding to SH groups or by replacement of metals ions in the catalytic center of enzymes. Excessive free radicals production can induce an inflammatory response. The aim of this study was to review the information on the induction of inflammation by metals present in tobacco smoke such as lead (Pb), cadmium (Cd), arsenic (As), aluminum (Al), nickel (Ni) and mercury (Hg). In cellular immune response, it was demonstrated that radicals induced by metals can disrupt the transcription signaling pathway mediated by the mitogen-activated protein kinase (induced by Pb), NLRP3-ASC-caspase 1 (induced by Ni), tyrosine kinase Src (induced by As) and the nuclear factor κB (induced by Pb, Ni, Hg). The result of this is a gene transcription for early inflammatory cytokines, such as Interleukine 1β, Interleukine 6, and Tumor necrosis factor α). These cytokines can cause leukocytes recruitment and secretions of other pro-inflammatory cytokines and chemokines, which intensifies the inflammatory response. Some metals, such as cadmium (Cd), can activate an inflammatory response through tissue damage induction mediated by free radicals, which also results in leukocytes recruitment and cytokines secretions. Inflammation generated by metals can be reduced by metallothionein, which has the ability to scavenge free radicals and bind toxic metals through the release of Zn and oxidation of SH groups. Copyright © 2014 Elsevier GmbH. All rights reserved.

Modulating supramolecular binding of carbon dioxide in a redox-active porous metal-organic framework

DOE PAGES

Lu, Zhenzhong; Godfrey, Harry G. W.; da Silva, Ivan; ...

2017-02-13

Hydrogen bonds dominate many chemical and biological processes, and chemical modification enables control and modulation of host–guest systems. Here in this paper we report a targeted modification of hydrogen bonding and its effect on guest binding in redox-active materials. MFM-300(V III) {[V III 2(OH) 2(L)], LH 4=biphenyl-3,3',5,5'-tetracarboxylic acid} can be oxidized to isostructural MFM-300(V IV), [V IV 2O 2(L)], in which deprotonation of the bridging hydroxyl groups occurs. MFM-300(V III) shows the second highest CO 2 uptake capacity in metal-organic framework materials at 298 K and 1 bar (6.0 mmol g -1) and involves hydrogen bonding between the OH group of the host and the O-donor of CO 2, which binds in an end-on manner, OH∙∙∙ =1.863(1) Å. In contrast, CO 2-loaded MFM-300(V IV) shows CO 2 bound side-on to the oxy group and sandwiched between two phenyl groups involving a unique O COmore » $$_2$$···c.g.phenyl interaction [3.069(2), 3.146(3) Å]. Lastly, the macroscopic packing of CO 2 in the pores is directly influenced by these primary binding sites.« less

Metal ion-dependent DNAzymes and their applications as biosensors.

PubMed

Lan, Tian; Lu, Yi

2012-01-01

Long considered to serve solely as the genetic information carrier, DNA has been shown in 1994 to be able to act as DNA catalysts capable of catalyzing a trans-esterification reaction similar to the action of ribozymes and protein enzymes. Although not yet found in nature, numerous DNAzymes have been isolated through in vitro selection for catalyzing many different types of reactions in the presence of different metal ions and thus become a new class of metalloenzymes. What remains unclear is how DNA can carry out catalysis with simpler building blocks and fewer functional groups than ribozymes and protein enzymes and how DNA can bind metal ions specifically to perform these functions. In the past two decades, many biochemical and biophysical studies have been carried out on DNAzymes, especially RNA-cleaving DNAzymes. Important insights have been gained regarding their metal-dependent activity, global folding, metal binding sites, and catalytic mechanisms for these DNAzymes. Because of their high metal ion selectivity, one of the most important practical applications for DNAzymes is metal ion detection, resulting in highly sensitive and selective fluorescent, colorimetric, and electrochemical sensors for a wide range of metal ions such as Pb(2+), UO2 2 +,[Formula: see text] including paramagnetic metal ions such as Cu(2+). This chapter summarizes recent progresses in in vitro selection of metal ion-selective DNAzymes, their biochemical and biophysical studies and sensing applications.

Drug Delivery Systems For Anti-Cancer Active Complexes of Some Coinage Metals.

PubMed

Zhang, Ming; Saint-Germain, Camille; He, Guiling; Sun, Raymond Wai-Yin

2018-02-12

Although cisplatin and a number of platinum complexes have widely been used for the treatment of neoplasia, patients receiving these treatments have frequently suffered from their severe toxic side effects, the development of resistance with consequent relapse. In the recent decades, numerous complexes of coinage metals including that of gold, copper and silver have been reported to display promising in vitro and/or in vivo anti-cancer activities as well as potent activities towards cisplatin-resistant tumors. Nevertheless, the medical development of these metal complexes has been hampered by their instability in aqueous solutions and the nonspecific binding in biological systems. One of the approaches to overcome these problems is to design and develop adequate drug delivery systems (DDSs) for the transport of these complexes. By functionalization, encapsulation or formulation of the metal complexes, several types of DDSs have been reported to improve the desired pharmacological profile of the metal complexes, improving their overall stability, bioavailability, anti-cancer activity and reducing their toxicity towards normal cells. In this review, we summarized the recent findings for different DDSs for various anti- cancer active complexes of some coinage metals. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Crystal structure, DNA binding, cleavage, antioxidant and antibacterial studies of Cu(II), Ni(II) and Co(III) complexes with 2-((furan-2-yl)methylimino)methyl)-6-ethoxyphenol Schiff base

NASA Astrophysics Data System (ADS)

Venkateswarlu, Kadtala; Kumar, Marri Pradeep; Rambabu, Aveli; Vamsikrishna, Narendrula; Daravath, Sreenu; Rangan, Krishnan; Shivaraj

2018-05-01

Three novel binary metal complexes; 1 [Cu(L)2], 2 [Ni(L)2] and 3 [Co(L)3] where, L (2-(((furan-2-yl) methylimino)methyl)-6-ethoxyphenol, C14H15NO3), were synthesized and characterized by various spectral techniques. Based on spectral studies square planar geometry is assigned for Cu(II) and Ni(II) complexes, whereas Co(III) owned octahedral geometry. Ligand, [Cu(L)2] and [Ni(L)2] are crystallized and found to be monoclinic crystal systems. CT-DNA absorption binding studies revealed that the complexes show good binding propensity (Kb = 5.02 × 103 M-1, 2.77 × 103 M-1, 1.63 × 104 M-1 for 1, 2 and 3 respectively). The role of these complexes in the oxidative and photolytic cleavage of supercoiled pBR322 DNA was studied and found that the complexes cleave the pBR322 DNA effectively. The catalytic ability of 1, 2 and 3 follows the order: 3 > 1 >2. Antioxidant studies of the new complexes revealed that they exhibit significant antioxidant activity against DPPH radical. The Schiff base and its metal complexes have been screened for antibacterial studies by Minimum Inhibitory Concentration method. It is observed that all metal complexes showed more activity than free ligand.

Gain-of-function mutations identify amino acids within transmembrane domains of the yeast vacuolar transporter Zrc1 that determine metal specificity

PubMed Central

Lin, Huilan; Burton, Damali; Li, Liangtao; Warner, David E.; Phillips, John D.; Ward, Diane McVEY; Kaplan, Jerry

2015-01-01

Cation diffusion facilitator transporters are found in all three Kingdoms of life and are involved in transporting transition metals out of the cytosol. The metals they transport include Zn2+, Co2+, Fe2+, Cd2+, Ni2+ and Mn2+; however, no single transporter transports all metals. Previously we showed that a single amino acid mutation in the yeast vacuolar zinc transporter Zrc1 changed its substrate specificity from Zn2+ to Fe2+ and Mn2+ [Lin, Kumanovics, Nelson, Warner, Ward and Kaplan (2008) J. Biol. Chem. 283, 33865–33873]. Mutant Zrc1 that gained iron transport activity could protect cells with a deletion in the vacuolar iron transporter (CCC1) from high iron toxicity. Utilizing suppression of high iron toxicity and PCR mutagenesis of ZRC1, we identified other amino acid substitutions within ZRC1 that changed its metal specificity. All Zrc1 mutants that transported Fe2+ could also transport Mn2+. Some Zrc1 mutants lost the ability to transport Zn2+, but others retained the ability to transport Zn2+. All of the amino acid substitutions that resulted in a gain in Fe2+ transport activity were found in transmembrane domains. In addition to alteration of residues adjacent to the putative metal-binding site in two transmembrane domains, alteration of residues distant from the binding site affected substrate specificity. These results suggest that substrate selection involves co-operativity between transmembrane domains. PMID:19538181

AIE active multianalyte fluorescent probe for the detection of Cu2+, Ni2+ and Hg2+ ions.

PubMed

Pannipara, Mehboobali; Al-Sehemi, Abdullah G; Irfan, Ahmad; Assiri, Mohammed; Kalam, Abul; Al-Ammari, Yahya S

2018-08-05

A novel pyrazolyl chromene derivative (Probe 1) displaying aggregation induced emission (AIE) properties that capable of sensing of multiple metal ions has been designed and synthesized. The multi analyte probe exhibits selective sensing for Cu 2+ and Ni 2+ ions via fluorescence turn-off mechanism and ratiometric selectivity for Hg 2+ ions in aqueous media. The extent of binding of the probe with sensitive metal ions has been demonstrated. The experimental results were further investigated by computational means by optimizing the ground state geometries of Probe 1 and its various metal complexes for Probe 1-Ni, Probe 1-Hg and Probe 1-Cu using density functional theory (DFT) at B3LYP/6-31+g(d,p) (LANL2DZ) level. On the basis of binding energies, the stability of metal complexes has been studied. In Probe 1-Ni and Probe 1-Cu complexes, charge transfer has been observed from Probe 1 to metal ions revealing ligand to metal charge transfer (LMCT) while in Probe1-Hg complex LMCT as well as intra-molecular charge tranfer (ICT) within Probe 1. Copyright © 2018 Elsevier B.V. All rights reserved.

Biosorption of heavy metal ions to brown algae, Macrocystis pyrifera, Kjellmaniella crassiforia, and Undaria pinnatifida

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seki, Hideshi; Suzuki, Akira

1998-10-01

A fundamental study of the application of brown algae to the aqueous-phase separation of toxic heavy metals was carried out. The biosorption characteristics of cadmium and lead ions were determined with brown algae, Macrocystis pyrifera, Kjellmaniella crassiforia, and Undaria pinnatifida. A metal binding model proposed by the authors was used for the description of metal binding data. The results showed that the biosorption of bivalent metal ions to brown algae was due to bivalent binding to carboxylic groups on alginic acid in brown algae.

Selectivity of externally facing ion-binding sites in the Na/K pump to alkali metals and organic cations

PubMed Central

Ratheal, Ian M.; Virgin, Gail K.; Yu, Haibo; Roux, Benoît; Gatto, Craig; Artigas, Pablo

2010-01-01

The Na/K pump is a P-type ATPase that exchanges three intracellular Na+ ions for two extracellular K+ ions through the plasmalemma of nearly all animal cells. The mechanisms involved in cation selection by the pump's ion-binding sites (site I and site II bind either Na+ or K+; site III binds only Na+) are poorly understood. We studied cation selectivity by outward-facing sites (high K+ affinity) of Na/K pumps expressed in Xenopus oocytes, under voltage clamp. Guanidinium+, methylguanidinium+, and aminoguanidinium+ produced two phenomena possibly reflecting actions at site III: (i) voltage-dependent inhibition (VDI) of outwardly directed pump current at saturating K+, and (ii) induction of pump-mediated, guanidinium-derivative–carried inward current at negative potentials without Na+ and K+. In contrast, formamidinium+ and acetamidinium+ induced K+-like outward currents. Measurement of ouabain-sensitive ATPase activity and radiolabeled cation uptake confirmed that these cations are external K+ congeners. Molecular dynamics simulations indicate that bound organic cations induce minor distortion of the binding sites. Among tested metals, only Li+ induced Na+-like VDI, whereas all metals tested except Na+ induced K+-like outward currents. Pump-mediated K+-like organic cation transport challenges the concept of rigid structural models in which ion specificity at site I and site II arises from a precise and unique arrangement of coordinating ligands. Furthermore, actions by guanidinium+ derivatives suggest that Na+ binds to site III in a hydrated form and that the inward current observed without external Na+ and K+ represents cation transport when normal occlusion at sites I and II is impaired. These results provide insights on external ion selectivity at the three binding sites. PMID:20937860

Universal binding energy relations in metallic adhesion

NASA Technical Reports Server (NTRS)

Ferrante, J.; Smith, J. R.; Rose, J. H.

1981-01-01

Scaling relations which map metallic adhesive binding energy onto a single universal binding energy curve are discussed in relation to adhesion, friction, and wear in metals. The scaling involved normalizing the energy to the maximum binding energy and normalizing distances by a suitable combination of Thomas-Fermi screening lengths. The universal curve was found to be accurately represented by E*(A*)= -(1+beta A) exp (-Beta A*) where E* is the normalized binding energy, A* is the normalized separation, and beta is the normalized decay constant. The calculated cohesive energies of potassium, barium, copper, molybdenum, and samarium were also found to scale by similar relations, suggesting that the universal relation may be more general than for the simple free electron metals.

Glyceraldehyde-3-phosphate dehydrogenase from Chironomidae showed differential activity towards metals.

PubMed

Chong, Isaac K W; Ho, Wing S

2013-09-01

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is known to interact with different biomolecules and was implicated in many novel cellular activities including programmed cell death, nuclear RNA transport unrelated to the commonly known carbohydrate metabolism. We reported here the purification of GAPDH from Chironomidae larvae (Insecta, Diptera) that showed different biologic activity towards heavy metals. It was inhibited by copper, cobalt nickel, iron and lead but was activated by zinc. The GAPDH was purified by ammonium sulphate fractionation and Chelating Sepharose CL-6B chromatography followed by Blue Sepharose CL-6B chromatography. The 150-kDa tetrameric GAPDH showed optimal activity at pH 8.5 and 37°C. The multiple alignment of sequence of the Chironomidae GAPDH with other known species showed 78 - 88% identity to the conserved regions of the GADPH. Bioinformatic analysis unveils substantial N-terminal sequence similarity of GAPDH of Chironomidae larvae to mammalian GADPHs. However, the GADPH of Chironomidae larvae showed different biologic activities and cytotoxicity towards heavy metals. The GAPDH enzyme would undergo adaptive molecular changes through binding at the active site leading to higher tolerance to heavy metals.

Metal binding proteins, recombinant host cells and methods

DOEpatents

Summers, Anne O.; Caguiat, Jonathan J.

2004-06-15

The present disclosure provides artificial heavy metal binding proteins termed chelons by the inventors. These chelons bind cadmium and/or mercuric ions with relatively high affinity. Also disclosed are coding sequences, recombinant DNA molecules and recombinant host cells comprising those recombinant DNA molecules for expression of the chelon proteins. In the recombinant host cells or transgenic plants, the chelons can be used to bind heavy metals taken up from contaminated soil, groundwater or irrigation water and to concentrate and sequester those ions. Recombinant enteric bacteria can be used within the gastrointestinal tracts of animals or humans exposed to toxic metal ions such as mercury and/or cadmium, where the chelon recombinantly expressed in chosen in accordance with the ion to be rededicated. Alternatively, the chelons can be immobilized to solid supports to bind and concentrate heavy metals from a contaminated aqueous medium including biological fluids.

Crown oxygen-doping graphene with embedded main-group metal atoms

NASA Astrophysics Data System (ADS)

Wu, Liyuan; Wang, Qian; Yang, Chuanghua; Quhe, Ruge; Guan, Pengfei; Lu, Pengfei

2018-02-01

Different main-group metal atoms embedded in crown oxygen-doping graphene (metal@OG) systems are studied by the density functional theory. The binding energies and electronic structures are calculated by using first-principles calculations. The binding energy of metal@OG system mainly depends on the electronegativity of the metal atom. The lower the value of the electronegativity, the larger the binding energy, indicating the more stable the system. The electronic structure of metal@OG arouses the emergence of bandgap and shift of Dirac point. It is shown that interaction between metal atom and crown oxygen-doping graphene leads to the graphene's stable n-doping, and the metal@OG systems are stable semiconducting materials, which can be used in technological applications.

A comparative study for Hydrogen storage in metal decorated graphyne nanotubes and graphyne monolayers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Jinlian; Guo, Yanhua; Zhang, Yun

A comparative study for hydrogen storage in metal decorated graphyne nanotubes and graphyne monolayers has been investigated within the framework of first-principle calculations. Our results show that the binding energies of Li, Ca, Sc, Ti on graphyne nanotubes are stronger than that on graphyne monolayers. Such strong binding would prevent the formation of metal clusters on graphyne nanotubes. From the charge transfer and partial density of states, it is found that the curvature effect of nanotubes plays an important role for the strong binding strength of metal on graphyne nanotubes. And the hydrogen storage capacity is 4.82 wt%, 5.08 wt%,more » 4.88 wt%, 4.76 wt% for Li, Ca, Sc, Ti decorated graphyne nanotubes that promise a potential material for storing hydrogen. - Graphical abstract: Metal atoms (Li, Ca, Sc and Ti) can strongly bind to graphyne nanotubes to avoid the formation of metal clusters, and a capacity of Ca@graphyne nanotube is 5.08 wt% which is close to the requirement of DOE in 2015. Twenty-four hydrogen molecules absorb to Ti-decorated graphyne nanotube. - Highlights: • The binding strength for metal on graphyne nanotubes is much stronger than that on γ-graphyne monolayer. • Metal atoms can strongly bind to the curving triangle acetylenes rings to avoid the formation of metal clusters. • A capacity of Ca@graphyne nanotube is 5.08 wt% which is close to the requirement of DOE in 2015.« less

Lead-binding capacity of calcium pectates with different molecular weight.

PubMed

Khotimchenko, Maksim; Makarova, Ksenia; Khozhaenko, Elena; Kovalev, Valeri

2017-04-01

Nowadays, heavy metal contamination of environment is considered as a serious threat to public health because of toxicity of these pollutants and the lack of effective materials with metal-binding properties. Some biopolymers such as pectins were proposed for removal of metal ions from industrial water disposals. Chemical structure of pectins is quite variable and substantially affects their metal binding properties. In this work, relationship between molecular weight and Pb(II)-binding capacity of calcium pectates was investigated in a batch sorption system. The results showed that all pectate samples are able to form complexes with Pb(II) ions. The effects of contact time, pH of the media and equilibrium metal concentration on metal-binding process were tested in experiments. The equilibrium time min required for uptake of Pb(II) by pectate compounds was found to be 60min. Langmuir and Freundlich models were applied for description of interactions between pectates and metal ions. Binding capacity of low molecular pectate was highest among all the samples tested. Langmuir model was figured out to be the best fit within the whole range of pH values. These results demonstrate that calcium pectate with low molecular weight is more promising agent for elimination of Pb(II) ions from contaminated wastewaters. Copyright © 2017 Elsevier B.V. All rights reserved.

Mercury(II) binds to both of chymotrypsin's histidines, causing inhibition followed by irreversible denaturation/aggregation.

PubMed

Stratton, Amanda; Ericksen, Matthew; Harris, Travis V; Symmonds, Nick; Silverstein, Todd P

2017-02-01

The toxicity of mercury is often attributed to its tight binding to cysteine thiolate anions in vital enzymes. To test our hypothesis that Hg(II) binding to histidine could be a significant factor in mercury's toxic effects, we studied the enzyme chymotrypsin, which lacks free cysteine thiols; we found that chymotrypsin is not only inhibited, but also denatured by Hg(II). We followed the aggregation of denatured enzyme by the increase in visible absorbance due to light scattering. Hg(II)-induced chymotrypsin precipitation increased dramatically above pH 6.5, and free imidazole inhibited this precipitation, implicating histidine-Hg(II) binding in the process of chymotrypsin denaturation/aggregation. Diethylpyrocarbonate (DEPC) blocked chymotrypsin's two histidines (his 40 and his 57 ) quickly and completely, with an IC 50 of 35 ± 6 µM. DEPC at 350 µM reduced the hydrolytic activity of chymotrypsin by 90%, suggesting that low concentrations of DEPC react with his 57 at the active site catalytic triad; furthermore, DEPC below 400 µM enhanced the Hg(II)-induced precipitation of chymotrypsin. We conclude that his 57 reacts readily with DEPC, causing enzyme inhibition and enhancement of Hg(II)-induced aggregation. Above 500 µM, DEPC inhibited Hg(II)-induced precipitation, and [DEPC] >2.5 mM completely protected chymotrypsin against precipitation. This suggests that his 40 reacts less readily with DEPC, and that chymotrypsin denaturation is caused by Hg(II) binding specifically to the his 40 residue. Finally, we show that Hg(II)-histidine binding may trigger hemoglobin aggregation as well. Because of results with these two enzymes, we suggest that metal-histidine binding may be key to understanding all heavy metal-induced protein aggregation. © 2017 The Protein Society.

Cyclic Tetrapyrrole Sulfonation, Metals, and Oligomerization in Antiprion Activity▿

PubMed Central

Caughey, Winslow S.; Priola, Suzette A.; Kocisko, David A.; Raymond, Lynne D.; Ward, Anne; Caughey, Byron

2007-01-01

Cyclic tetrapyrroles are among the most potent compounds with activity against transmissible spongiform encephalopathies (TSEs; or prion diseases). Here the effects of differential sulfonation and metal binding to cyclic tetrapyrroles were investigated. Their potencies in inhibiting disease-associated protease-resistant prion protein were compared in several types of TSE-infected cell cultures. In addition, prophylactic antiscrapie activities were determined in scrapie-infected mice. The activity of phthalocyanine was relatively insensitive to the number of peripheral sulfonate groups but varied with the type of metal bound at the center of the molecule. The tendency of the various phthalocyanine sulfonates to oligomerize (i.e., stack) correlated with anti-TSE activity. Notably, aluminum(III) phthalocyanine tetrasulfonate was both the poorest anti-TSE compound and the least prone to oligomerization in aqueous media. Similar comparisons of iron- and manganese-bound porphyrin sulfonates confirmed that stacking ability correlates with anti-TSE activity among cyclic tetrapyrroles. PMID:17709470

Price To Be Paid for Two-Metal Catalysis: Magnesium Ions That Accelerate Chemistry Unavoidably Limit Product Release from a Protein Kinase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacobsen, Douglas M.; Bao, Zhao-Qin; O'’Brien, Patrick

Incorporation of divalent metal ions into an active site is a fundamental catalytic tool used by diverse enzymes. Divalent cations are used by protein kinases to both stabilize ATP binding and accelerate chemistry. Kinetic analysis establishes that Cyclin-dependent kinase 2 (CDK2) requires simultaneous binding of two Mg 2+ ions for catalysis of phosphoryl transfer. This tool, however, comes with a price: the rate-acceleration effects are opposed by an unavoidable rate-limiting consequence of the use of two Mg 2+ ions by CDK2. The essential metal ions stabilize ADP product binding and limit the overall rate of the reaction. We demonstrate thatmore » product release is rate limiting for activated CDK2 and evaluate the effects of the two catalytically essential Mg 2+ ions on the stability of the ADP product within the active site. We present two new crystal structures of CDK2 bound to ADP showing how the phosphate groups can be coordinated by either one or two Mg 2+ ions, with the occupancy of one site in a weaker equilibrium. Molecular dynamics simulations indicate that ADP phosphate mobility is more restricted when ADP is coordinated by two Mg 2+ ions compared to one. The structural similarity between the rigid ADP·2Mg product and the cooperatively assembled transition state provides a mechanistic rational for the rate-limiting ADP release that is observed. We demonstrate that although the simultaneous binding of two Mg 2+ ions is essential for efficient phosphoryl transfer, the presence of both Mg 2+ ions in the active site also cooperatively increases ADP affinity and opposes its release. Evolution of protein kinases must have involved careful tuning of the affinity for the second Mg 2+ ion in order to balance the needs to stabilize the chemical transition state and allow timely product release. The link between Mg 2+ site affinity and activity presents a chemical handle that may be used by regulatory factors as well as explain some mutational effects.« less

Interfacial and Alloying Effects on Activation of Ethanol from First-Principles

DOE PAGES

An, Wei; Men, Yong; Wang, Jinguo; ...

2017-02-24

Here, we present a first-principles density-functional theory study of ethanol activation at oxide/Rh(111) interface and the alloying effect on mitigating carbon deposition, which are essential to direct ethanol fuel cell (DEFC) anode reaction and steam reforming of ethanol (SRE) reaction. Our calculated results show that charge can transfer from Rh(111) substrate to MO x chain (e.g., MoO 3 and MnO 2), or from MO x chain (e.g., MgO, SnO 2, ZrO 2, and TiO 2) to Rh(111) substrate. The OH-binding strength is increased exponentially with M δ+ charge ranging from 1.4 to 2.2, which renders MnO 2/Rh(111) and MgO/Rh(111) interfacesmore » weaker OH-binding, and thereby enhanced oxidizing functionality of OH* for promoting ethanol oxidation reaction (EOR) at DEFC anode. For efficient C–C bond breaking, a large number of Rh ensemble sizes are critically needed at the interface of MO x/Rh(111). We found that Rh 1Au 3 near surface alloy has the weakest C* and CO* binding, followed by Rh 1Cu 3 and Rh 1Pd 3 near surface alloys, while Rh 1Ir 3 and Rh 1Ru 3 surface alloys have C* and CO* binding strength similar to that of pure Rh metal. The general implication of this study is that by engineering alloyed structure of weakened C* and CO* binding complemented with metal oxides of weakened OH-binding, high-performance DEFC anode or SRE catalysts can be identified.« less

Metal complexation properties of freshwater dissolved organic matter are explained by its aromaticity and by anthropogenic ligands.

PubMed

Baken, Stijn; Degryse, Fien; Verheyen, Liesbeth; Merckx, Roel; Smolders, Erik

2011-04-01

Dissolved organic matter (DOM) in surface waters affects the fate and environmental effects of trace metals. We measured variability in the Cd, Cu, Ni, and Zn affinity of 23 DOM samples isolated by reverse osmosis from freshwaters in natural, agricultural, and urban areas. Affinities at uniform pH and ionic composition were assayed at low, environmentally relevant free Cd, Cu, Ni, and Zn activities. The C-normalized metal binding of DOM varied 4-fold (Cu) or about 10-fold (Cd, Ni, Zn) among samples. The dissolved organic carbon concentration ranged only 9-fold in the waters, illustrating that DOM quality is an equally important parameter for metal complexation as DOM quantity. The UV-absorbance of DOM explained metal affinity only for waters receiving few urban inputs, indicating that in those waters, aromatic humic substances are the dominant metal chelators. Larger metal affinities were found for DOM from waters with urban inputs. Aminopolycarboxylate ligands (mainly EDTA) were detected at concentrations up to 0.14 μM and partly explained the larger metal affinity. Nickel concentrations in these surface waters are strongly related to EDTA concentrations (R2=0.96) and this is underpinned by speciation calculations. It is concluded that metal complexation in waters with anthropogenic discharges is larger than that estimated with models that only take into account binding on humic substances.

Modulation of nitric oxide-mediated metal release from metallothionein by the redox state of glutathione in vitro.

PubMed

Khatai, Leila; Goessler, Walter; Lorencova, Helena; Zangger, Klaus

2004-06-01

Metallothioneins (MTs) release bound metals when exposed to nitric oxide. At inflammatory sites, both metallothionein and inducible nitric oxide synthase (iNOS) are induced by the same factors and the zinc released from metallothionein by NO suppresses both the induction and activity of iNOS. In a search for a possible modulatory mechanism of this coexpression of counteracting proteins, we investigated the role of the glutathione redox state in vitro because the oxidation state of thiols is involved in the metal binding in Cd-S or Zn-S clusters found in metallothioneins, and NO also binds to reduced glutathione via S-nitrosation. Using a variety of techniques, we found that NO and also ONOO(-)-mediated metal release from purified MTs is suppressed by reduced glutathione (GSH), but not by oxidized glutathione. Considering the millimolar concentrations of GSH present in mammalian cells, the metal release from MTs by NO should play no role in living systems. Therefore, the fact that it has been observed in vivo points to a hitherto unknown mechanism or additional compound(s) being involved in this physiologically relevant reaction and as long as this additional factor is not found experimental results on the MT-NO interaction should be treated with caution. Contrary to the peroxynitrite-induced activation of guanylyl cyclase, where GSH is needed, we found that the metal release from metallothionein by peroxynitrite is not enhanced, but also suppressed by reduced glutathione. In addition, we show that zinc, the major natural metal ligand in mammalian MTs and suppressor of iNOS, is released more readily under the influence of NO than cadmium, but in contrast to the MT isoform 1, the amount of metal released from the beta-domain of MT-2 is comparable to that from the alpha-domain.

The Novel Helicobacter pylori CznABC Metal Efflux Pump Is Required for Cadmium, Zinc, and Nickel Resistance, Urease Modulation, and Gastric Colonization

PubMed Central

Stähler, Frank Nils; Odenbreit, Stefan; Haas, Rainer; Wilrich, Julia; Vliet, Arnoud H. M. Van; Kusters, Johannes G.; Kist, Manfred; Bereswill, Stefan

2006-01-01

Maintaining metal homeostasis is crucial for the adaptation of Helicobacter pylori to the gastric environment. Iron, copper, and nickel homeostasis has recently been demonstrated to be required for the establishment of H. pylori infection in animal models. Here we demonstrate that the HP0969-0971 gene cluster encoding the Czc-type metal export pump homologs HP0969, HP0970, and the H. pylori-specific protein HP0971 forms part of a novel H. pylori metal resistance determinant, which is required for gastric colonization and for the modulation of urease activity. Insertional mutagenesis of the HP0971, HP0970, or HP0969 genes in H. pylori reference strain 26695 resulted in increased sensitivity to cadmium, zinc, and nickel (czn), suggesting that the encoded proteins constitute a metal-specific export pump. Accordingly, the genes were designated cznC (HP0971), cznB (HP0970), and cznA (HP0969). The CznC and CznA proteins play a predominant role in nickel homeostasis, since only the cznC and cznA mutants but not the cznB mutant displayed an 8- to 10-fold increase in urease activity. Nickel-specific affinity chromatography demonstrated that recombinant versions of CznC and CznB can bind to nickel and that the purified CznB protein interacted with cadmium and zinc, since both metals competitively inhibited nickel binding. Finally, single cznA, cznB, and cznC mutants did not colonize the stomach in a Mongolian gerbil-based animal model. This demonstrates that the metal export functions of H. pylori cznABC are essential for gastric colonization and underlines the extraordinary importance of metal ion homeostasis for the survival of H. pylori in the gastric environment. PMID:16790756

Biological Nanoplatforms for Self-Assembled Electronics

DTIC Science & Technology

2015-03-24

as M13 , a virus that infects Escherichia coli. Approximately one billion different amino acid sequences are displayed on different viruses in the...sequence when contained within a phage M13 coat protein sequence, not chemically linked to the surface of phage MS2 VLPs. Thus, binding properties may...gallium arsenide in a bacteriophage M13 phage display library, MS2 VLPs modified with the metal binding peptides do not display the same activity

Inhibition of Monometalated Methionine Aminopeptidase: Inhibitor Discovery and Crystallographic Analysis†

PubMed Central

Huang, Min; Xie, Sheng-Xue; Ma, Ze-Qiang; Huang, Qing-Qing; Nan, Fa-Jun; Ye, Qi-Zhuang

2008-01-01

Two divalent metal ions are commonly seen in the active site cavity of methionine aminopeptidase, and at least one of the metal ions is directly involved in catalysis. Although ample structural and functional information is available for dimetalated enzyme, methionine aminopeptidase likely functions as a monometalated enzyme under physiological conditions. Information on structure, as well as catalysis and inhibition, of the monometalated enzyme is lacking. By improving conditions of high throughput screening, we identified a unique inhibitor with specificity toward the monometalated enzyme. Kinetic characterization indicates a mutual exclusivity in binding between the inhibitor and the second metal ion at the active site. This is confirmed by X-ray structure, and this inhibitor coordinates with the first metal ion and occupies the space normally occupied by the second metal ion. Kinetic and structural analyses of the inhibition by this and other inhibitors provide insight in designing effective inhibitors of methionine aminopeptidase. PMID:17948983

Catalytic zinc site and mechanism of the metalloenzyme PR-AMP cyclohydrolase.

PubMed

D'Ordine, Robert L; Linger, Rebecca S; Thai, Carolyn J; Davisson, V Jo

2012-07-24

The enzyme N(1)-(5'-phosphoribosyl) adenosine-5'-monophosphate cyclohydrolase (PR-AMP cyclohydrolase) is a Zn(2+) metalloprotein encoded by the hisI gene. It catalyzes the third step of histidine biosynthesis, an uncommon ring-opening of a purine heterocycle for use in primary metabolism. A three-dimensional structure of the enzyme from Methanobacterium thermoautotrophicum has revealed that three conserved cysteine residues occur at the dimer interface and likely form the catalytic site. To investigate the functions of these cysteines in the enzyme from Methanococcus vannielii, a series of biochemical studies were pursued to test the basic hypothesis regarding their roles in catalysis. Inactivation of the enzyme activity by methyl methane thiosulfonate (MMTS) or 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) also compromised the Zn(2+) binding properties of the protein inducing loss of up to 90% of the metal. Overall reaction stoichiometry and the potassium cyanide (KCN) induced cleavage of the protein suggested that all three cysteines were modified in the process. The enzyme was protected from DTNB-induced inactivation by inclusion of the substrate N(1)-(5'-phosphoribosyl)adenosine 5'-monophosphate; (PR-AMP), while Mg(2+), a metal required for catalytic activity, enhanced the rate of inactivation. Site-directed mutations of the conserved C93, C109, C116 and the double mutant C109/C116 were prepared and analyzed for catalytic activity, Zn(2+) content, and reactivity with DTNB. Substitution of alanine for each of the conserved cysteines showed no measurable catalytic activity, and only the C116A was still capable of binding Zn(2+). Reactions of DTNB with the C109A/C116A double mutant showed that C93 is completely modified within 0.5 s. A model consistent with these data involves a DTNB-induced mixed disulfide linkage between C93 and C109 or C116, followed by ejection of the active site Zn(2+) and provides further evidence that the Zn(2+) coordination site involves the three conserved cysteine residues. The C93 reactivity is modulated by the presence of the Zn(2+) and Mg(2+) and substantiates the role of this residue as a metal ligand. In addition, Mg(2+) ligand binding site(s) indicated by the structural analysis were probed by site-directed mutagenesis of three key aspartate residues flanking the conserved C93 which were shown to have a functional impact on catalysis, cysteine activation, and metal (zinc) binding capacity. The unique amino acid sequence, the dynamic properties of the cysteine ligands involved in Zn(2+) coordination, and the requirement for a second metal (Mg(2+)) are discussed in the context of their roles in catalysis. The results are consistent with a Zn(2+)-mediated activation of H(2)O mechanism involving histidine as a general base that has features similar to but distinct from those of previously characterized purine and pyrimidine deaminases.

Structural analysis of substrate recognition by glucose isomerase in Mn2+ binding mode at M2 site in S. rubiginosus.

PubMed

Bae, Ji-Eun; Hwang, Kwang Yeon; Nam, Ki Hyun

2018-06-16

Glucose isomerase (GI) catalyzes the reversible enzymatic isomerization of d-glucose and d-xylose to d-fructose and d-xylulose, respectively. This is one of the most important enzymes in the production of high-fructose corn syrup (HFCS) and biofuel. We recently determined the crystal structure of GI from S. rubiginosus (SruGI) complexed with a xylitol inhibitor in one metal binding mode. Although we assessed inhibitor binding at the M1 site, the metal binding at the M2 site and the substrate recognition mechanism for SruGI remains the unclear. Here, we report the crystal structure of the two metal binding modes of SruGI and its complex with glucose. This study provides a snapshot of metal binding at the SruGI M2 site in the presence of Mn 2+ , but not in the presence of Mg 2+ . Metal binding at the M2 site elicits a configuration change at the M1 site. Glucose molecule can only bind to the M1 site in presence of Mn 2+ at the M2 site. Glucose and Mn 2+ at the M2 site were bridged by water molecules using a hydrogen bonding network. The metal binding geometry of the M2 site indicates a distorted octahedral coordination with an angle of 55-110°, whereas the M1 site has a relatively stable octahedral coordination with an angle of 85-95°. We suggest a two-step sequential process for SruGI substrate recognition, in Mn 2+ binding mode, at the M2 site. Our results provide a better understanding of the molecular role of the M2 site in GI substrate recognition. Copyright © 2018. Published by Elsevier Inc.

Structure of the endonuclease IV homologue from Thermotoga maritima in the presence of active-site divalent metal ions

PubMed Central

Tomanicek, Stephen J.; Hughes, Ronny C.; Ng, Joseph D.; Coates, Leighton

2010-01-01

The most frequent lesion in DNA is at apurinic/apyrimidinic (AP) sites resulting from DNA-base losses. These AP-site lesions can stall DNA replication and lead to genome instability if left unrepaired. The AP endonucleases are an important class of enzymes that are involved in the repair of AP-site intermediates during damage-general DNA base-excision repair pathways. These enzymes hydrolytically cleave the 5′-phosphodiester bond at an AP site to generate a free 3′-­hydroxyl group and a 5′-terminal sugar phosphate using their AP nuclease activity. Specifically, Thermotoga maritima endonuclease IV is a member of the second conserved AP endonuclease family that includes Escherichia coli endonuclease IV, which is the archetype of the AP endonuclease superfamily. In order to more fully characterize the AP endonuclease family of enzymes, two X-­ray crystal structures of the T. maritima endonuclease IV homologue were determined in the presence of divalent metal ions bound in the active-site region. These structures of the T. maritima endonuclease IV homologue further revealed the use of the TIM-barrel fold and the trinuclear metal binding site as important highly conserved structural elements that are involved in DNA-binding and AP-site repair processes in the AP endonuclease superfamily. PMID:20823514

Isolation and characterization of iron chelators from turmeric (Curcuma longa): selective metal binding by curcuminoids.

PubMed

Messner, Donald J; Surrago, Christine; Fiordalisi, Celia; Chung, Wing Yin; Kowdley, Kris V

2017-10-01

Iron overload disorders may be treated by chelation therapy. This study describes a novel method for isolating iron chelators from complex mixtures including plant extracts. We demonstrate the one-step isolation of curcuminoids from turmeric, the medicinal food spice derived from Curcuma longa. The method uses iron-nitrilotriacetic acid (NTA)-agarose, to which curcumin binds rapidly, specifically, and reversibly. Curcumin, demethoxycurcumin, and bisdemethoxycurcumin each bound iron-NTA-agarose with comparable affinities and a stoichiometry near 1. Analyses of binding efficiencies and purity demonstrated that curcuminoids comprise the primary iron binding compounds recovered from a crude turmeric extract. Competition of curcuminoid binding to the iron resin was used to characterize the metal binding site on curcumin and to detect iron binding by added chelators. Curcumin-Iron-NTA-agarose binding was inhibited by other metals with relative potency: (>90% inhibition) Cu 2+  ~ Al 3+  > Zn 2+  ≥ Ca 2+  ~ Mg 2+  ~ Mn 2+ (<20% inhibition). Binding was also inhibited by pharmaceutical iron chelators (desferoxamine or EDTA) or by higher concentrations of weak iron chelators (citrate or silibinin). Investigation of the physiological effects of iron binding by curcumin revealed that curcumin uptake by cultured cells was reduced >80% by addition of iron to the media; uptake was completely restored by desferoxamine. Ranking of metals by relative potencies for blocking curcumin uptake agreed with their relative potencies in blocking curcumin binding to iron-NTA-agarose. We conclude that curcumin can selectively bind toxic metals including iron in a physiological setting, and propose inhibition of curcumin binding to iron-NTA-agarose for iron chelator screening.

Computational scheme for the prediction of metal ion binding by a soil fulvic acid

USGS Publications Warehouse

Marinsky, J.A.; Reddy, M.M.; Ephraim, J.H.; Mathuthu, A.S.

1995-01-01

The dissociation and metal ion binding properties of a soil fulvic acid have been characterized. Information thus gained was used to compensate for salt and site heterogeneity effects in metal ion complexation by the fulvic acid. An earlier computational scheme has been modified by incorporating an additional step which improves the accuracy of metal ion speciation estimates. An algorithm is employed for the prediction of metal ion binding by organic acid constituents of natural waters (once the organic acid is characterized in terms of functional group identity and abundance). The approach discussed here, currently used with a spreadsheet program on a personal computer, is conceptually envisaged to be compatible with computer programs available for ion binding by inorganic ligands in natural waters.

Articles including thin film monolayers and multilayers

DOEpatents

Li, DeQuan; Swanson, Basil I.

1995-01-01

Articles of manufacture including: (a) a base substrate having an oxide surface layer, and a multidentate ligand, capable of binding a metal ion, attached to the oxide surface layer of the base substrate, (b) a base substrate having an oxide surface layer, a multidentate ligand, capable of binding a metal ion, attached to the oxide surface layer of the base substrate, and a metal species attached to the multidentate ligand, (c) a base substrate having an oxide surface layer, a multidentate ligand, capable of binding a metal ion, attached to the oxide surface layer of the base substrate, a metal species attached to the multidentate ligand, and a multifunctional organic ligand attached to the metal species, and (d) a base substrate having an oxide surface layer, a multidentate ligand, capable of binding a metal ion, attached to the oxide surface layer of the base substrate, a metal species attached to the multidentate ligand, a multifunctional organic ligand attached to the metal species, and a second metal species attached to the multifunctional organic ligand, are provided, such articles useful in detecting the presence of a selected target species, as nonliear optical materials, or as scavengers for selected target species.

Method of making maximally dispersed heterogeneous catalysts

DOEpatents

Jennison, Dwight R [Albuquerque, NM

2005-11-15

A method of making a catalyst with monolayer or sub-monolayer metal by controlling the wetting characteristics on the support surface and increasing the adhesion between the catalytic metal and an oxide layer. There are two methods that have been demonstrated by experiment and supported by theory. In the first method, which is useful for noble metals as well as others, a negatively-charged species is introduced to the surface of a support in sub-ML coverage. The layer-by-layer growth of metal deposited onto the oxide surface is promoted because the adhesion strength of the metal-oxide interface is increased. This method can also be used to achieve nanoislands of metal upon sub-ML deposition. The negatively-charged species can either be deposited onto the oxide surface or a compound can be deposited that dissociates on, or reacts with, the surface to form the negatively-charged species. The deposited metal adatoms can thereby bond laterally to the negatively-charged species as well as vertically to the oxide surface. Thus the negatively-charged species serve as anchors for the metal. In the second method, a chemical reaction that occurs when most metals are deposited on a fully hydroxylated oxide surface is used to create cationic metal species that bind strongly both to the substrate and to metallic metal atoms. These are incorporated into the top layer of the substrate and bind strongly both to the substrate and to metallic metal atoms. In this case, these oxidized metal atoms serve as the anchors. Here, as in the previous method, nanoislands of catalytic metal can be achieved to increase catalytic activity, or monolayers or bilayers of reactive metal can also be made.

SPECT/CT localization of oral radioiodine activity: a retrospective study and in-vitro assessment

PubMed Central

Burlison, Jared S.; Hartshorne, Michael F.; Voda, Alan M.; Cocks, Franklin H.

2013-01-01

Purpose We sought to further localize radioiodine activity in the mouth on post-thyroid cancer therapy imaging using single-photon emission computed tomography/computed tomography (SPECT/CT). Materials and methods We retrospectively reviewed all patients (58) who underwent thyroid cancer therapy with iodine-131 (131I) at our institution from August 2009 to March 2011 whose post-therapy radioiodine imaging included neck SPECT/CT. A small group (six) of diagnostic 123I scans including SPECT/CT was also reviewed. Separately, we performed in-vitro 131I (sodium iodide) binding assays with amalgam and Argenco HP 77 (77% dental gold alloy) as proof of principle for these interactions. Results Of the 58 post-therapy patients, 45 (78%) had undergone metallic dental restorations, and of them 41 (91%) demonstrated oral 131I activity localizing preferentially to those restorations. It was observed that radioiodine also localized to other dental restorations and to orthodontic hardware. Gum-line activity in edentulous patients suggests radioiodine interaction with denture adhesive. In vitro, dental amalgam and Argenco HP 77 bound 131I in a time-dependent manner over 1–16 days of exposure. Despite subsequent washings with normal saline, significant 131I activity (maximally 12% for amalgam and 68% for Argenco HP 77) was retained by these metals. Subsequent soaking in a saturated solution of potassium iodide partially displaced 131I from amalgam, with near-total displacement of 131I from Argenco HP 77. Conclusion SPECT/CT shows that radioiodine in the oral cavity localizes to metallic dental restorations. Furthermore, in-vitro studies demonstrate partially reversible binding of 131I to common dental metals. PMID:24128897

Proton NMR spectroscopic characterization of binary and ternary complexes of cobalt(II) carboxypeptidase A with inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bertini, I.; Luchinat, C.; Messori, L.

The binding of L- and D-phenylalanine and carboxylate inhibitors to cobalt(II)-substituted carboxypeptidase A, Co(II)CPD (E), in the presence and absence of pseudohalogens (X = N/sub 3//sup -/, NCO/sup -/, and NCS/sup -/) has been studied by /sup 1/H NMR spectroscopy. This technique monitors the proton signals of histidine residues bound to cobalt(II) and is therefore sensitive to the interactions of inhibitors that perturb the coordination sphere of the metal. Enzyme-inhibitor complexes, E/times/I, E/times/I/sub 2/, and E/times/I/times/X, each with characteristic NMR features, have been identified. The NMR data suggest that when the carboxylate group of a substrate of inhibitor binds atmore » the active site, a conformational change occurs that allows a second ligand molecule to bind to the metal ion, altering its coordination sphere and thereby attenuating the bidentate behavior of Glu-72. The /sup 1/H NMR signals also reflect alterations in the histidine interactions with the metal upon inhibitor binding. Isotropic shifts in the signals for the C-4 (c) and N protons (a) of one of the histidine ligands are readily observed in all of these complexes. These signals are relatively constant for all E/times/I and E/times/I/times/X complexes, indicating that this ligand is in a relatively fixed or buried conformation. However in the 2:1 carboxylate inhibitor (E/times/I/sub 2/) complexes, both signals are shifted upfield, suggesting a disturbance in the interaction of this histidine with the metal.« less

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence.

PubMed

Haley, Kathryn P; Delgado, Alberto G; Piazuelo, M Blanca; Mortensen, Brittany L; Correa, Pelayo; Damo, Steven M; Chazin, Walter J; Skaar, Eric P; Gaddy, Jennifer A

2015-07-01

During infectious processes, antimicrobial proteins are produced by both epithelial cells and innate immune cells. Some of these antimicrobial molecules function by targeting transition metals and sequestering these metals in a process referred to as "nutritional immunity." This chelation strategy ultimately starves invading pathogens, limiting their growth within the vertebrate host. Recent evidence suggests that these metal-binding antimicrobial molecules have the capacity to affect bacterial virulence, including toxin secretion systems. Our previous work showed that the S100A8/S100A9 heterodimer (calprotectin, or calgranulin A/B) binds zinc and represses the elaboration of the H. pylori cag type IV secretion system (T4SS). However, there are several other S100 proteins that are produced in response to infection. We hypothesized that the zinc-binding protein S100A12 (calgranulin C) is induced in response to H. pylori infection and also plays a role in controlling H. pylori growth and virulence. To test this, we analyzed gastric biopsy specimens from H. pylori-positive and -negative patients for S100A12 expression. These assays showed that S100A12 is induced in response to H. pylori infection and inhibits bacterial growth and viability in vitro by binding nutrient zinc. Furthermore, the data establish that the zinc-binding activity of the S100A12 protein represses the activity of the cag T4SS, as evidenced by the gastric cell "hummingbird" phenotype, interleukin 8 (IL-8) secretion, and CagA translocation assays. In addition, high-resolution field emission gun scanning electron microscopy (FEG-SEM) was used to demonstrate that S100A12 represses biogenesis of the cag T4SS. Together with our previous work, these data reveal that multiple S100 proteins can repress the elaboration of an oncogenic bacterial surface organelle. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Synthesis, characterization and DNA-binding studies of mono and heterobimetallic complexes Cu sbnd Sn 2/Zn sbnd Sn 2 and their DNA cleavage activity

NASA Astrophysics Data System (ADS)

Arjmand, Farukh; Sayeed, Fatima

2010-02-01

Heterobimetallic complexes C 6H 24N 4O 6CuSn 2Cl 63, C 6H 24N 4O 6ZnSn 2Cl 64 have been synthesized from their monometallic analogs C 6H 16N 4O 2CuCl 21, C 6H 16N 4O 2ZnCl 22, and were characterized by various spectroscopic and analytical methods. The complexes 1-4 reveal an octahedral geometry for both central metal ions Cu/Zn as well as for Sn metal ion. The interaction of complexes 1-4 with CT-DNA, were investigated by using absorption, emission, cyclic voltammetry, viscometry and DNA cleavage studies. The emission quenching of 3 and 4 by [Fe(CN) 6] 4- depressed greatly when bound to CT-DNA. The results of spectroscopic, viscometric and cyclic voltammetry of complexes 3 and 4 revealed electrostatic mode of binding of the complexes with CT-DNA. These results revealed that 4 bind more avidly in comparison to 3 with CT-DNA. Gel electrophoresis of DNA with complexes 3 and 4 demonstrated that the complexes exhibit excellent cleavage activity under physiological conditions.

A novel heavy metal ATPase peptide from Prosopis juliflora is involved in metal uptake in yeast and tobacco.

PubMed

Keeran, Nisha S; Ganesan, G; Parida, Ajay K

2017-04-01

Heavy metal pollution of agricultural soils is one of the most severe ecological problems in the world. Prosopis juliflora, a phreatophytic tree species, grows well in heavy metal laden industrial sites and is known to accumulate heavy metals. Heavy Metal ATPases (HMAs) are ATP driven heavy metal pumps that translocate heavy metals across biological membranes thus helping the plant in heavy metal tolerance and phytoremediation. In the present study we have isolated and characterized a novel 28.9 kDa heavy metal ATPase peptide (PjHMT) from P. juliflora which shows high similarity to the C-terminal region of P 1B ATPase HMA1. It also shows the absence of the invariant signature sequence DKTGT, and the metal binding CPX motif but the presence of conserved regions like MVGEGINDAPAL (ATP binding consensus sequence), HEGGTLLVCLNS (metal binding domain) and MLTGD, GEGIND and HEGG motifs which play important roles in metal transport or ATP binding. PjHMT, was found to be upregulated under cadmium and zinc stress. Heterologous expression of PjHMT in yeast showed a higher accumulation and tolerance of heavy metals in yeast. Further, transgenic tobacco plants constitutively expressing PjHMT also showed increased accumulation and tolerance to cadmium. Thus, this study suggests that the transport peptide from P. juliflora may have an important role in Cd uptake and thus in phytoremediation.

Revealing the role of the product metal in DNA polymerase β catalysis

PubMed Central

Freudenthal, Bret D.; Beard, William A.; Pedersen, Lee G.; Wilson, Samuel H.

2017-01-01

Abstract DNA polymerases catalyze a metal-dependent nucleotidyl transferase reaction during extension of a DNA strand using the complementary strand as a template. The reaction has long been considered to require two magnesium ions. Recently, a third active site magnesium ion was identified in some DNA polymerase product crystallographic structures, but its role is not known. Using quantum mechanical/ molecular mechanical calculations of polymerase β, we find that a third magnesium ion positioned near the newly identified product metal site does not alter the activation barrier for the chemical reaction indicating that it does not have a role in the forward reaction. This is consistent with time-lapse crystallographic structures following insertion of Sp-dCTPαS. Although sulfur substitution deters product metal binding, this has only a minimal effect on the rate of the forward reaction. Surprisingly, monovalent sodium or ammonium ions, positioned in the product metal site, lowered the activation barrier. These calculations highlight the impact that an active site water network can have on the energetics of the forward reaction and how metals or enzyme side chains may interact with the network to modulate the reaction barrier. These results also are discussed in the context of earlier findings indicating that magnesium at the product metal position blocks the reverse pyrophosphorolysis reaction. PMID:28108654

A dye-binding assay for measurement of the binding of Cu(II) to proteins.

PubMed

Wilkinson-White, Lorna E; Easterbrook-Smith, Simon B

2008-10-01

We analysed the theory of the coupled equilibria between a metal ion, a metal ion-binding dye and a metal ion-binding protein in order to develop a procedure for estimating the apparent affinity constant of a metal ion:protein complex. This can be done by analysing from measurements of the change in the concentration of the metal ion:dye complex with variation in the concentration of either the metal ion or the protein. Using experimentally determined values for the affinity constant of Cu(II) for the dye, 2-(5-bromo-2-pyridylaxo)-5-(N-propyl-N-sulfopropylamino) aniline (5-Br-PSAA), this procedure was used to estimate the apparent affinity constants for formation of Cu(II):transthyretin, yielding values which were in agreement with literature values. An apparent affinity constant for Cu(II) binding to alpha-synuclein of approximately 1 x 10(9)M(-1) was obtained from measurements of tyrosine fluorescence quenching by Cu(II). This value was in good agreement with that obtained using 5-Br-PSAA. Our analysis and data therefore show that measurement of changes in the equilibria between Cu(II) and 5-Br-PSAA by Cu(II)-binding proteins provides a general procedure for estimating the affinities of proteins for Cu(II).

Biotic ligand modeling approach: Synthesis of the effect of major cations on the toxicity of metals to soil and aquatic organisms.

PubMed

Ardestani, Masoud M; van Straalen, Nico M; van Gestel, Cornelis A M

2015-10-01

The biotic ligand model (BLM) approach is used to assess metal toxicity, taking into account the competition of other cations with the free metal ions for binding to the biotic ligand sites of aquatic and soil organisms. The bioavailable fraction of metals, represented by the free metal ion, is a better measure than the total concentration for assessing their potential risk to the environment. Because BLMs are relating toxicity to the fraction of biotic ligands occupied by the metal, they can be useful for investigating factors affecting metal bioaccumulation and toxicity. In the present review, the effects of major cations on the toxicity of metals to soil and aquatic organisms were comprehensively studied by performing a meta-analysis of BLM literature data. Interactions at the binding sites were shown to be species- and metal-specific. The main factors affecting the relationships between toxicity and conditional binding constants for metal binding at the biotic ligand appeared to be Ca(2+) , Mg(2+) , and protons. Other important characteristics of the exposure medium, such as levels of dissolved organic carbon and concentrations of other cations, should also be considered to obtain a proper assessment of metal toxicity to soil and aquatic organisms. © 2015 SETAC.

Engineered Mononuclear Variants in Bacillus cereus Metallo-β-lactamase BcII Are Inactive†

PubMed Central

Abriata, Luciano A.; González, Lisandro J.; Llarrull, Leticia I.; Tomatis, Pablo E.; Myers, William K.; Costello, Alison L.; Tierney, David L.; Vila, Alejandro J.

2008-01-01

Metallo-β-lactamases (MβLs) are zinc enzymes able to hydrolyze almost all β-lactam antibiotics, rendering them inactive, at the same time endowing bacteria high levels of resistance. The design of inhibitors active against all classes of MβLs has been hampered by their structural diversity and by the heterogeneity in metal content in enzymes from different sources. BcII is the metallo-β-lactamase from Bacillus cereus, which is found in both the mononuclear and dinuclear forms. Despite extensive studies, there is still controversy about the nature of the active BcII species. Here we have designed two mutant enzymes in which each one of the metal binding sites was selectively removed. Both mutants were almost inactive, despite preserving most of the structural features of each metal site. These results reveal that neither site isolated in the MβL scaffold is sufficient to render a fully active enzyme. This suggests that only the dinuclear species is active or that the mononuclear variants can be active only if aided by other residues that would be metal ligands in the dinuclear species. PMID:18652482

Development of an iron chelating polyethylene film for active packaging applications.

PubMed

Tian, Fang; Decker, Eric A; Goddard, Julie M

2012-02-29

Metal-promoted oxidation reactions are a major cause of food quality deterioration. Active packaging offers novel approaches to controlling such oxidation for the purpose of extending shelf life. Herein, we report modification of the surface of polyethylene (PE) films to possess metal chelating activity. Metal chelating carboxylic acids were introduced to the film surface using cross-linking agents [polyethylenimine (PEI) or ethylenediamine (ED)] to increase the number of available carboxylic acids. ATR-FTIR, contact angle, dye assay, and iron chelating assay were used to characterize changes in surface chemistry after each functionalization step. The chelator poly(acrylic acid) (PAA) was attached to the surface at a density of 9.12 ± 0.71 nmol carboxyl groups/cm², and exhibited an iron chelating activity. The results indicate that PAA-modified PE films might have a higher affinity to Fe³⁺ than Fe²⁺ with the optimum binding pH at 5.0. Such inexpensive active packaging materials are promising in food industry for the preservation of liquid and semiliquid food products and have application in heavy metal chelation therapy for biomedical materials as well.

The Study of the Successive Metal-ligand Binding Energies for Fe(+), Fe(-), V(+) and Co(+)

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Ricca, Alessandra; Maitre, Philippe; Langhoff, Stephen R. (Technical Monitor)

1994-01-01

The successive binding energies of CO and H2O to Fe(+), CO to Fe(-), and H2 to Co(+) and V(+) are presented. Overall the computed results are in good agreement with experiment. The trends in binding energies are analyzed in terms of metal to ligand donation, ligand to metal donation, ligand-ligand repulsion, and changes in the metal atom, such as hybridization, promotion, and spin multiplicity. The geometry and vibrational frequencies are also shown to be directly affected by these effects.

The Study Of The Successive Metal-Ligand Binding Energies For Fe+, Fe-, V+ and Co+

NASA Technical Reports Server (NTRS)

Bauschicher, Charles W., Jr.; Ricca, Alessandra; Maitre, Philippe; Langhoff, Stephen R. (Technical Monitor)

1995-01-01

The successive binding energies of CO and H2O to Fe(+), CO to Fe(-), and H2 to Co(+) and V(+) are presented. Overall the computed results are in good agreement with experiment. The trends in binding energies are analyzed in terms of metal to ligand donation, ligand to metal donation, ligand-ligand repulsion, and changes in the metal atom, such as hybridization, promotion, and spin multiplicity. The geometry and vibrational frequencies are also shown to be directly affected by these effects.

Manipulation and measurement of pH sensitive metal-ligand binding using electrochemical proton generation and metal detection.

PubMed

Read, Tania L; Joseph, Maxim B; Macpherson, Julie V

2016-01-31

Generator-detector electrodes can be used to both perturb and monitor pH dependant metal-ligand binding equilibria, in situ. In particular, protons generated at the generator locally influence the speciation of metal (Cu(2+)) in the presence of ligand (triethylenetetraamine), with the detector employed to monitor, in real time, free metal (Cu(2+)) concentrations.

Biosorption of heavy metal ions on Rhodobacter sphaeroides and Alcaligenes eutrophus H16

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seki, Hideshi; Suzuki, Akira; Mitsueda, Shinichiro

1998-01-15

A fundamental study of the application of bacteria to the recovery of toxic heavy metals from aqueous environments was carried out. The biosorption characteristics of cadmium and lead ions were determined with purple nonsulfur bacteria, Rhodobacter sphaeroides and hydrogen bacteria, Alcaligenes eutrophus H16 that were inactivated by steam sterilization. A simplified version of the metal binding model proposed by Plette et al. was used for the description of meal binding data. The results showed that the biosorption of bivalent metal ions to whole cell bodies of the bacteria was due to monodentate binding to two different types of acidic sites:more » carboxilic and phosphatic-type sites. The number of metal binding sites of A. eutrophus was 2.4-fold larger than that of R. sphaeroides.« less

Identification of a Pyridoxine-Derived Small-Molecule Inhibitor Targeting Dengue Virus RNA-Dependent RNA Polymerase.

PubMed

Xu, Hong-Tao; Colby-Germinario, Susan P; Hassounah, Said; Quashie, Peter K; Han, Yingshan; Oliveira, Maureen; Stranix, Brent R; Wainberg, Mark A

2016-01-01

The viral RNA-dependent RNA polymerase (RdRp) activity of the dengue virus (DENV) NS5 protein is an attractive target for drug design. Here, we report the identification of a novel class of inhibitor (i.e., an active-site metal ion chelator) that acts against DENV RdRp activity. DENV RdRp utilizes a two-metal-ion mechanism of catalysis; therefore, we constructed a small library of compounds, through mechanism-based drug design, aimed at chelating divalent metal ions in the catalytic site of DENV RdRp. We now describe a pyridoxine-derived small-molecule inhibitor that targets DENV RdRp and show that 5-benzenesulfonylmethyl-3-hydroxy-4-hydroxymethyl-pyridine-2-carboxylic acid hydroxyamide (termed DMB220) inhibited the RdRp activity of DENV serotypes 1 to 4 at low micromolar 50% inhibitory concentrations (IC50s of 5 to 6.7 μM) in an enzymatic assay. The antiviral activity of DMB220 against DENV infection was also verified in a cell-based assay and showed a 50% effective concentration (EC50) of <3 μM. Enzyme assays proved that DMB220 was competitive with nucleotide incorporation. DMB220 did not inhibit the enzymatic activity of recombinant HIV-1 reverse transcriptase and showed only weak inhibition of HIV-1 integrase strand transfer activity, indicating high specificity for DENV RdRp. S600T substitution in the DENV RdRp, which was previously shown to confer resistance to nucleoside analogue inhibitors (NI), conferred 3-fold hypersusceptibility to DMB220, and enzymatic analyses showed that this hypersusceptibility may arise from the decreased binding/incorporation efficiency of the natural NTP substrate without significantly impacting inhibitor binding. Thus, metal ion chelation at the active site of DENV RdRp represents a viable anti-DENV strategy, and DMB220 is the first of a new class of DENV inhibitor. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Simultaneous determination of Ca, Cu, Ni, Zn and Cd binding strengths with fulvic acid fractions by Schubert's method

USGS Publications Warehouse

Brown, G.K.; MacCarthy, P.; Leenheer, J.A.

1999-01-01

The equilibrium binding of Ca2+, Ni2+, Cd2+, Cu2+ and Zn2+ with unfractionated Suwannee river fulvic acid (SRFA) and an enhanced metal binding subfraction of SRFA was measured using Schubert's ion-exchange method at pH 6.0 and at an ionic strength (??) of 0.1 (NaNO3). The fractionation and subfractionation were directed towards obtaining an isolate with an elevated metal binding capacity or binding strength as estimated by Cu2+ potentiometry (ISE). Fractions were obtained by stepwise eluting an XAD-8 column loaded with SRFA with water eluents of pH 1.0 to pH 12.0. Subfractions were obtained by loading the fraction eluted from XAD-8 at pH 5.0 onto a silica gel column and eluting with solvents of increasing polarity. Schuberts ion exchange method was rigorously tested by measuring simultaneously the conditional stability constants (K) of citric acid complexed with the five metals at pH 3.5 and 6.0. The logK of SRFA with Ca2+, Ni2+, Cd2+, Cu2+ and Zn2+ determined simultaneously at pH 6.0 follow the sequence of Cu2+>Cd2+>Ni2+>Zn2+>Ca2+ while all logK values increased for the enhanced metal binding subfraction and followed a different sequence of Cu2+>Cd2+>Ca2+>Ni2+>Zn2+. Both fulvic acid samples and citric acid exhibited a 1:1 metal to ligand stochiometry under the relatively low metal loading conditions used here. Quantitative 13C nuclear magnetic resonance spectroscopy showed increases in aromaticity and ketone content and decreases in aliphatic carbon for the elevated metal binding fraction while the carboxyl carbon, and elemental nitrogen, phosphorus, and sulfur content did not change. The more polar, elevated metal binding fraction did show a significant increase in molecular weight over the unfractionated SRFA. Copyright (C) 1999 Elsevier Science B.V.

Metal Preferences and Metallation*

PubMed Central

Foster, Andrew W.; Osman, Deenah; Robinson, Nigel J.

2014-01-01

The metal binding preferences of most metalloproteins do not match their metal requirements. Thus, metallation of an estimated 30% of metalloenzymes is aided by metal delivery systems, with ∼25% acquiring preassembled metal cofactors. The remaining ∼70% are presumed to compete for metals from buffered metal pools. Metallation is further aided by maintaining the relative concentrations of these pools as an inverse function of the stabilities of the respective metal complexes. For example, magnesium enzymes always prefer to bind zinc, and these metals dominate the metalloenzymes without metal delivery systems. Therefore, the buffered concentration of zinc is held at least a million-fold below magnesium inside most cells. PMID:25160626

Electron Spin Resonance Studies of Carbonic Anhydrase: Transition Metal Ions and Spin-Labeled Sulfonamides*

PubMed Central

Taylor, June S.; Mushak, Paul; Coleman, Joseph E.

1970-01-01

Electron spin resonance (esr) spectra of Cu(II) and Co(II) carbonic anhydrase, and a spin-labeled sulfonamide complex of the Zn(II) enzyme, are reported. The coordination geometry of Cu(II) bound in the enzyme appears to have approximately axial symmetry. Esr spectra of enzyme complexes with metal-binding anions also show axial symmetry and greater covalency, in the order ethoxzolamide < SH- < N3- ≤ CN-. Well-resolved superhyperfine structure in the spectrum of the cyanide complex suggests the presence of two, and probably three, equivalent nitrogen ligands from the protein. Esr spectra of the Co(II) enzyme and its complexes show two types of Co(II) environment, one typical of the native enzyme and the 1:1 CN- complex, and one typical of a 2:1 CN- complex. Co(II) in the 2:1 complex appears to be low-spin and probably has a coordination number of 5. Binding of a spin-labeled sulfonamide to the active center immobilizes the free radical. The similarity of the esr spectra of spin-labeled Zn(II) and Co(II) carbonic anhydrases suggests that the conformation at the active center is similar in the two metal derivatives. PMID:4320976

Water-soluble Manganese and Iron Mesotetrakis(carboxyl)porphyrin: DNA Binding, Oxidative Cleavage, and Cytotoxic Activities.

PubMed

Shi, Lei; Jiang, Yi-Yu; Jiang, Tao; Yin, Wei; Yang, Jian-Ping; Cao, Man-Li; Fang, Yu-Qi; Liu, Hai-Yang

2017-06-29

Two new water-soluble metal carboxyl porphyrins, manganese (III) meso -tetrakis (carboxyl) porphyrin and iron (III) meso -tetrakis (carboxyl) porphyrin, were synthesized and characterized. Their interactions with ct-DNA were investigated by UV-Vis titration, fluorescence spectra, viscosity measurement and CD spectra. The results showed they can strongly bind to ct-DNA via outside binding mode. Electrophoresis experiments revealed that both complexes can cleave pBR322 DNA efficiently in the presence of hydrogen peroxide, albeit 2-Mn exhibited a little higher efficiency. The inhibitor tests suggest the oxidative DNA cleavage by these two complexes may involve hydroxyl radical active intermediates. Notably, 2-Mn exhibited considerable photocytotoxicity against Hep G2 cell via triggering a significant generation of ROS and causing disruption of MMP after irradiation.

A VARIABLE REACTIVITY MODEL FOR ION BINDING TO ENVIRONMENTAL SORBENTS

EPA Science Inventory

The conceptual and mathematical basis for a new general-composite modeling approach for ion binding to environmental sorbents is presented. The work extends the Simple Metal Sorption (SiMS) model previously presented for metal and proton binding to humic substances. A surface com...

Phosphate forms an unusual tripodal complex with the Fe–Mn center of sweet potato purple acid phosphatase

PubMed Central

Schenk, Gerhard; Gahan, Lawrence R.; Carrington, Lyle E.; Mitić, Nataša; Valizadeh, Mohsen; Hamilton, Susan E.; de Jersey, John; Guddat, Luke W.

2005-01-01

Purple acid phosphatases (PAPs) are a family of binuclear metalloenzymes that catalyze the hydrolysis of phosphoric acid esters and anhydrides. A PAP in sweet potato has a unique, strongly antiferromagnetically coupled Fe(III)–Mn(II) center and is distinguished from other PAPs by its increased catalytic efficiency for a range of activated and unactivated phosphate esters, its strict requirement for Mn(II), and the presence of a μ-oxo bridge at pH 4.90. This enzyme displays maximum catalytic efficiency (kcat/Km) at pH 4.5, whereas its catalytic rate constant (kcat) is maximal at near-neutral pH, and, in contrast to other PAPs, its catalytic parameters are not dependent on the pKa of the leaving group. The crystal structure of the phosphate-bound Fe(III)–Mn(II) PAP has been determined to 2.5-Å resolution (final Rfree value of 0.256). Structural comparisons of the active site of sweet potato, red kidney bean, and mammalian PAPs show several amino acid substitutions in the sweet potato enzyme that can account for its increased catalytic efficiency. The phosphate molecule binds in an unusual tripodal mode to the two metal ions, with two of the phosphate oxygen atoms binding to Fe(III) and Mn(II), a third oxygen atom bridging the two metal ions, and the fourth oxygen pointing toward the substrate binding pocket. This binding mode is unique among the known structures in this family but is reminiscent of phosphate binding to urease and of sulfate binding to λ protein phosphatase. The structure and kinetics support the hypothesis that the bridging oxygen atom initiates hydrolysis. PMID:15625111

Zn(II) and Hg(II) binding to a designed peptide that accommodates different coordination geometries.

PubMed

Szunyogh, Dániel; Gyurcsik, Béla; Larsen, Flemming H; Stachura, Monika; Thulstrup, Peter W; Hemmingsen, Lars; Jancsó, Attila

2015-07-28

Designed metal ion binding peptides offer a variety of applications in both basic science as model systems of more complex metalloproteins, and in biotechnology, e.g. in bioremediation of toxic metal ions, biomining or as artificial enzymes. In this work a peptide (HS: Ac-SCHGDQGSDCSI-NH2) has been specifically designed for binding of both Zn(II) and Hg(II), i.e. metal ions with different preferences in terms of coordination number, coordination geometry, and to some extent ligand composition. It is demonstrated that HS accommodates both metal ions, and the first coordination sphere, metal ion exchange between peptides, and speciation are characterized as a function of pH using UV-absorption-, synchrotron radiation CD-, (1)H-NMR-, and PAC-spectroscopy as well as potentiometry. Hg(II) binds to the peptide with very high affinity in a {HgS2} coordination geometry, bringing together the two cysteinates close to each end of the peptide in a loop structure. Despite the high affinity, Hg(II) is kinetically labile, exchanging between peptides on the subsecond timescale, as indicated by line broadening in (1)H-NMR. The Zn(II)-HS system displays more complex speciation, involving monomeric species with coordinating cysteinates, histidine, and a solvent water molecule, as well as HS-Zn(II)-HS complexes. In summary, the HS peptide displays conformational flexibility, contains many typical metal ion binding groups, and is able to accommodate metal ions with different structural and ligand preferences with high affinity. As such, the HS peptide may be a scaffold offering binding of a variety of metal ions, and potentially serve for metal ion sequestration in biotechnological applications.

Methanobactin and the Link between Copper and Bacterial Methane Oxidation

PubMed Central

Semrau, Jeremy D.; Murrell, J. Colin; Gallagher, Warren H.; Dennison, Christopher; Vuilleumier, Stéphane

2016-01-01

SUMMARY Methanobactins (mbs) are low-molecular-mass (<1,200 Da) copper-binding peptides, or chalkophores, produced by many methane-oxidizing bacteria (methanotrophs). These molecules exhibit similarities to certain iron-binding siderophores but are expressed and secreted in response to copper limitation. Structurally, mbs are characterized by a pair of heterocyclic rings with associated thioamide groups that form the copper coordination site. One of the rings is always an oxazolone and the second ring an oxazolone, an imidazolone, or a pyrazinedione moiety. The mb molecule originates from a peptide precursor that undergoes a series of posttranslational modifications, including (i) ring formation, (ii) cleavage of a leader peptide sequence, and (iii) in some cases, addition of a sulfate group. Functionally, mbs represent the extracellular component of a copper acquisition system. Consistent with this role in copper acquisition, mbs have a high affinity for copper ions. Following binding, mbs rapidly reduce Cu2+ to Cu1+. In addition to binding copper, mbs will bind most transition metals and near-transition metals and protect the host methanotroph as well as other bacteria from toxic metals. Several other physiological functions have been assigned to mbs, based primarily on their redox and metal-binding properties. In this review, we examine the current state of knowledge of this novel type of metal-binding peptide. We also explore its potential applications, how mbs may alter the bioavailability of multiple metals, and the many roles mbs may play in the physiology of methanotrophs. PMID:26984926

Nitric oxide activation by distal redox modulation in tetranuclear iron nitrosyl complexes.

PubMed

de Ruiter, Graham; Thompson, Niklas B; Lionetti, Davide; Agapie, Theodor

2015-11-11

A series of tetranuclear iron complexes displaying a site-differentiated metal center was synthesized. Three of the metal centers are coordinated to our previously reported ligand, based on a 1,3,5-triarylbenzene motif with nitrogen and oxygen donors. The fourth (apical) iron center is coordinatively unsaturated and appended to the trinuclear core through three bridging pyrazolates and an interstitial μ4-oxide moiety. Electrochemical studies of complex [LFe3(PhPz)3OFe][OTf]2 revealed three reversible redox events assigned to the Fe(II)4/Fe(II)3Fe(III) (-1.733 V), Fe(II)3Fe(III)/Fe(II)2Fe(III)2 (-0.727 V), and Fe(II)2Fe(III)2/Fe(II)Fe(III)3 (0.018 V) redox couples. Combined Mössbauer and crystallographic studies indicate that the change in oxidation state is exclusively localized at the triiron core, without changing the oxidation state of the apical metal center. This phenomenon is assigned to differences in the coordination environment of the two metal sites and provides a strategy for storing electron and hole equivalents without affecting the oxidation state of the coordinatively unsaturated metal. The presence of a ligand-binding site allowed the effect of redox modulation on nitric oxide activation by an Fe(II) metal center to be studied. Treatment of the clusters with nitric oxide resulted in binding of NO to the apical iron center, generating a {FeNO}(7) moiety. As with the NO-free precursors, the three reversible redox events are localized at the iron centers distal from the NO ligand. Altering the redox state of the triiron core resulted in significant change in the NO stretching frequency, by as much as 100 cm(-1). The increased activation of NO is attributed to structural changes within the clusters, in particular, those related to the interaction of the metal centers with the interstitial atom. The differences in NO activation were further shown to lead to differential reactivity, with NO disproportionation and N2O formation performed by the more electron-rich cluster.

Competition effects in cation binding to humic acid: Conditional affinity spectra for fixed total metal concentration conditions

NASA Astrophysics Data System (ADS)

David, Calin; Mongin, Sandrine; Rey-Castro, Carlos; Galceran, Josep; Companys, Encarnació; Garcés, José Luis; Salvador, José; Puy, Jaume; Cecilia, Joan; Lodeiro, Pablo; Mas, Francesc

2010-09-01

Information on the Pb and Cd binding to a purified Aldrich humic acid (HA) is obtained from the influence of different fixed total metal concentrations on the acid-base titrations of this ligand. NICA (Non-Ideal Competitive Adsorption) isotherm has been used for a global quantitative description of the binding, which has then been interpreted by plotting the Conditional Affinity Spectra of the H + binding at fixed total metal concentrations (CAScTM). This new physicochemical tool, here introduced, allows the interpretation of binding results in terms of distributions of proton binding energies. A large increase in the acidity of the phenolic sites as the total metal concentration increases, especially in presence of Pb, is revealed from the shift of the CAScTM towards lower affinities. The variance of the CAScTM distribution, which can be used as a direct measure of the heterogeneity, also shows a significant dependence on the total metal concentration. A discussion of the factors that influence the heterogeneity of the HA under the conditions of each experiment is provided, so that the smoothed pattern exhibited by the titration curves can be justified.

Generic NICA-Donnan model parameters for metal-ion binding by humic substances.

PubMed

Milne, Christopher J; Kinniburgh, David G; van Riemsdijk, Willem H; Tipping, Edward

2003-03-01

A total of 171 datasets of literature and experimental data for metal-ion binding by fulvic and humic acids have been digitized and re-analyzed using the NICA-Donnan model. Generic parameter values have been derived that can be used for modeling in the absence of specific metalion binding measurements. These values complement the previously derived generic descriptions of proton binding. For ions where the ranges of pH, concentration, and ionic strength conditions are well covered by the available data,the generic parameters successfully describe the metalion binding behavior across a very wide range of conditions and for different humic and fulvic acids. Where published data for other metal ions are too sparse to constrain the model well, generic parameters have been estimated by interpolating trends observable in the parameter values of the well-defined data. Recommended generic NICA-Donnan model parameters are provided for 23 metal ions (Al, Am, Ba, Ca, Cd, Cm, Co, CrIII, Cu, Dy, Eu, FeII, FeIII, Hg, Mg, Mn, Ni, Pb, Sr, Thv, UVIO2, VIIIO, and Zn) for both fulvic and humic acids. These parameters probably represent the best NICA-Donnan description of metal-ion binding that can be achieved using existing data.

Cytotoxicity and DNA cleavage with core-shell nanocomposites functionalized by a KH domain DNA binding peptide

NASA Astrophysics Data System (ADS)

Bazak, Remon; Ressl, Jan; Raha, Sumita; Doty, Caroline; Liu, William; Wanzer, Beau; Salam, Seddik Abdel; Elwany, Samy; Paunesku, Tatjana; Woloschak, Gayle E.

2013-11-01

A nanoconjugate was composed of metal oxide nanoparticles decorated with peptides and fluorescent dye and tested for DNA cleavage following UV light activation. The peptide design was based on a DNA binding domain, the so called KH domain of the hnRNPK protein. This ``KH peptide'' enabled cellular uptake of nanoconjugates and their entry into cell nuclei. The control nanoconjugate carried no peptide; it consisted only of the metal oxide nanoparticle prepared as Fe3O4@TiO2 nanocomposite and the fluorescent dye alizarin red S. These components of either construct are responsible for nanoconjugate activation by UV light and the resultant production of reactive oxygen species (ROS). Production of ROS at different subcellular locations causes damage to different components of cells: only nanoconjugates inside cell nuclei can be expected to cause DNA cleavage. Degradation of cellular DNA with KH peptide decorated nanoconjugates exceeded the DNA damage obtained from control, no-peptide nanoconjugate counterparts. Moreover, caspase activation and cell death were more extensive in the same cells.A nanoconjugate was composed of metal oxide nanoparticles decorated with peptides and fluorescent dye and tested for DNA cleavage following UV light activation. The peptide design was based on a DNA binding domain, the so called KH domain of the hnRNPK protein. This ``KH peptide'' enabled cellular uptake of nanoconjugates and their entry into cell nuclei. The control nanoconjugate carried no peptide; it consisted only of the metal oxide nanoparticle prepared as Fe3O4@TiO2 nanocomposite and the fluorescent dye alizarin red S. These components of either construct are responsible for nanoconjugate activation by UV light and the resultant production of reactive oxygen species (ROS). Production of ROS at different subcellular locations causes damage to different components of cells: only nanoconjugates inside cell nuclei can be expected to cause DNA cleavage. Degradation of cellular DNA with KH peptide decorated nanoconjugates exceeded the DNA damage obtained from control, no-peptide nanoconjugate counterparts. Moreover, caspase activation and cell death were more extensive in the same cells. Electronic supplementary information (ESI) available: http://janus.northwestern.edu/wololab/auxiliary/supplementary_data_2013.docx. See DOI: 10.1039/c3nr02203j

The Fe(III) and Ga(III) coordination chemistry of 3-(1-hydroxymethylidene) and 3-(1-hydroxydecylidene)-5-(2-hydroxyethyl)pyrrolidine-2,4-dione: Novel tetramic acid degradation products of homoserine lactone bacterial quorum sensing molecules

PubMed Central

Romano, Ariel A.; Hahn, Tobias; Davis, Nicole; Lowery, Colin A.; Struss, Anjali K.; Janda, Kim D.; Böttger, Lars H.; Matzanke, Berthold F.; Carrano, Carl J.

2011-01-01

Bacteria use small diffusible molecules to exchange information in a process called quorum sensing (QS). An important class of quorum sensing molecules used by Gram-negative bacteria is the family of N-acylhomoserine lactones (HSL). It was recently discovered that a degradation product of the QS molecule 3-oxo-C12-homoserine lactone, the tetramic acid 3-(1-hydroxydecylidene)-5-(2-hydroxyethyl)pyrrolidine-2,4-dione, is a potent antibacterial agent, thus implying roles for QS outside of simply communication. Because these tetramic acids also appear to bind iron with appreciable affinity it was suggested that metal binding might contribute to their biological activity. Here, using a variety of spectroscopic tools, we describe the coordination chemistry of both the methylidene and decylidene tetramic acid derivatives with Fe(III) and Ga(III) and discuss the potential biological significance of such metal binding. PMID:22178671

[Influence of liquid ceramic additive on binding of heavy metal during the vitrification of fly ash from municipal solid waste incinerator].

PubMed

Li, Run-dong; Nie, Yong-feng; Li, Ai-min; Wang, Lei; Chi, Yong; Cen, Ke-fa

2004-09-01

Vitrification process can effectively control the leachability of heavy metals in fly ash generated from municipal solid waste incinerator (MWSI). The use of liquid ceramic (LC) additive as a heavy metal chemical stabilization agent was evaluated for MSWI fly ash. The residuals of chromium, lead and zinc in slag increase by different degree with liquid ceramic additive at 1400 degrees C, while those of cadmium and copper decreases. The migrating characteristic of nickel is hardly affected by the additive less than 10%. The volatilization of Cr and Zn occurs after 61 minute with 10% addition of LC, and the binding efficiency of Cr decreases with increasing of melting temperature. The results indicate that the binding efficiency of heavy metals was affected greatly by LC additive and showed significant differences according to type of heavy metal during melting process. The short melting time (no longer than 33 min) is useful to obtain high binding efficiency of heavy metals.

Insight into the Interaction of Metal Ions with TroA from Streptococcus suis

PubMed Central

Zheng, Beiwen; Zhang, Qiangmin; Gao, Jia; Han, Huiming; Li, Ming; Zhang, Jingren; Qi, Jianxun; Yan, Jinghua; Gao, George F.

2011-01-01

Background The scavenging ability of sufficient divalent metal ions is pivotal for pathogenic bacteria to survive in the host. ATP-binding cassette (ABC)-type metal transporters provide a considerable amount of different transition metals for bacterial growth. TroA is a substrate binding protein for uptake of multiple metal ions. However, the function and structure of the TroA homologue from the epidemic Streptococcus suis isolates (SsTroA) have not been characterized. Methodology/Principal Findings Here we determined the crystal structure of SsTroA from a highly pathogenic streptococcal toxic shock syndrome (STSS)-causing Streptococcus suis in complex with zinc. Inductively coupled plasma mass spectrometry (ICP-MS) analysis revealed that apo-SsTroA binds Zn2+ and Mn2+. Both metals bind to SsTroA with nanomolar affinity and stabilize the protein against thermal unfolding. Zn2+ and Mn2+ induce distinct conformational changes in SsTroA compared with the apo form as confirmed by both circular dichroism (CD) and nuclear magnetic resonance (NMR) spectra. NMR data also revealed that Zn2+/Mn2+ bind to SsTroA in either the same site or an adjacent region. Finally, we found that the folding of the metal-bound protein is more compact than the corresponding apoprotein. Conclusions/Significance Our findings reveal a mechanism for uptake of metal ions in S. suis and this mechanism provides a reasonable explanation as to how SsTroA operates in metal transport. PMID:21611125

Micron-size metal-binding hydrogel particles improve germination and radicle elongation of Australian metallophyte grasses in mine waste rock and tailings.

PubMed

Guterres, J; Rossato, L; Pudmenzky, A; Doley, D; Whittaker, M; Schmidt, S

2013-03-15

Metal contamination of landscapes as a result of mining and other industrial activities is a pervasive problem worldwide. Metal contaminated soils often lack effective vegetation cover and are prone to contaminant leaching and dispersion through erosion, leading to contamination of the environment. Metal-binding hydrogel particle amendments could ameliorate mine wastes prior to planting and enhance seedling emergence. In this study, micron-size thiol functional cross-linked acrylamide polymer hydrogel particles (X3) were synthesised and tested in laboratory-scale experiments on phytotoxic mine wastes to determine their capacity to: (i) increase substrate water holding capacity (WHC); (ii) reduce metal availability to plants to below the phytotoxicity threshold; and (iii) enhance germination characteristics and early radicle development of two Australian metallophyte grasses under limiting and non-limiting water conditions. Addition of X3 to mine wastes significantly increased their WHC and lowered toxic soluble metal concentrations in mine waste leachates. Germination percentages and radicle elongation of both grasses in wastes were significantly increased. Highest germination percentages and greater radicle development recorded in X3 amended wastes under water limited conditions suggests that X3 was able to ameliorate metal toxicity to radicles, and provide moisture, which improved the imbibition and consequent germination of the seeds. Copyright © 2013 Elsevier B.V. All rights reserved.

Identification of metal ion binding sites based on amino acid sequences

PubMed Central

Cao, Xiaoyong; Zhang, Xiaojin; Gao, Sujuan; Ding, Changjiang; Feng, Yonge; Bao, Weihua

2017-01-01

The identification of metal ion binding sites is important for protein function annotation and the design of new drug molecules. This study presents an effective method of analyzing and identifying the binding residues of metal ions based solely on sequence information. Ten metal ions were extracted from the BioLip database: Zn2+, Cu2+, Fe2+, Fe3+, Ca2+, Mg2+, Mn2+, Na+, K+ and Co2+. The analysis showed that Zn2+, Cu2+, Fe2+, Fe3+, and Co2+ were sensitive to the conservation of amino acids at binding sites, and promising results can be achieved using the Position Weight Scoring Matrix algorithm, with an accuracy of over 79.9% and a Matthews correlation coefficient of over 0.6. The binding sites of other metals can also be accurately identified using the Support Vector Machine algorithm with multifeature parameters as input. In addition, we found that Ca2+ was insensitive to hydrophobicity and hydrophilicity information and Mn2+ was insensitive to polarization charge information. An online server was constructed based on the framework of the proposed method and is freely available at http://60.31.198.140:8081/metal/HomePage/HomePage.html. PMID:28854211

Identification of metal ion binding sites based on amino acid sequences.

PubMed

Cao, Xiaoyong; Hu, Xiuzhen; Zhang, Xiaojin; Gao, Sujuan; Ding, Changjiang; Feng, Yonge; Bao, Weihua

2017-01-01

The identification of metal ion binding sites is important for protein function annotation and the design of new drug molecules. This study presents an effective method of analyzing and identifying the binding residues of metal ions based solely on sequence information. Ten metal ions were extracted from the BioLip database: Zn2+, Cu2+, Fe2+, Fe3+, Ca2+, Mg2+, Mn2+, Na+, K+ and Co2+. The analysis showed that Zn2+, Cu2+, Fe2+, Fe3+, and Co2+ were sensitive to the conservation of amino acids at binding sites, and promising results can be achieved using the Position Weight Scoring Matrix algorithm, with an accuracy of over 79.9% and a Matthews correlation coefficient of over 0.6. The binding sites of other metals can also be accurately identified using the Support Vector Machine algorithm with multifeature parameters as input. In addition, we found that Ca2+ was insensitive to hydrophobicity and hydrophilicity information and Mn2+ was insensitive to polarization charge information. An online server was constructed based on the framework of the proposed method and is freely available at http://60.31.198.140:8081/metal/HomePage/HomePage.html.

Reversible surface binding of cadmium and lead by lactic acid and bifidobacteria.

PubMed

Teemu, Halttunen; Seppo, Salminen; Jussi, Meriluoto; Raija, Tahvonen; Kalle, Lertola

2008-07-15

Extensive cadmium and lead contamination of water has been reported to occur locally as a result of human activities. Lactic acid bacteria have been reported to remove cadmium and lead from water. The aim of this work was to clarify the mechanisms of cadmium and lead removal from water. In addition, the effect of other metals, reversibility of binding and recyclability of the biomass was assessed. Based on our earlier data, the two most promising lactic acid bacteria, Lactobacillus fermentum ME3 and Bifidobacterium longum 46, were selected for these experiments. The results showed that the presence of other cationic metals and blocking of carboxyl and phosphoryl groups reduced cadmium and lead removal. These results suggest involvement of electrostatic interactions in cadmium and lead removal, and support our earlier findings. Transmission electron micrographs showed large deposits of lead on the bacterial surface suggesting formation of metallic lead precipitates. Both cadmium and lead removal were reversible processes established by full recovery of removed metal after desorption with dilute solutions of EDTA and HNO(3). Resorption capacity of both biomasses tested was reduced after regeneration with 10 mM EDTA and 15 mM HNO(3). Taken together, the results suggest involvement of several reversible mechanisms such as ion exchange and precipitation in cadmium and lead binding by lactic acid bacteria. The results show that specific lactic acid bacteria have the potential for removal of cadmium and lead from water although reduction in resorption capacity after regeneration of the biomass may form a problem. Since the studies so far have mainly focused on removal of single metals from pure water, metal removal in conditions of natural waters should be assessed in further experiments.

Promotional effects of chemisorbed oxygen and hydroxide in the activation of C-H and O-H bonds over transition metal surfaces

NASA Astrophysics Data System (ADS)

Hibbitts, David; Neurock, Matthew

2016-08-01

Electronegative coadsorbates such as atomic oxygen (O*) and hydroxide (OH*) can act as Brønsted bases when bound to Group 11 as well as particular Group 8-10 metal surfaces and aid in the activation of X-H bonds. First-principle density functional theory calculations were carried out to systematically explore the reactivity of the C-H bonds of methane and surface methyl intermediates as well as the O-H bond of methanol directly and with the assistance of coadsorbed O* and OH* intermediates over Group 11 (Cu, Ag, and Au) and Group 8-10 transition metal (Ru, Rh, Pd, Os, Ir, and Pt) surfaces. C-H as well as O-H bond activation over the metal proceeds via a classic oxidative addition type mechanism involving the insertion of the metal center into the C-H or O-H bond. O* and OH* assist C-H and O-H activation over particular Group 11 and Group 8-10 metal surfaces via a σ-bond metathesis type mechanism involving the oxidative addition of the C-H or O-H bond to the metal along with a reductive deprotonation of the acidic C-H and O-H bond over the M-O* or M-OH* site pair. The O*- and OH*-assisted C-H activation paths are energetically preferred over the direct metal catalyzed C-H scission for all Group 11 metals (Cu, Ag, and Au) with barriers that are 0.4-1.5 eV lower than those for the unassisted routes. The barriers for O*- and OH*-assisted C-H activation of CH4 on the Group 8-10 transition metals, however, are higher than those over the bare transition metal surfaces by as much as 1.4 eV. The C-H activation of adsorbed methyl species show very similar trends to those for CH4 despite the differences in structure between the weakly bound methane and the covalently adsorbed methyl intermediates. The activation of the O-H bond of methanol is significantly promoted by O* as well as OH* intermediates over both the Group 11 metals (Cu, Ag, and Au) as well as on all Group 8-10 metals studied (Ru, Rh, Pd, Os, Ir, and Pt). The O*- and OH*-assisted CH3O-H barriers are 0.6 to 2.0 eV lower than unassisted barriers, with the largest differences occurring on Group 11 metals. The higher degree of O*- and OH*-promotion in activating methanol over that in methane and methyl is due to the stronger interaction between the basic O* and OH* sites and the acidic proton in the O-H bond of methanol versus the non-acidic H in the C-H bond of methane. A detailed analysis of the binding energies and the charges for O* and OH* on different metal surfaces indicates that the marked differences in the properties and reactivity of O* and OH* between the Group 11 and Group 8-10 metals is due to the increased negative charge on the O-atoms (in O* as well as OH*) bound to Group 11 metals. The promotional effects of O* and OH* are consistent with a proton-coupled electron transfer and the cooperative role of the metal-O* or metal-OH* pair in carrying out the oxidative addition and reductive deprotonation of the acidic C-H and O-H bonds. Ultimately, the ability of O* or OH* to act as a Brønsted base depends upon its charge, its binding energy on the metal surface (due to shifts in its position during X-H activation), and the acidity of the H-atom being abstracted.

Amelioration of municipal sludge by Pistia stratiotes L.: role of antioxidant enzymes in detoxification of metals.

PubMed

Tewari, Anamika; Singh, Ragini; Singh, Naveen Kumar; Rai, U N

2008-12-01

Pistia stratiotes when exposed to mixture of municipal sludge and effluent accumulated substantial amount of metals in the fronds and roots. With the increase in the metal accumulation by the plants, a reduction in the concentration of metals was found in leachates. The treated plants showed reduced level in chlorophylls but enhanced level of carotenoids and protein. The plant showed a concomitant increase in the activities of antioxidant enzymes; superoxide dismutase, guiacol peroxidase and also an enhanced level of lipid peroxidation. The activities were more in the root tissues as compared to frond. Initially a reduced level of cysteine content in roots of sludge treated plant as compared to control was found, but with time duration it increased progressively. The level of non-protein thiols also increased gradually at all the durations in both fronds and roots. Thus, beside the reduction of metals from municipal sludge, the ability of P. stratiotes to combat metal generated damages by induced synthesis of antioxidant enzymes and other metal binding ligands shows its suitability for the phytoremediation of the waste.

Carbon dioxide hydrogenation on Ni(110).

PubMed

Vesselli, Erik; De Rogatis, Loredana; Ding, Xunlei; Baraldi, Alessandro; Savio, Letizia; Vattuone, Luca; Rocca, Mario; Fornasiero, Paolo; Peressi, Maria; Baldereschi, Alfonso; Rosei, Renzo; Comelli, Giovanni

2008-08-27

We demonstrate that the key step for the reaction of CO 2 with hydrogen on Ni(110) is a change of the activated molecule coordination to the metal surface. At 90 K, CO 2 is negatively charged and chemically bonded via the carbon atom. When the temperature is increased and H approaches, the H-CO 2 complex flips and binds to the surface through the two oxygen atoms, while H binds to the carbon atom, thus yielding formate. We provide the atomic-level description of this process by means of conventional ultrahigh vacuum surface science techniques combined with density functional theory calculations and corroborated by high pressure reactivity tests. Knowledge about the details of the mechanisms involved in this reaction can yield a deeper comprehension of heterogeneous catalytic organic synthesis processes involving carbon dioxide as a reactant. We show why on Ni the CO 2 hydrogenation barrier is remarkably smaller than that on the common Cu metal-based catalyst. Our results provide a possible interpretation of the observed high catalytic activity of NiCu alloys.

Calorimetric studies of the interactions of metalloenzyme active site mimetics with zinc-binding inhibitors.

PubMed

Robinson, Sophia G; Burns, Philip T; Miceli, Amanda M; Grice, Kyle A; Karver, Caitlin E; Jin, Lihua

2016-07-19

The binding of drugs to metalloenzymes is an intricate process that involves several interactions, including binding of the drug to the enzyme active site metal, as well as multiple interactions between the drug and the enzyme residues. In order to determine the free energy contribution of Zn(2+) binding by known metalloenzyme inhibitors without the other interactions, valid active site zinc structural mimetics must be formed and binding studies need to be performed in biologically relevant conditions. The potential of each of five ligands to form a structural mimetic with Zn(2+) was investigated in buffer using Isothermal Titration Calorimetry (ITC). All five ligands formed strong 1 : 1 (ligand : Zn(2+)) binary complexes. The complexes were used in further ITC experiments to study their interaction with 8-hydroxyquinoline (8-HQ) and/or acetohydroxamic acid (AHA), two bidentate anionic zinc-chelating enzyme inhibitors. It was found that tetradentate ligands were not suitable for creating zinc structural mimetics for inhibitor binding in solution due to insufficient coordination sites remaining on Zn(2+). A stable binary complex, [Zn(BPA)](2+), which was formed by a tridentate ligand, bis(2-pyridylmethyl)amine (BPA), was found to bind one AHA in buffer or a methanol : buffer mixture (60 : 40 by volume) at pH 7.25 or one 8-HQ in the methanol : buffer mixture at pH 6.80, making it an effective structural mimetic for the active site of zinc metalloenzymes. These results are consistent with the observation that metalloenzyme active site zinc ions have three residues coordinated to them, leaving one or two sites open for inhibitors to bind. Our findings indicate that Zn(BPA)X2 can be used as an active site structural mimetic for zinc metalloenzymes for estimating the free energy contribution of zinc binding to the overall inhibitor active site interactions. Such use will help aid in the rational design of inhibitors to a variety of zinc metalloenzymes.

Identification of a magnesium-dependent NAD(P)(H)-binding domain in the nicotinoprotein methanol dehydrogenase from Bacillus methanolicus.

PubMed

Hektor, Harm J; Kloosterman, Harm; Dijkhuizen, Lubbert

2002-12-06

The Bacillus methanolicus methanol dehydrogenase (MDH) is a decameric nicotinoprotein alcohol dehydrogenase (family III) with one Zn(2+) ion, one or two Mg(2+) ions, and a tightly bound cofactor NAD(H) per subunit. The Mg(2+) ions are essential for binding of cofactor NAD(H) in MDH. A B. methanolicus activator protein strongly stimulates the relatively low coenzyme NAD(+)-dependent MDH activity, involving hydrolytic removal of the NMN(H) moiety of cofactor NAD(H) (Kloosterman, H., Vrijbloed, J. W., and Dijkhuizen, L. (2002) J. Biol. Chem. 277, 34785-34792). Members of family III of NAD(P)-dependent alcohol dehydrogenases contain three unique, conserved sequence motifs (domains A, B, and C). Domain C is thought to be involved in metal binding, whereas the functions of domains A and B are still unknown. This paper provides evidence that domain A constitutes (part of) a new magnesium-dependent NAD(P)(H)-binding domain. Site-directed mutants D100N and K103R lacked (most of the) bound cofactor NAD(H) and had lost all coenzyme NAD(+)-dependent MDH activity. Also mutants G95A and S97G were both impaired in cofactor NAD(H) binding but retained coenzyme NAD(+)-dependent MDH activity. Mutant G95A displayed a rather low MDH activity, whereas mutant S97G was insensitive to activator protein but displayed "fully activated" MDH reaction rates. The various roles of these amino acid residues in coenzyme and/or cofactor NAD(H) binding in MDH are discussed.

Thermodynamics of Pb(ii) and Zn(ii) binding to MT-3, a neurologically important metallothionein.

PubMed

Carpenter, M C; Shami Shah, A; DeSilva, S; Gleaton, A; Su, A; Goundie, B; Croteau, M L; Stevenson, M J; Wilcox, D E; Austin, R N

2016-06-01

Isothermal titration calorimetry (ITC) was used to quantify the thermodynamics of Pb(2+) and Zn(2+) binding to metallothionein-3 (MT-3). Pb(2+) binds to zinc-replete Zn7MT-3 displacing each zinc ion with a similar change in free energy (ΔG) and enthalpy (ΔH). EDTA chelation measurements of Zn7MT-3 and Pb7MT-3 reveal that both metal ions are extracted in a tri-phasic process, indicating that they bind to the protein in three populations with different binding thermodynamics. Metal binding is entropically favoured, with an enthalpic penalty that reflects the enthalpic cost of cysteine deprotonation accompanying thiolate ligation of the metal ions. These data indicate that Pb(2+) binding to both apo MT-3 and Zn7MT-3 is thermodynamically favourable, and implicate MT-3 in neuronal lead biochemistry.

The Study of the Successive Metal-Ligand Binding Energies for Fe(sup +), Fe(sup -), V(sup +) and Co(sup +)

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Ricca, Alessandra; Maitre, Philippe; Langhoff, Stephen R. (Technical Monitor)

1995-01-01

The successive binding energies of CO and H2O to Fe(sup +), CO to Fe(sup -), and H2 to Co(sup +) and V(sup +) are presented. Overall the computed results are in good agreement with experiment. The trends in binding energies are analyzed in terms of metal to ligand donation, ligand to metal donation, ligand-ligand repulsion, and changes in the metal atom, such as hybridization, promotion, and spin multiplicity. The geometry and vibrational frequencies are also shown to be directly affected by these effects.

The structure of the TsaB/TsaD/TsaE complex reveals an unexpected mechanism for the bacterial t6A tRNA-modification.

PubMed

Missoury, Sophia; Plancqueel, Stéphane; Li de la Sierra-Gallay, Ines; Zhang, Wenhua; Liger, Dominique; Durand, Dominique; Dammak, Raoudha; Collinet, Bruno; van Tilbeurgh, Herman

2018-05-08

The universal N6-threonylcarbamoyladenosine (t6A) modification at position A37 of ANN-decoding tRNAs is essential for translational fidelity. In bacteria the TsaC enzyme first synthesizes an l-threonylcarbamoyladenylate (TC-AMP) intermediate. In cooperation with TsaB and TsaE, TsaD then transfers the l-threonylcarbamoyl-moiety from TC-AMP onto tRNA. We determined the crystal structure of the TsaB-TsaE-TsaD (TsaBDE) complex of Thermotoga maritima in presence of a non-hydrolysable AMPCPP. TsaE is positioned at the entrance of the active site pocket of TsaD, contacting both the TsaB and TsaD subunits and prohibiting simultaneous tRNA binding. AMPCPP occupies the ATP binding site of TsaE and is sandwiched between TsaE and TsaD. Unexpectedly, the binding of TsaE partially denatures the active site of TsaD causing loss of its essential metal binding sites. TsaE interferes in a pre- or post-catalytic step and its binding to TsaBD is regulated by ATP hydrolysis. This novel binding mode and activation mechanism of TsaE offers good opportunities for antimicrobial drug development.

Metals in proteins: correlation between the metal-ion type, coordination number and the amino-acid residues involved in the coordination.

PubMed

Dokmanić, Ivan; Sikić, Mile; Tomić, Sanja

2008-03-01

Metal ions are constituents of many metalloproteins, in which they have either catalytic (metalloenzymes) or structural functions. In this work, the characteristics of various metals were studied (Cu, Zn, Mg, Mn, Fe, Co, Ni, Cd and Ca in proteins with known crystal structure) as well as the specificity of their environments. The analysis was performed on two data sets: the set of protein structures in the Protein Data Bank (PDB) determined with resolution <1.5 A and the set of nonredundant protein structures from the PDB. The former was used to determine the distances between each metal ion and its electron donors and the latter was used to assess the preferred coordination numbers and common combinations of amino-acid residues in the neighbourhood of each metal. Although the metal ions considered predominantly had a valence of two, their preferred coordination number and the type of amino-acid residues that participate in the coordination differed significantly from one metal ion to the next. This study concentrates on finding the specificities of a metal-ion environment, namely the distribution of coordination numbers and the amino-acid residue types that frequently take part in coordination. Furthermore, the correlation between the coordination number and the occurrence of certain amino-acid residues (quartets and triplets) in a metal-ion coordination sphere was analysed. The results obtained are of particular value for the identification and modelling of metal-binding sites in protein structures derived by homology modelling. Knowledge of the geometry and characteristics of the metal-binding sites in metalloproteins of known function can help to more closely determine the biological activity of proteins of unknown function and to aid in design of proteins with specific affinity for certain metals.

Hidden relationships between metalloproteins unveiled by structural comparison of their metal sites

NASA Astrophysics Data System (ADS)

Valasatava, Yana; Andreini, Claudia; Rosato, Antonio

2015-03-01

Metalloproteins account for a substantial fraction of all proteins. They incorporate metal atoms, which are required for their structure and/or function. Here we describe a new computational protocol to systematically compare and classify metal-binding sites on the basis of their structural similarity. These sites are extracted from the MetalPDB database of minimal functional sites (MFSs) in metal-binding biological macromolecules. Structural similarity is measured by the scoring function of the available MetalS2 program. Hierarchical clustering was used to organize MFSs into clusters, for each of which a representative MFS was identified. The comparison of all representative MFSs provided a thorough structure-based classification of the sites analyzed. As examples, the application of the proposed computational protocol to all heme-binding proteins and zinc-binding proteins of known structure highlighted the existence of structural subtypes, validated known evolutionary links and shed new light on the occurrence of similar sites in systems at different evolutionary distances. The present approach thus makes available an innovative viewpoint on metalloproteins, where the functionally crucial metal sites effectively lead the discovery of structural and functional relationships in a largely protein-independent manner.

Cysteine residue 911 in C-terminal tail of human BK(Ca)α channel subunit is crucial for its activation by carbon monoxide.

PubMed

Telezhkin, Vsevolod; Brazier, Stephen P; Mears, Ruth; Müller, Carsten T; Riccardi, Daniela; Kemp, Paul J

2011-06-01

The large conductance, voltage- and calcium-activated potassium channel, BK(Ca), is a known target for the gasotransmitter, carbon monoxide (CO). Activation of BK(Ca) by CO modulates cellular excitability and contributes to the physiology of a diverse array of processes, including vascular tone and oxygen-sensing. Currently, there is no consensus regarding the molecular mechanisms underpinning reception of CO by the BK(Ca). Here, employing voltage-clamped, inside-out patches from HEK293 cells expressing single, double and triple cysteine mutations in the BK(Ca) α-subunit, we test the hypothesis that CO regulation is conferred upon the channel by interactions with cysteine residues within the RCK2 domain. In physiological [Ca(2+)](i), all mutants carrying a cysteine substitution at position 911 (C911G) demonstrated significantly reduced CO sensitivity; the C911G mutant did not express altered Ca(2+)-sensitivity. In contrast, histidine residues in RCK1 domain, previously shown to ablate CO activation in low [Ca(2+)](i), actually increased CO sensitivity when [Ca(2+)](i) was in the physiological range. Importantly, cyanide, employed here as a substituent for CO at potential metal centres, occluded activation by CO; this effect was freely reversible. Taken together, these data suggest that a specific cysteine residue in the C-terminal domain, which is close to the Ca(2+) bowl but which is not involved in Ca(2+) activation, confers significant CO sensitivity to BK(Ca) channels. The rapid reversibility of CO and cyanide binding, coupled to information garnered from other CO-binding proteins, suggests that C911 may be involved in formation of a transition metal cluster which can bind and, thereafter, activate BK(Ca).

Structural and Thermodynamic Consequences of the Replacement of Zinc with Environmental Metals on ERα-DNA Interactions

PubMed Central

Deegan, Brian J.; Bona, Anna M.; Bhat, Vikas; Mikles, David C.; McDonald, Caleb B.; Seldeen, Kenneth L.; Farooq, Amjad

2011-01-01

Estrogen receptor α (ERα) acts as a transcription factor by virtue of the ability of its DNA-binding (DB) domain, comprised of a tandem pair of zinc fingers, to recognize the estrogen response element (ERE) within the promoters of target genes. Herein, using an array of biophysical methods, we probe structural consequences of the replacement of zinc within the DB domain of ERα with various environmental metals and their effects on the thermodynamics of binding to DNA. Our data reveal that while the DB domain reconstituted with divalent ions of zinc, cadmium, mercury and cobalt binds to DNA with affinities in the nanomolar range, divalent ions of barium, copper, iron, lead, manganese, nickel and tin are unable to regenerate DB domain with DNA-binding potential though they can compete with zinc for coordinating the cysteine ligands within the zinc fingers. We also show that the metal-free DB domain is a homodimer in solution and that the binding of various metals only results in subtle secondary and tertiary structural changes, implying that metal-coordination may only be essential for DNA-binding. Collectively, our findings provide mechanistic insights into how environmental metals may modulate the physiological function of a key nuclear receptor involved in mediating a plethora of cellular functions central to human health and disease. PMID:22038807

Carbon-Coated Core-Shell Fe-Cu Nanoparticles as Highly Active and Durable Electrocatalysts for a Zn-Air Battery.

PubMed

Nam, Gyutae; Park, Joohyuk; Choi, Min; Oh, Pilgun; Park, Suhyeon; Kim, Min Gyu; Park, Noejung; Cho, Jaephil; Lee, Jang-Soo

2015-06-23

Understanding the interaction between a catalyst and oxygen has been a key step in designing better electrocatalysts for the oxygen reduction reaction (ORR) as well as applying them in metal-air batteries and fuel cells. Alloying has been studied to finely tune the catalysts' electronic structures to afford proper binding affinities for oxygen. Herein, we synthesized a noble-metal-free and nanosized transition metal CuFe alloy encapsulated with a graphitic carbon shell as a highly efficient and durable electrocatalyst for the ORR in alkaline solution. Theoretical models and experimental results demonstrated that the CuFe alloy has a more moderate binding strength for oxygen molecules as well as the final product, OH(-), thus facilitating the oxygen reduction process. Furthermore, the nitrogen-doped graphitic carbon-coated layer, formed catalytically under the influence of iron, affords enhanced charge transfer during the oxygen reduction process and superior durability. These benefits were successfully confirmed by realizing the catalyst application in a mechanically rechargeable Zn-air battery.

Kinetic study of sunflower phospholipase Dα: interactions with micellar substrate, detergents and metals.

PubMed

Abdelkafi, Slim; Abousalham, Abdelkarim

2011-07-01

Phospholipase Dα (PLDα) purified from six-day post-germinated sunflower seeds was inactive in vitro on bilamellar substrates. It was fully active on mixed micelles made with phospholipids and a mixture of Triton-X100 and SDS at equal concentrations. It had an absolute need for divalent ions and calcium ions at millimolar concentration were the most efficient. Calcium had two effects. Firstly, using the fluorescent probe 2-p-toluidinylnaphtalene-6-sulfonate, we showed that the enzyme was able to bind calcium with a dissociation constant of 40-50 mM. This high value is probably due to the modification of the C2 domain which lacks some coordination residues allowing the binding of the metal. Secondly, using turbidity measurements, we showed that the metal ions interact with the SDS contained in the mixed micelles thus leading to an aggregated form of the substrate which is more easily hydrolyzed by PLDα. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Zinc-binding structure of a catalytic amyloid from solid-state NMR.

PubMed

Lee, Myungwoon; Wang, Tuo; Makhlynets, Olga V; Wu, Yibing; Polizzi, Nicholas F; Wu, Haifan; Gosavi, Pallavi M; Stöhr, Jan; Korendovych, Ivan V; DeGrado, William F; Hong, Mei

2017-06-13

Throughout biology, amyloids are key structures in both functional proteins and the end product of pathologic protein misfolding. Amyloids might also represent an early precursor in the evolution of life because of their small molecular size and their ability to self-purify and catalyze chemical reactions. They also provide attractive backbones for advanced materials. When β-strands of an amyloid are arranged parallel and in register, side chains from the same position of each chain align, facilitating metal chelation when the residues are good ligands such as histidine. High-resolution structures of metalloamyloids are needed to understand the molecular bases of metal-amyloid interactions. Here we combine solid-state NMR and structural bioinformatics to determine the structure of a zinc-bound metalloamyloid that catalyzes ester hydrolysis. The peptide forms amphiphilic parallel β-sheets that assemble into stacked bilayers with alternating hydrophobic and polar interfaces. The hydrophobic interface is stabilized by apolar side chains from adjacent sheets, whereas the hydrated polar interface houses the Zn 2+ -binding histidines with binding geometries unusual in proteins. Each Zn 2+ has two bis-coordinated histidine ligands, which bridge adjacent strands to form an infinite metal-ligand chain along the fibril axis. A third histidine completes the protein ligand environment, leaving a free site on the Zn 2+ for water activation. This structure defines a class of materials, which we call metal-peptide frameworks. The structure reveals a delicate interplay through which metal ions stabilize the amyloid structure, which in turn shapes the ligand geometry and catalytic reactivity of Zn 2 .

Utilization of ICP/OES for the determination of trace metal binding to different humic fractions.

PubMed

de la Rosa, G; Peralta-Videa, J R; Gardea-Torresdey, J L

2003-02-28

In this study, the use of inductively coupled plasma/optical emission spectrometry (ICP/OES) to determine multi-metal binding to three biomasses, Sphagnum peat moss, humin and humic acids is reported. All the investigations were performed under part per billion (ppb) concentrations. Batch pH profile experiments were performed using multi-metal solutions of Cd(II), Cu(II), Pb(II), Ni(II), Cr(III) and Cr(VI). The results showed that at pH 2 and 3, the metal affinity of the three biomasses exposed to the multi-metal solution that included Cr(III) presented the following order: Cu(II), Pb(II)>Ni(II)>Cr(III)>Cd(II). On the other hand, when Cr(VI) was in the heavy metal mixture, Sphagnum peat moss and humin showed the following affinity: Cu(II), Pb(II)>Ni(II)>Cr(VI)>Cd(II); however, the affinity of the humic acids was: Cu(II)>Pb(II), Cr(VI)>Ni(II)>Cd(II). The results demonstrated that pH values of 4 and 5 were the most favorable for the heavy metal binding process. At pH 5, all the metals, except for Cr(VI), were bound between 90 and 100% to the three biomasses. However, the binding capacity of humic acids decreased at pH 6 in the presence of Cr(VI). The results showed that the ICP/OES permits the determination of heavy metal binding to organic matter at ppb concentration. These results will be very useful in understanding the role of humic substances in the fate and transport of heavy metals, and thus could provide information to develop new methodologies for the removal of low concentrations of toxic heavy metals from contaminated waters.

Water-gas-shift over metal-free nanocrystalline ceria: An experimental and theoretical study

DOE PAGES

Guild, Curtis J.; Vovchok, Dimitriy; Kriz, David A.; ...

2017-01-23

A tandem experimental and theoretical investigation of a mesoporous ceria catalyst reveals the properties of the metal oxide are conducive for activity typically ascribed to metals, suggesting reduced Ce 3+ and oxygen vacancies are responsible for the inherent bi-functionality of CO oxidation and dissociation of water required for facilitating the production of H 2. The degree of reduction of the ceria, specifically the (1 0 0) face, is found to significantly influence the binding of reagents, suggesting reduced surfaces harbor the necessary reactive sites. The metal-free catalysis of the reaction is significant for catalyst design considerations, and the suite ofmore » in situ analyses provides a comprehensive study of the dynamic nature of the high surface area catalyst system. Finally, this study postulates feasible improvements in catalytic activity may redirect the purpose of the water-gas shift reaction from CO purification to primary hydrogen production.« less

Water-gas-shift over metal-free nanocrystalline ceria: An experimental and theoretical study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guild, Curtis J.; Vovchok, Dimitriy; Kriz, David A.

A tandem experimental and theoretical investigation of a mesoporous ceria catalyst reveals the properties of the metal oxide are conducive for activity typically ascribed to metals, suggesting reduced Ce 3+ and oxygen vacancies are responsible for the inherent bi-functionality of CO oxidation and dissociation of water required for facilitating the production of H 2. The degree of reduction of the ceria, specifically the (1 0 0) face, is found to significantly influence the binding of reagents, suggesting reduced surfaces harbor the necessary reactive sites. The metal-free catalysis of the reaction is significant for catalyst design considerations, and the suite ofmore » in situ analyses provides a comprehensive study of the dynamic nature of the high surface area catalyst system. Finally, this study postulates feasible improvements in catalytic activity may redirect the purpose of the water-gas shift reaction from CO purification to primary hydrogen production.« less

Why copper is preferred over iron for oxygen activation and reduction in haem-copper oxidases.

PubMed

Bhagi-Damodaran, Ambika; Michael, Matthew A; Zhu, Qianhong; Reed, Julian; Sandoval, Braddock A; Mirts, Evan N; Chakraborty, Saumen; Moënne-Loccoz, Pierre; Zhang, Yong; Lu, Yi

2017-03-01

Haem-copper oxidase (HCO) catalyses the natural reduction of oxygen to water using a haem-copper centre. Despite decades of research on HCOs, the role of non-haem metal and the reason for nature's choice of copper over other metals such as iron remains unclear. Here, we use a biosynthetic model of HCO in myoglobin that selectively binds different non-haem metals to demonstrate 30-fold and 11-fold enhancements in the oxidase activity of Cu- and Fe-bound HCO mimics, respectively, as compared with Zn-bound mimics. Detailed electrochemical, kinetic and vibrational spectroscopic studies, in tandem with theoretical density functional theory calculations, demonstrate that the non-haem metal not only donates electrons to oxygen but also activates it for efficient O-O bond cleavage. Furthermore, the higher redox potential of copper and the enhanced weakening of the O-O bond from the higher electron density in the d orbital of copper are central to its higher oxidase activity over iron. This work resolves a long-standing question in bioenergetics, and renders a chemical-biological basis for the design of future oxygen-reduction catalysts.

A universal surface complexation framework for modeling proton binding onto bacterial surfaces in geologic settings

USGS Publications Warehouse

Borrok, D.; Turner, B.F.; Fein, J.B.

2005-01-01

Adsorption onto bacterial cell walls can significantly affect the speciation and mobility of aqueous metal cations in many geologic settings. However, a unified thermodynamic framework for describing bacterial adsorption reactions does not exist. This problem originates from the numerous approaches that have been chosen for modeling bacterial surface protonation reactions. In this study, we compile all currently available potentiometric titration datasets for individual bacterial species, bacterial consortia, and bacterial cell wall components. Using a consistent, four discrete site, non-electrostatic surface complexation model, we determine total functional group site densities for all suitable datasets, and present an averaged set of 'universal' thermodynamic proton binding and site density parameters for modeling bacterial adsorption reactions in geologic systems. Modeling results demonstrate that the total concentrations of proton-active functional group sites for the 36 bacterial species and consortia tested are remarkably similar, averaging 3.2 ?? 1.0 (1??) ?? 10-4 moles/wet gram. Examination of the uncertainties involved in the development of proton-binding modeling parameters suggests that ignoring factors such as bacterial species, ionic strength, temperature, and growth conditions introduces relatively small error compared to the unavoidable uncertainty associated with the determination of cell abundances in realistic geologic systems. Hence, we propose that reasonable estimates of the extent of bacterial cell wall deprotonation can be made using averaged thermodynamic modeling parameters from all of the experiments that are considered in this study, regardless of bacterial species used, ionic strength, temperature, or growth condition of the experiment. The average site densities for the four discrete sites are 1.1 ?? 0.7 ?? 10-4, 9.1 ?? 3.8 ?? 10-5, 5.3 ?? 2.1 ?? 10-5, and 6.6 ?? 3.0 ?? 10-5 moles/wet gram bacteria for the sites with pKa values of 3.1, 4.7, 6.6, and 9.0, respectively. It is our hope that this thermodynamic framework for modeling bacteria-proton binding reactions will also provide the basis for the development of an internally consistent set of bacteria-metal binding constants. 'Universal' constants for bacteria-metal binding reactions can then be used in conjunction with equilibrium constants for other important metal adsorption and complexation reactions to calculate the overall distribution of metals in realistic geologic systems.

Adsorption of biometals to monosodium titanate in biological environments

DOE Office of Scientific and Technical Information (OSTI.GOV)

HOBBS, D.T.; MESSER, R. L. W.; LEWIS, J. B.

2005-06-06

Monosodium titanate (MST) is an inorganic sorbent/ion exchanger developed for the removal of radionuclides from nuclear wastes. We investigated the ability of MST to bind Cd(II), Hg(II), or Au(III) to establish the utility of MST for applications in environmental decontamination or medical therapy (drug delivery). Adsorption isotherms for MST were determined at pH 7-7.5 in water or phosphate-buffered saline. The extent of metal binding was determined spectroscopically by measuring the concentrations of the metals in solution before and after contact with the MST. Cytotoxic responses to MST were assessed using THP1 monocytes and succinate dehydrogenase activity. Monocytic activation by MSTmore » was assessed by TNF{alpha} secretion (ELISA) with or without lipopolysaccharide (LPS) activation. MST sorbed Cd(II), Hg(II), and Au(III) under conditions similar to that in physiological systems. MST exhibited the highest affinity for Cd(II) followed by Hg(II) and Au (III). MST (up to 100 mg/L) exhibited only minor (< 25% suppression of succinate dehydrogenase) cytotoxicity and did not trigger TNF{alpha} secretion nor modulate LPS-induced TNF{alpha} secretion from monocytes. MST exhibits high affinity for biometals with no significant biological liabilities in these introductory studies. MST deserves further scrutiny as a substance with the capacity to decontaminate biological environments or deliver metals in a controlled fashion.« less

Isohelical DNA-Binding Oligomers: Antiviral Activity and Application for the Design of Nanostructured Devices

NASA Astrophysics Data System (ADS)

Gursky, Georgy; Nikitin, Alexei; Surovaya, Anna; Grokhovsky, Sergey; Andronova, Valeria; Galegov, Georgy

We performed a systematic search for new structural motifs isohelical to double-stranded DNA and found five motifs that can be used for the design and synthesis of new DNA-binding oligomers. Some of the DNA-binding oligomers can be equipped with fluorescence chromophores and metal-chelating groups and may serve as conductive wires in nano-scaled electric circuits. A series of new DNA-binding ligands were synthesized by a modular assembly of pyrrole carboxamides and novel pseudopeptides of the form (XY)n. Here, Y is a glycine residue; n is the degree of polymerization. X is an unusual amino acid residue containing a five-membered aromatic ring. Antiviral activity of bis-linked netropsin derivatives is studied. Bis-netropsins containing 15 and 31 lysine residues at the N-termini inhibit most effectively reproduction of the herpes virus type 1 in the Vero cell culture, including virus variants resistant to acyclovir and its analogues. Antiviral activity of bis-linked netropsin derivatives is correlated with their ability to interact with long clusters of AT-base pairs in the origin of replication of the viral DNA.

Diels-Alder active-template synthesis of rotaxanes and metal-ion-switchable molecular shuttles.

PubMed

Crowley, James D; Hänni, Kevin D; Leigh, David A; Slawin, Alexandra M Z

2010-04-14

A synthesis of [2]rotaxanes in which Zn(II) or Cu(II) Lewis acids catalyze a Diels-Alder cycloaddition to form the axle while simultaneously acting as the template for the assembly of the interlocked molecules is described. Coordination of the Lewis acid to a multidentate endotopic 2,6-di(methyleneoxymethyl)pyridyl- or bipyridine-containing macrocycle orients a chelated dienophile through the macrocycle cavity. Lewis acid activation of the double bond causes it to react with an incoming "stoppered" diene, affording the [2]rotaxane in up to 91% yield. Unusually for an active-template synthesis, the metal binding site "lives on" in these rotaxanes. This was exploited in the synthesis of a molecular shuttle containing two different ligating sites in which the position of the macrocycle could be switched by complexation with metal ions [Zn(II) and Pd(II)] with different preferred coordination geometries.

Patterns of metal distribution in hypersaline microbialites during early diagenesis: Implications for the fossil record.

PubMed

Sforna, M C; Daye, M; Philippot, P; Somogyi, A; van Zuilen, M A; Medjoubi, K; Gérard, E; Jamme, F; Dupraz, C; Braissant, O; Glunk, C; Visscher, P T

2017-03-01

The use of metals as biosignatures in the fossil stromatolite record requires understanding of the processes controlling the initial metal(loid) incorporation and diagenetic preservation in living microbialites. Here, we report the distribution of metals and the organic fraction within the lithifying microbialite of the hypersaline Big Pond Lake (Bahamas). Using synchrotron-based X-ray microfluorescence, confocal, and biphoton microscopies at different scales (cm-μm) in combination with traditional geochemical analyses, we show that the initial cation sorption at the surface of an active microbialite is governed by passive binding to the organic matrix, resulting in a homogeneous metal distribution. During early diagenesis, the metabolic activity in deeper microbialite layers slows down and the distribution of the metals becomes progressively heterogeneous, resulting from remobilization and concentration as metal(loid)-enriched sulfides, which are aligned with the lamination of the microbialite. In addition, we were able to identify globules containing significant Mn, Cu, Zn, and As enrichments potentially produced through microbial activity. The similarity of the metal(loid) distributions observed in the Big Pond microbialite to those observed in the Archean stromatolites of Tumbiana provides the foundation for a conceptual model of the evolution of the metal distribution through initial growth, early diagenesis, and fossilization of a microbialite, with a potential application to the fossil record. © 2016 John Wiley & Sons Ltd.

Positive evolutionary selection of an HD motif on Alzheimer precursor protein orthologues suggests a functional role.

PubMed

Miklós, István; Zádori, Zoltán

2012-02-01

HD amino acid duplex has been found in the active center of many different enzymes. The dyad plays remarkably different roles in their catalytic processes that usually involve metal coordination. An HD motif is positioned directly on the amyloid beta fragment (Aβ) and on the carboxy-terminal region of the extracellular domain (CAED) of the human amyloid precursor protein (APP) and a taxonomically well defined group of APP orthologues (APPOs). In human Aβ HD is part of a presumed, RGD-like integrin-binding motif RHD; however, neither RHD nor RXD demonstrates reasonable conservation in APPOs. The sequences of CAEDs and the position of the HD are not particularly conserved either, yet we show with a novel statistical method using evolutionary modeling that the presence of HD on CAEDs cannot be the result of neutral evolutionary forces (p<0.0001). The motif is positively selected along the evolutionary process in the majority of APPOs, despite the fact that HD motif is underrepresented in the proteomes of all species of the animal kingdom. Position migration can be explained by high probability occurrence of multiple copies of HD on intermediate sequences, from which only one is kept by selective evolutionary forces, in a similar way as in the case of the "transcription binding site turnover." CAED of all APP orthologues and homologues are predicted to bind metal ions including Amyloid-like protein 1 (APLP1) and Amyloid-like protein 2 (APLP2). Our results suggest that HDs on the CAEDs are most probably key components of metal-binding domains, which facilitate and/or regulate inter- or intra-molecular interactions in a metal ion-dependent or metal ion concentration-dependent manner. The involvement of naturally occurring mutations of HD (Tottori (D7N) and English (H6R) mutations) in early onset Alzheimer's disease gives additional support to our finding that HD has an evolutionary preserved function on APPOs.

Positive Evolutionary Selection of an HD Motif on Alzheimer Precursor Protein Orthologues Suggests a Functional Role

PubMed Central

Miklós, István; Zádori, Zoltán

2012-01-01

HD amino acid duplex has been found in the active center of many different enzymes. The dyad plays remarkably different roles in their catalytic processes that usually involve metal coordination. An HD motif is positioned directly on the amyloid beta fragment (Aβ) and on the carboxy-terminal region of the extracellular domain (CAED) of the human amyloid precursor protein (APP) and a taxonomically well defined group of APP orthologues (APPOs). In human Aβ HD is part of a presumed, RGD-like integrin-binding motif RHD; however, neither RHD nor RXD demonstrates reasonable conservation in APPOs. The sequences of CAEDs and the position of the HD are not particularly conserved either, yet we show with a novel statistical method using evolutionary modeling that the presence of HD on CAEDs cannot be the result of neutral evolutionary forces (p<0.0001). The motif is positively selected along the evolutionary process in the majority of APPOs, despite the fact that HD motif is underrepresented in the proteomes of all species of the animal kingdom. Position migration can be explained by high probability occurrence of multiple copies of HD on intermediate sequences, from which only one is kept by selective evolutionary forces, in a similar way as in the case of the “transcription binding site turnover.” CAED of all APP orthologues and homologues are predicted to bind metal ions including Amyloid-like protein 1 (APLP1) and Amyloid-like protein 2 (APLP2). Our results suggest that HDs on the CAEDs are most probably key components of metal-binding domains, which facilitate and/or regulate inter- or intra-molecular interactions in a metal ion-dependent or metal ion concentration-dependent manner. The involvement of naturally occurring mutations of HD (Tottori (D7N) and English (H6R) mutations) in early onset Alzheimer's disease gives additional support to our finding that HD has an evolutionary preserved function on APPOs. PMID:22319430

Three-Dimensional Structure and Biophysical Characterization of Staphylococcus aureus Cell Surface Antigen-Manganese Transporter MntC

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gribenko, Alexey; Mosyak, Lidia; Ghosh, Sharmistha

MntC is a metal-binding protein component of the Mn 2 +-specific mntABC transporter from the pathogen Staphylococcus aureus. The protein is expressed during the early stages of infection and was proven to be effective at reducing both S. aureus and Staphylococcus epidermidis infections in a murine animal model when used as a vaccine antigen. MntC is currently being tested in human clinical trials as a component of a multiantigen vaccine for the prevention of S. aureus infections. To better understand the biological function of MntC, we are providing structural and biophysical characterization of the protein in this work. The three-dimensionalmore » structure of the protein was solved by X-ray crystallography at 2.2 Å resolution and suggests two potential metal binding modes, which may lead to reversible as well as irreversible metal binding. Precise Mn 2 +-binding affinity of the protein was determined from the isothermal titration calorimetry experiments using a competition approach. Differential scanning calorimetry experiments confirmed that divalent metals can indeed bind to MntC reversibly as well as irreversibly. Finally, Mn 2 +-induced structural and dynamics changes have been characterized using spectroscopic methods and deuterium–hydrogen exchange mass spectroscopy. Results of the experiments show that these changes are minimal and are largely restricted to the structural elements involved in metal coordination. Therefore, it is unlikely that antibody binding to this antigen will be affected by the occupancy of the metal-binding site by Mn 2 +.« less

Metal based biologically active compounds: Design, synthesis, DNA binding and antidiabetic activity of 6-methyl-3-formyl chromone derived hydrazones and their metal (II) complexes.

PubMed

Philip, Jessica Elizabeth; Shahid, Muhammad; Prathapachandra Kurup, M R; Velayudhan, Mohanan Puzhavoorparambil

2017-10-01

Two chromone hydrazone ligands HL 1 and HL 2 were synthesized and characterized by elemental analyses, IR, 1 H NMR & 13 C NMR, electronic absorption and mass spectra. The reactions of the chromone hydrazones with transition metals such as Ni, Cu, and Zn (II) salts of acetate afforded mononuclear metal complexes. Characterization and structure elucidation of the prepared chromone hydrazone metal (II) complexes were done by elemental, IR, electronic, EPR spectra and thermo gravimetric analyses as well as conductivity and magnetic susceptibility measurements. The spectroscopic data showed that the ligand acts as a mono basic bidentate with coordination sites are azomethine nitrogen and hydrazonic oxygen, and they exhibited distorted geometry. The biological studies involved antidiabetic activity i.e. enzyme inhibition of α-amylase and α-glucosidase, Calf Thymus - DNA (CT-DNA) interaction and molecular docking. Potential capacity of synthesized compounds to inhibit the α-amylase and α-glucosidase activity was assayed whereas DNA interaction studies were carried out with the help UV-Vis absorption titration and viscosity method. The docking studies of chromone hydrazones show that they are minor groove binders. Complexes were found to be good DNA - intercalates. Chromone hydrazones and its transition metal complexes have shown comparable antidiabetic activity with a standard drug acarbose. Copyright © 2017 Elsevier B.V. All rights reserved.

Cerium Binding Activity of Pectins Isolated from the Seagrasses Zostera marina and Phyllospadix iwatensis

PubMed Central

Khotimchenko, Yuri; Khozhaenko, Elena; Kovalev, Valeri; Khotimchenko, Maxim

2012-01-01

Cerium binding activity of three different water soluble pectin compounds of different origin was studied in a batch sorption system. The Langmuir, Freundlich and BET sorption models were adopted to describe the binding reactions between metal ions and pectin molecules. The Langmuir model provided the best fit. Within the pH range from 4.0 to 6.0, the largest amount of the cerium ions was bound by pectin isolated from the seagrass Phylospadix iwatensis in comparison to pectin extracted from the seagrass Zostera marina and pectin obtained from citrus peel (commercial grade). The Langmuir constants were also highest for the pectin samples isolated from the seagrass P. iwatensis. The results obtained from this study suggest that pectin is a prospective source for the development of radioisotope-removing pharmaceuticals. PMID:22690146

2-(Hetero(aryl)methylene)hydrazine-1-carbothioamides as potent urease inhibitors.

PubMed

Saeed, Aamer; Imran, Aqeel; Channar, Pervaiz A; Shahid, Mohammad; Mahmood, Wajahat; Iqbal, Jamshed

2015-02-01

A small series of 2-(hetero(aryl)methylene) hydrazine-1-carbothioamides including two aryl derivatives was synthesized and tested for their inhibitory activity against urease. Compound (E)-2-(Furan-2-ylmethylene) hydrazine-1-carbothioamide (3f), having a furan ring, was the most potent inhibitor of urease with an IC50 value of 0.58 μM. Molecular modeling was carried out through docking the designed compounds into the urease binding site to predict whether these derivatives have analogous binding mode to the urease inhibitors. The study revealed that all of the tested compounds bind with both metal atoms at the active site of the enzyme. The aromatic ring of the compounds forms ionic interactions with the residues, Ala(440), Asp(494), Ala(636), and Met(637). © 2014 John Wiley & Sons A/S.

Characterization of a conformationally sensitive murine monoclonal antibody directed to the metal ion-dependent adhesion site face of integrin CD11b.

PubMed

Li, Rui; Haruta, Ikuko; Rieu, Philippe; Sugimori, Takashi; Xiong, Jian-Ping; Arnaout, M Amin

2002-02-01

Integrin binding to physiologic ligands requires divalent cations and an inside-out-driven switch of the integrin to a high-affinity state. Divalent cations at the metal ion-dependent adhesion site (MIDAS) face of the alpha subunit-derived A domain provide a direct bridge between ligands and the integrin, and it has been proposed that activation dependency is caused by reorientation of the surrounding residues relative to the metal ion, forming an optimal binding interface. To gain more insight into the functional significance of the protein movements on the MIDAS face, we raised and characterized a murine mAb 107 directed against the MIDAS face of the A domain from integrin CD11b. We find that mAb 107 behaves as a ligand mimic. It binds in a divalent-cation-dependent manner to solvent-exposed residues on the MIDAS face of CD11b, blocks interaction of 11bA or the holoreceptor with ligands, and inhibits spreading and phagocytosis by human neutrophils. However, in contrast to physiologic ligands, mAb 107 preferentially binds to the inactive low-affinity form of the integrin, suggesting that its antagonistic effects are exerted in part by stabilizing the receptor in the low-affinity state. These data support a functional relevance of the protein movements on the MIDAS face and suggest that stabilizing the A domain in the low-affinity state may have therapeutic benefit.

Effect of Zn2+ binding and enzyme active site on the transition state for RNA 2′-O-transphosphorylation interpreted through kinetic isotope effects

PubMed Central

Chen, Haoyuan; Piccirilli, Joseph A.; Harris, Michael E.; York, Darrin M.

2016-01-01

Divalent metal ions, due to their ability to stabilize high concentrations of negative charge, are important for RNA folding and catalysis. Detailed models derived from the structures and kinetics of enzymes and from computational simulations have been developed. However, in most cases the specific catalytic modes involving metal ions and their mechanistic roles and effects on transition state structures remains controversial. Valuable information about the nature of the transition state is provided by measurement of kinetic isotope effects (KIEs). However, KIEs reflect changes in all bond vibrational modes that differ between the ground state and transition state. QM calculations are therefore essential for developing structural models of the transition state and evaluating mechanistic alternatives. Herein, we present computational models for Zn2+ binding to RNA 2′O-transphosphorylation reaction models that aid in the interpretation of KIE experiments. Different Zn2+ binding modes produce distinct KIE signatures, and one binding mode involving two zinc ions is in close agreement with KIEs measured for non-enzymatic catalysis by Zn2+ aquo ions alone. Interestingly, the KIE signatures in this specific model are also very close to those in RNase A catalysis. These results allow a quantitative connection to be made between experimental KIE measurements and transition state structure and bonding, and provide insight into RNA 2′O-transphosphorylation reactions catalyzed by metal ions and enzymes. PMID:25812974

Interactions of trace metals with hydrogels and filter membranes used in DET and DGT techniques.

PubMed

Garmo, Oyvind A; Davison, William; Zhang, Hao

2008-08-01

Equilibrium partitioning of trace metals between bulk solution and hydrogels/filter was studied. Under some conditions, trace metal concentrations were higher in the hydrogels or filter membranes compared to bulk solution (enrichment). In synthetic soft water, enrichment of cationic trace metals in polyacrylamide hydrogels decreased with increasing trace metal concentration. Enrichment was little affected by Ca and Mg in the concentration range typically encountered in natural freshwaters, indicating high affinity but low capacity binding of trace metals to solid structure in polyacrylamide gels. The apparent binding strength decreased in the sequence: Cu > Pb > Ni approximately to Cd approximately to Co and a low concentration of cationic Cu eliminated enrichment of weakly binding trace metal cations. The polyacrylamide gels also had an affinity for fulvic acid and/or its trace metal complexes. Enrichment of cationic Cd in agarose gel and hydrophilic polyethersulfone filter was independent of concentration (10 nM to 5 microM) but decreased with increasing Ca/ Mg concentration and ionic strength, suggesting that it is mainly due to electrostatic interactions. However, Cu and Pb were enriched even after equilibration in seawater, indicating that these metals additionally bind to sites within the agarose gel and filter. Compared to the polyacrylamide gels, agarose gel had a lower affinity for metal-fulvic complexes. Potential biases in measurements made with the diffusive equilibration in thin-films (DET) technique, identified by this work, are discussed.

Application of poly (ethyleneimine) solution as a binding agent in DGT technique for measurement of heavy metals in water.

PubMed

Sui, Dian-Peng; Fan, Hong-Tao; Li, Jing; Li, You; Li, Qiong; Sun, Ting

2013-09-30

A 0.050 mol L(-1) solution of poly (ethyleneimine) (PEI), had been used as a novel binding agent of diffusive gradients in thin-films (DGT) technique (PEI-DGT) for measuring the concentrations of labile Cu(2+), Cd(2+) and Pb(2+) in waters. The binding capacities of the PEI-DGT for Cu(2+), Cd(2+) and Pb(2+) were 11.8, 10.2 and 10.6 μmol L(-1), respectively. The performance of PEI-DGT was independence of pH in the range of 4-8 and ionic strength in the range from 1×10(-4) to 0.1 mol L(-1) (as NaNO3). PEI-DGT could measure 104.7±5.2% of the total concentration of Cd(2+) (0.500 mg L(-1)), 95.2±4.3% of the total Cu(2+) (0.500 mg L(-1)) and 99.2±3.4% of the total Pb(2+) (0.500 mg L(-1)) in synthetic solution. Effects of the ligands on the measurement of labile metals were also investigated in synthetic solutions containing the various concentrations of EDTA and humic acid. In EDTA solution, the concentrations of labile metals measured by PEI-DGT showed good agreement with the theoretical concentrations of free metal ions. In humic acid solution, the concentrations of labile metals measured by PEI-DGT decreased with the increase of the concentrations of humic acid. Several DGT devices with various binding agents, including PEI, sodium polyacrylate and poly(4-styrenesulfonate) solution, were used for the measurement of labile fractions of Cu(2+), Cd(2+) and Pb(2+) in the spiked waters and in mine wastewaters. The results showed that the concentrations of labile metal measured by DGT devices with different binding agents could be significantly different, indicating that the labile fractions of metals were dependent on the binding strength of the binding agents with metals. By choosing binding agents, the useful information on the speciation and bioavailability of the analytes can be provided. Copyright © 2013 Elsevier B.V. All rights reserved.

Cd and proton adsorption onto bacterial consortia grown from industrial wastes and contaminated geologic settings.

PubMed

Borrok, David M; Fein, Jeremy B; Kulpa, Charles F

2004-11-01

To model the effects of bacterial metal adsorption in contaminated environments, results from metal adsorption experiments involving individual pure stains of bacteria must be extrapolated to systems in which potentially dozens of bacterial species are present. This extrapolation may be made easier because bacterial consortia from natural environments appear to exhibit similar metal binding properties. However, bacteria that thrive in highly perturbed contaminated environments may exhibit significantly different adsorptive behavior. Here we measure proton and Cd adsorption onto a range of bacterial consortia grown from heavily contaminated industrial wastes, groundwater, and soils. We model the results using a discrete site surface complexation approach to determine binding constants and site densities for each consortium. The results demonstrate that bacterial consortia from different contaminated environments exhibit a range of total site densities (approximately a 3-fold difference) and Cd-binding constants (approximately a 10-fold difference). These ranges for Cd binding constants may be small enough to suggest that bacteria-metal adsorption in contaminated environments can be described using relatively few "averaged" bacteria-metal binding constants (in conjunction with the necessary binding constants for competing surfaces and ligands). However, if additional precision is necessary, modeling parameters must be developed separately for each contaminated environment of interest.

Interactions of the GM2 activator protein with phosphatidylcholine bilayers: a site-directed spin-labeling power saturation study.

PubMed

Mathias, Jordan D; Ran, Yong; Carter, Jeffery D; Fanucci, Gail E

2009-09-02

The GM2 activator protein (GM2AP) is an accessory protein that is an essential component in the catabolism of the ganglioside GM2. A function of GM2AP is to bind and extract GM2 from intralysosomal vesicles, forming a soluble protein-lipid complex, which interacts with the hydrolase Hexosaminidase A, the enzyme that cleaves the terminal sugar group of GM2. Here, we used site-directed spin labeling with power saturation electron paramagnetic resonance to determine the surface-bound orientation of GM2AP upon phosphatidylcholine vesicles. Because GM2AP extracts lipid ligands from the vesicle and is undergoing exchange on and off the vesicle surface, we utilized a nickel-chelating lipid to localize the paramagnetic metal collider to the lipid bilayer-aqueous interface. Spin-labeled sites that collide with the lipid-bound metal relaxing agent provide a means for mapping sites of the protein that interact with the lipid bilayer interface. Results show that GM2AP binds to lipid bilayers such that the residues lining the lipid-binding cavity lie on the vesicle surface. This orientation creates a favorable microenvironment that can allow for the lipid tails to flip out of the bilayer directly into the hydrophobic pocket of GM2AP.

The role of metals in production and scavenging of reactive oxygen species in photosystem II.

PubMed

Pospíšil, Pavel

2014-07-01

Metal ions play a crucial role in enzymatic reactions in all photosynthetic organisms such as cyanobacteria, algae and plants. It well known that metal ions maintain the binding of substrate in the active site of the metalloenzymes and control the redox activity of the metalloenzyme in the enzymatic reaction. A large pigment-protein complex, PSII, known to serve as a water-plastoquinone oxidoreductase, contains three metal centers comprising non-heme iron, heme iron of Cyt b559 and the water-splitting manganese complex. Metal ions bound to PSII proteins maintain the electron transport from water to plastoquinone and regulate the pro-oxidant and antioxidant activity in PSII. In this review, attention is focused on the role of PSII metal centers in (i) the formation of superoxide anion and hydroxyl radicals by sequential one-electron reduction of molecular oxygen and the formation of hydrogen peroxide by incomplete two-electron oxidation of water; and (ii) the elimination of superoxide anion radical by one-electron oxidation and reduction (superoxide dismutase activity) and of hydrogen peroxide by two-electron oxidation and reduction (catalase activity). The balance between the formation and elimination of reactive oxygen species by PSII metal centers is discussed as an important aspect in the prevention of photo-oxidative damage of PSII proteins and lipids. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Heavy metal-immobilizing organoclay facilitates polycyclic aromatic hydrocarbon biodegradation in mixed-contaminated soil.

PubMed

Biswas, Bhabananda; Sarkar, Binoy; Mandal, Asit; Naidu, Ravi

2015-11-15

Soils contaminated with a mixture of heavy metals and polycyclic aromatic hydrocarbons (PAHs) pose toxic metal stress to native PAH-degrading microorganisms. Adsorbents such as clay and modified clay minerals can bind the metal and reduce its toxicity to microorganisms. However, in a mixed-contaminated soil, an adsorption process more specific to the metals without affecting the bioavailability of PAHs is desired for effective degradation. Furthermore, the adsorbent should enhance the viability of PAH-degrading microorganisms. A metal-immobilizing organoclay (Arquad(®) 2HT-75-bentonite treated with palmitic acid) (MIOC) able to reduce metal (cadmium (Cd)) toxicity and enhance PAH (phenanthrene) biodegradation was developed and characterized in this study. The MIOC differed considerably from the parent clay in terms of its ability to reduce metal toxicity (MIOC>unmodified bentonite>Arquad-bentonite). The MIOC variably increased the microbial count (10-43%) as well as activities (respiration 3-44%; enzymatic activities up to 68%), and simultaneously maintained phenanthrene in bioavailable form in a Cd-phenanthrene mixed-contaminated soil over a 21-day incubation period. This study may lead to a new MIOC-assisted bioremediation technique for PAHs in mixed-contaminated soils. Copyright © 2015 Elsevier B.V. All rights reserved.

Dredging-related mobilisation of trace metals: a case study in The Netherlands.

PubMed

van den Berg, G A; Meijers, G G; van der Heijdt, L M; Zwolsman, J J

2001-06-01

Mobilisation of contaminants is an important issue in environmental risk assessment of dredging projects. This study has aimed at identifying the effects of dredging on mobilisation of trace metals (Zn, Cu, Cd and Pb). The intensities and time scales of trace metal mobilisation were investigated during an experimental dredging project conducted under field conditions. The loss of contaminated dredge spoil is mainly reflected by increasing levels of trace metals in the suspended matter, dissolved trace metal concentrations in the water column are not significantly influenced by the dredging activities. This indicates a strong binding mechanism of trace metals to the solid phase or a fast redistribution over sorptive phases in response to oxidation of e.g. trace metal sulphides. Given the differences in levels of reactive phases (Mn, Fe, sulphides and organic matter) between the riverine suspended matter and the sediments, changes in the levels of these parameters in the suspended matter upon dredging may give information on the processes influencing the behaviour of trace metals and on the potential loss of sediment during dredging operations. Therefore, we recommend to routinely measure these parameters in studies on contaminant behaviour related to dredging activities.

Metal complexes of quinolone antibiotics and their applications: an update.

PubMed

Uivarosi, Valentina

2013-09-11

Quinolones are synthetic broad-spectrum antibiotics with good oral absorption and excellent bioavailability. Due to the chemical functions found on their nucleus (a carboxylic acid function at the 3-position, and in most cases a basic piperazinyl ring (or another N-heterocycle) at the 7-position, and a carbonyl oxygen atom at the 4-position) quinolones bind metal ions forming complexes in which they can act as bidentate, as unidentate and as bridging ligand, respectively. In the polymeric complexes in solid state, multiple modes of coordination are simultaneously possible. In strongly acidic conditions, quinolone molecules possessing a basic side nucleus are protonated and appear as cations in the ionic complexes. Interaction with metal ions has some important consequences for the solubility, pharmacokinetics and bioavailability of quinolones, and is also involved in the mechanism of action of these bactericidal agents. Many metal complexes with equal or enhanced antimicrobial activity compared to the parent quinolones were obtained. New strategies in the design of metal complexes of quinolones have led to compounds with anticancer activity. Analytical applications of complexation with metal ions were oriented toward two main directions: determination of quinolones based on complexation with metal ions or, reversely, determination of metal ions based on complexation with quinolones.

Porous metals from sintering of nanoparticles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cappillino, Patrick J.; Robinson, David B.

A method including encapsulating or capping metallic nanoparticles by a dendrimer or a polymer with binding sites for metal particless or metal ions dispersed in a fluid; modifying the fluid to disrupt the interaction of the dendrimer or polymer with the particles; and subsequently or concomitantly sintering or partially consolidating the zero valent metal. A method including introducing a first metal salt and a second metal salt into a dendrimer or a polymer with binding sites for metals or metal ions; reducing a metal ion of the first metal salt to a zero valent first metal and a metal ionmore » of the second metal salt to a zero valend second metal; disrupting an interaction between the dendrimer or the polymer and the first metal and the second metal; and sintering or partially consolidating the first metal and the second metal.« less

Non-covalent and coordination interactions in Cu(II) systems with uridine, uridine 5'-monophosphate and triamine or tetramine as biogenic amine analogues in aqueous solutions.

PubMed

Łomozik, Lechosław; Jastrzab, Renata

2003-10-01

Reactions of metallation and non-covalent interactions have been studied in ternary systems of Cu(II) ions with uridine, uridine 5'-monophosphate and diamines or triamines. It has been found that in metal-free systems the reaction centres of the nucleoside with the polyamine are the donor nitrogen atoms N(3) and protonated -NH(x) groups of the amines. In comparison to systems with adenosine or cytidine, the pH range of complex formation is shifted towards higher values. It is a consequence of significantly higher basicity of uridine and in agreement with the ion-ion, ion-dipole interaction model assumed. Formation of molecular complexes of uridine 5'-monophosphate with polyamines at a low pH is the result of activity of the phosphate group which plays the role of a negatively charged reaction site. Non-covalent interactions interfere in processes of bioligand metallation. Centres of weak interactions are simultaneously binding sites of metal ions. In protonated Cu(Urd)(PA)H(x) complexes, coordination has been found to involve the N(3) atom from the nucleoside and two donor nitrogen atoms from the polyamine (PA). In the heteroligand species Cu(Urd)(PA), despite deprotonation of all amine groups, one of these groups is located outside the inner coordination sphere. In complexes with uridine-5'-monophosphate, the phosphate group is active in metallation. Moreover, in certain coordination compounds this group is engaged in non-covalent interactions with PA molecules, despite binding Cu ions, as has been shown on the basis of equilibrium and spectral studies.

Synthesis, spectroscopic, thermal and molecular modeling studies of Zn2+, Cd2+ and UO22+ complexes of Schiff bases containing triazole moiety. Antimicrobial, anticancer, antioxidant and DNA binding studies.

PubMed

Gaber, Mohamed; El-Ghamry, Hoda A; Fathalla, Shaimaa K; Mansour, Mohammed A

2018-02-01

A novel series of Zn 2+ , Cd 2+ and UO 2 2+ complexes of ligands namely 1-[(5-mercapto-1H-1,2,4-triazole-3-ylimino) methyl]naphthalene-2-ol (HL 1 ) and [(1H-1,2,4-triazole-3-ylimino) methyl] naphthalene-2-ol (HL 2 ) have been prepared and characterized by different analytical and spectral techniques. The stoichiometry, stereochemistry, conductivity measurements and mode of bonding of the complexes have been elucidated. Accurate comparison of the IR spectra of the ligands with their metal chelates proved the involvement of nitrogen atoms of the azomethine group and/or triazole ring in chelation in addition to the deprotonated hydroxyl oxygen. The UV-Vis and molar conductance data supported the octahedral geometry for the metal complexes. TGA technique has been used to study the thermal decomposition way of the metal complexes and the thermo kinetic parameters were estimated. Valuable information is obtained from calculations of molecular parameters using the molecular modeling techniques. The interaction between the metal complexes and CT-DNA has been studied from which the binding constants (k b ) were calculated. The Schiff bases and their metal chelates have shown potent antimicrobial, antioxidant and antitumor activities. The antitumor activities of the compounds have been tested in vitro against HEPG2 cell line and in silico by the molecular docking analysis with the VEGFR-2 receptor responsible for angiogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

Multiheteromacrocycles that Complex Metal Ions. Ninth Progress Report (includes results of last three years), 1 May 1980 -- 30 April 1983

DOE R&D Accomplishments Database

Cram, D. J.

1982-09-15

The overall objective of this research is to design, synthesize, and evaluate cyclic and polycyclic host organic compounds for the abilities to complex and lipophilize guest metal ions, their complexes, and their clusters. Host organic compounds consist of strategically placed solvating, coordinating, and ion-pairing sites tied together by covalent bonds through hydrocarbon units around cavities shaped to be occupied by guest metal ions, or by metal ions plus their ligands. Specificity in complexation is sought by matching the following properties of host and guest: cavity and metal ion sizes; geometric arrangements of binding sites; numbers of binding sites; characters of binding sites; and valences. The hope is to synthesize new classes of compounds useful in the separation of metal ions, their complexes, and their clusters.

Computational approaches for de novo design and redesign of metal-binding sites on proteins.

PubMed

Akcapinar, Gunseli Bayram; Sezerman, Osman Ugur

2017-04-28

Metal ions play pivotal roles in protein structure, function and stability. The functional and structural diversity of proteins in nature expanded with the incorporation of metal ions or clusters in proteins. Approximately one-third of these proteins in the databases contain metal ions. Many biological and chemical processes in nature involve metal ion-binding proteins, aka metalloproteins. Many cellular reactions that underpin life require metalloproteins. Most of the remarkable, complex chemical transformations are catalysed by metalloenzymes. Realization of the importance of metal-binding sites in a variety of cellular events led to the advancement of various computational methods for their prediction and characterization. Furthermore, as structural and functional knowledgebase about metalloproteins is expanding with advances in computational and experimental fields, the focus of the research is now shifting towards de novo design and redesign of metalloproteins to extend nature's own diversity beyond its limits. In this review, we will focus on the computational toolbox for prediction of metal ion-binding sites, de novo metalloprotein design and redesign. We will also give examples of tailor-made artificial metalloproteins designed with the computational toolbox. © 2017 The Author(s).

Further insights into the metal ion binding abilities and the metalation pathway of a plant metallothionein from Musa acuminata

PubMed Central

Cabral, Augusto C. S.; Jakovleska, Jovana; Deb, Aniruddha; Penner-Hahn, James E.; Pecoraro, Vincent L.

2017-01-01

The superfamily of metallothioneins (MTs) combines a diverse group of metalloproteins, sharing the characteristics of rather low molecular weight and high cysteine content. The latter provides MTs with the capability to coordinate thiophilic metal ions, in particular those with a d10 electron configuration. The sub-family of plant MT3 proteins is only poorly characterized and there is a complete lack of three-dimensional structure information. Building upon our previous results on the Musa acuminata MT3 (musMT3) protein, the focus of the present work is to understand the metal cluster formation process, the role of the single histidine residue present in musMT3, and the metal ion binding affinity. We concentrate our efforts on the coordination of ZnII and CdII ions, using CoII as a spectroscopic probe for ZnII binding. The overall protein-fold is analysed with a combination of limited proteolytic digestion, mass spectrometry, and dynamic light scattering. Histidine coordination of metal ions is probed with extended X-ray absorption fine structure spectroscopy and CoII titration experiments. Initial experiments with isothermal titration calorimetry provide insights into the thermodynamics of metal ion binding. PMID:29218632

Transition metal ion capture using functional mesoporous carbon made with 1,10-phenanthroline

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chouyyok, Wilaiwan; Yantasee, Wassana; Shin, Yongsoon

2009-11-01

Functional mesoporous carbon has been built using 1,10-phenanthroline as the fundamental building block, resulting in a nanoporous, high surface area sorbent capable of selectively binding transition metal ions. This material had a specific surface area of 870 m2/g, an average pore size of about 30Å, and contained as much as 8.2 weight percent N. Under acidic conditions, where the 1,10-phenanthroline ligand is protonated, this material was found to be an effective anion exchange material for transition metal anions like PdCl4-2 and H2VO4-1. 1,10-phenanthroline functionalized mesoporous carbon (“Phen-FMC”) was found to have a high affinity for Cu(II), even down to amore » pH of 1. At pHs above 5, Phen-FMC was found to bind a variety of transition metal cations (e.g. Co(II), Ni(II), Zn(II), etc.) from filtered ground water, river water and seawater. Phen-FMC displayed rapid sorption kinetics with Co(II) in filtered river water, reaching equilibrium in less than an hour, and easily lowering the [Co(II)] to sub-ppb levels. Phen-FMC was found to be more effective for transition metal ion capture than ion exchange resin or activated carbon.« less

Enhancing the use of waste activated sludge as bio-fuel through selectively reducing its heavy metal content.

PubMed

Dewil, Raf; Baeyens, Jan; Appels, Lise

2007-06-18

Power plant or cement kiln co-incineration are important disposal routes for the large amounts of waste activated sludge (WAS) which are generated annually. The presence of significant amounts of heavy metals in the sludge however poses serious problems since they are partly emitted with the flue gases (and collected in the flue gas dedusting) and partly incorporated in the ashes of the incinerator: in both cases, the disposal or reuse of the fly ash and bottom ashes can be jeopardized since subsequent leaching in landfill disposal can occur, or their "pozzolanic" incorporation in cement cannot be applied. The present paper studies some physicochemical methods for reducing the heavy metal content of WAS. The used techniques include acid and alkaline thermal hydrolysis and Fenton's peroxidation. By degrading the extracellular polymeric substances, binding sites for a large amount of heavy metals, the latter are released into the sludge water. The behaviour of several heavy metals (Cd, Cr, Cu, Hg, Pb, Ni, Zn) was assessed in laboratory tests. Results of these show a significant reduction of most heavy metals.

Nickel(II) and palladium(II) triphenylphosphine complexes incorporating tridentate Schiff base ligands: Synthesis, characterization and biocidal activities

NASA Astrophysics Data System (ADS)

Shabbir, Muhammad; Akhter, Zareen; Ashraf, Ahmad Raza; Ismail, Hammad; Habib, Anum; Mirza, Bushra

2017-12-01

Nickel(II) and palladium(II) triphenylphosphine complexes incorporating tridentate Schiff bases have been prepared and characterized by elemental analysis as well as by spectroscopic techniques (FTIR & NMR). The synthesized compounds were assessed to check their potential biocidal activity by using different biological assays (brine shrimp cytotoxicity, antimicrobial, antioxidant, antitumor and drug-DNA interaction). Results of brine shrimp cytotoxicity assay showed that ligand molecules are more bioactive than metal complexes with LD50 as low as 12.4 μg/mL. The prominent antitumor activity was shown by nickel complexes while the palladium complexes exhibited moderate activity. The synthesized compounds have shown high propensity for DNA binding either through intercalation or groove binding which represents the mechanism of antitumor effect of these compounds. Additionally, ligand molecules and nickel metal complexes showed significant antioxidant activity with IC50 values as low as 3.1 μg/mL and 18.9 μg/mL respectively while palladium complexes exhibited moderate activity. Moreover, in antimicrobial assays H2L1, Ni(L1)PPh3 and H2L3 showed dual inhibition against bacterial and fungal strains while for the rest of the compounds varying degree of activity was recorded against different strains. Overall comparison of results suggests that the synthesized compounds can be promising candidate for drug formulation and development.

Computational evaluation of sub-nanometer cluster activity of singly exposed copper atom with various coordinative environment in catalytic CO2 transformation

NASA Astrophysics Data System (ADS)

Shanmugam, Ramasamy; Thamaraichelvan, Arunachalam; Ganesan, Tharumeya Kuppusamy; Viswanathan, Balasubramanian

2017-02-01

Metal cluster, at sub-nanometer level has a unique property in the activation of small molecules, in contrast to that of bulk surface. In the present work, singly exposed active site of copper metal cluster at sub-nanometer level was designed to arrive at the energy minimised configurations, binding energy, electrostatic potential map, frontier molecular orbitals and partial density of states. The ab initio molecular dynamics was carried out to probe the catalytic nature of the cluster. Further, the stability of the metal cluster and its catalytic activity in the electrochemical reduction of CO2 to CO were evaluated by means of computational hydrogen electrode via calculation of the free energy profile using DFT/B3LYP level of theory in vacuum. The activity of the cluster is ascertained from the fact that the copper atom, present in a two coordinative environment, performs a more selective conversion of CO2 to CO at an applied potential of -0.35 V which is comparatively lower than that of higher coordinative sites. The present study helps to design any sub-nano level metal catalyst for electrochemical reduction of CO2 to various value added chemicals.

Single molecule junction conductance and binding geometry

NASA Astrophysics Data System (ADS)

Kamenetska, Maria

This Thesis addresses the fundamental problem of controlling transport through a metal-organic interface by studying electronic and mechanical properties of single organic molecule-metal junctions. Using a Scanning Tunneling Microscope (STM) we image, probe energy-level alignment and perform STM-based break junction (BJ) measurements on molecules bound to a gold surface. Using Scanning Tunneling Microscope-based break-junction (STM-BJ) techniques, we explore the effect of binding geometry on single-molecule conductance by varying the structure of the molecules, metal-molecule binding chemistry and by applying sub-nanometer manipulation control to the junction. These experiments are performed both in ambient conditions and in ultra high vacuum (UHV) at cryogenic temperatures. First, using STM imaging and scanning tunneling spectroscopy (STS) measurements we explore binding configurations and electronic properties of an amine-terminated benzene derivative on gold. We find that details of metal-molecule binding affect energy-level alignment at the interface. Next, using the STM-BJ technique, we form and rupture metal-molecule-metal junctions ˜104 times to obtain conductance-vs-extension curves and extract most likely conductance values for each molecule. With these measurements, we demonstrated that the control of junction conductance is possible through a choice of metal-molecule binding chemistry and sub-nanometer positioning. First, we show that molecules terminated with amines, sulfides and phosphines bind selectively on gold and therefore demonstrate constant conductance levels even as the junction is elongated and the metal-molecule attachment point is modified. Such well-defined conductance is also obtained with paracyclophane molecules which bind to gold directly through the pi system. Next, we are able to create metal-molecule-metal junctions with more than one reproducible conductance signatures that can be accessed by changing junction geometry. In the case of pyridine-linked molecules, conductance can be reliably switched between two distinct conductance states using sub-nanometer mechanical manipulation. Using a methyl sulfide linker attached to an oligoene backbone, we are able to create a 3-nm-long molecular potentiometer, whose resistance can be tuned exponentially with Angstom-scale modulations in metal-molecule configuration. These experiments points to a new paradigm for attaining reproducible electrical characteristics of metal-organic devices which involves controlling linker-metal chemistry rather than fabricating identically structured metal-molecule interfaces. By choosing a linker group which is either insensitive to or responds reproducibly to changes in metal-molecule configuration, one can design single molecule devices with functionality more complex than a simple resistor. These ambient temperature experiments were combined with UHV conductance measurements performed in a commercial STM on amine-terminated benzene derivatives which conduct through a non-resonant tunneling mechanism, at temperatures varying from 5 to 300 Kelvin. Our results indicate that while amine-gold binding remains selective irrespective of environment, conductance is not temperature independent, in contrast to what is expected for a tunneling mechanism. Furthermore, using temperature-dependent measurements in ambient conditions we find that HOMO-conducting amines and LUMO-conducting pyridines show opposite dependence of conductance on temperature. These results indicate that energy-level alignment between the molecule and the electrodes changes as a result of varying electrode structure at different temperatures. We find that temperature can serve as a knob with which to tune transport properties of single molecule-metal junctions.

Fine ambient particles induce oxidative stress and metal binding genes in human alveolar machrophages

EPA Science Inventory

Exposure to ambient pollutant particles (APP) increased respiratory morbidity and mortality. The alveolar macrophages (AMs) are one cell type in the lung directly exposed to APP. Upon contact with APP, AMs are activated and produce reactive oxygen species, but the scope ofthis ox...

Analysis of Cold Neutron Spectra of Metals.

DTIC Science & Technology

modes. The damping of lattice vibrations in metals is of the same order of magnitude as in dielectrics with ionic binding, i.e., much higher than the damping in dielectrics with covalent binding. (Author)

The Role of Hydroxide and Metal Concentration on the Viscoelastic Properties of Metal Coordinated Gels

NASA Astrophysics Data System (ADS)

Cazzell, Seth; Holten-Andersen, Niels

Nature uses metal binding amino acids to engineer mechanical properties. An example of this engineering can be found in the mussel byssal thread. This acellular thread contains reversible intermolecular protein-metal bonds, which allows the mussel to robustly anchor to rocks, while withstanding the mechanically demanding intertidal environment. Inspired by this metal-binding material, we present a synthetic hydrogel designed to mimic this bonding behavior. The mechanical properties of this hydrogel can be controlled independently by manipulating the amount of metal relative to the metal binding ligand, and the gel's pH. Here we report how high metal to ligand ratios and low pH can be used to induce the formation of a strong, slow relaxing gels. This gel has potential applications as an energy dissipating material, and furthers our understanding of the bio-inspired engineering techniques that are used to design viscoelastic soft materials. I was supported by the Department of Defense (DoD) through the National Defense Science & Engineering Graduate Fellowship (NDSEG) Program.

From basic physics to mechanisms of toxicity: the "liquid drop" approach applied to develop predictive classification models for toxicity of metal oxide nanoparticles.

PubMed

Sizochenko, Natalia; Rasulev, Bakhtiyor; Gajewicz, Agnieszka; Kuz'min, Victor; Puzyn, Tomasz; Leszczynski, Jerzy

2014-11-21

Many metal oxide nanoparticles are able to cause persistent stress to live organisms, including humans, when discharged to the environment. To understand the mechanism of metal oxide nanoparticles' toxicity and reduce the number of experiments, the development of predictive toxicity models is important. In this study, performed on a series of nanoparticles, the comparative quantitative-structure activity relationship (nano-QSAR) analyses of their toxicity towards E. coli and HaCaT cells were established. A new approach for representation of nanoparticles' structure is presented. For description of the supramolecular structure of nanoparticles the "liquid drop" model was applied. It is expected that a novel, proposed approach could be of general use for predictions related to nanomaterials. In addition, in our study fragmental simplex descriptors and several ligand-metal binding characteristics were calculated. The developed nano-QSAR models were validated and reliably predict the toxicity of all studied metal oxide nanoparticles. Based on the comparative analysis of contributed properties in both models the LDM-based descriptors were revealed to have an almost similar level of contribution to toxicity in both cases, while other parameters (van der Waals interactions, electronegativity and metal-ligand binding characteristics) have unequal contribution levels. In addition, the models developed here suggest different mechanisms of nanotoxicity for these two types of cells.

Metal Mixture Modeling Evaluation project: 2. Comparison of four modeling approaches

USGS Publications Warehouse

Farley, Kevin J.; Meyer, Joe; Balistrieri, Laurie S.; DeSchamphelaere, Karl; Iwasaki, Yuichi; Janssen, Colin; Kamo, Masashi; Lofts, Steve; Mebane, Christopher A.; Naito, Wataru; Ryan, Adam C.; Santore, Robert C.; Tipping, Edward

2015-01-01

As part of the Metal Mixture Modeling Evaluation (MMME) project, models were developed by the National Institute of Advanced Industrial Science and Technology (Japan), the U.S. Geological Survey (USA), HDR⎪HydroQual, Inc. (USA), and the Centre for Ecology and Hydrology (UK) to address the effects of metal mixtures on biological responses of aquatic organisms. A comparison of the 4 models, as they were presented at the MMME Workshop in Brussels, Belgium (May 2012), is provided herein. Overall, the models were found to be similar in structure (free ion activities computed by WHAM; specific or non-specific binding of metals/cations in or on the organism; specification of metal potency factors and/or toxicity response functions to relate metal accumulation to biological response). Major differences in modeling approaches are attributed to various modeling assumptions (e.g., single versus multiple types of binding site on the organism) and specific calibration strategies that affected the selection of model parameters. The models provided a reasonable description of additive (or nearly additive) toxicity for a number of individual toxicity test results. Less-than-additive toxicity was more difficult to describe with the available models. Because of limitations in the available datasets and the strong inter-relationships among the model parameters (log KM values, potency factors, toxicity response parameters), further evaluation of specific model assumptions and calibration strategies is needed.

Engineered Escherichia coli Silver-Binding Periplasmic Protein That Promotes Silver Tolerance

PubMed Central

Hall Sedlak, Ruth; Hnilova, Marketa; Grosh, Carolynn; Fong, Hanson; Baneyx, Francois; Schwartz, Dan; Sarikaya, Mehmet; Tamerler, Candan

2012-01-01

Silver toxicity is a problem that microorganisms face in medical and environmental settings. Through exposure to silver compounds, some bacteria have adapted to growth in high concentrations of silver ions. Such adapted microbes may be dangerous as pathogens but, alternatively, could be potentially useful in nanomaterial-manufacturing applications. While naturally adapted isolates typically utilize efflux pumps to achieve metal resistance, we have engineered a silver-tolerant Escherichia coli strain by the use of a simple silver-binding peptide motif. A silver-binding peptide, AgBP2, was identified from a combinatorial display library and fused to the C terminus of the E. coli maltose-binding protein (MBP) to yield a silver-binding protein exhibiting nanomolar affinity for the metal. Growth experiments performed in the presence of silver nitrate showed that cells secreting MBP-AgBP2 into the periplasm exhibited silver tolerance in a batch culture, while those expressing a cytoplasmic version of the fusion protein or MBP alone did not. Transmission electron microscopy analysis of silver-tolerant cells revealed the presence of electron-dense silver nanoparticles. This is the first report of a specifically engineered metal-binding peptide exhibiting a strong in vivo phenotype, pointing toward a novel ability to manipulate bacterial interactions with heavy metals by the use of short and simple peptide motifs. Engineered metal-ion-tolerant microorganisms such as this E. coli strain could potentially be used in applications ranging from remediation to interrogation of biomolecule-metal interactions in vivo. PMID:22286990

Theoretical study of transition-metal ions bound to benzene

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Partridge, Harry; Langhoff, Stephen R.

1992-01-01

Theoretical binding energies are reported for all first-row and selected second-row transition metal ions (M+) bound to benzene. The calculations employ basis sets of at least double-zeta plus polarization quality and account for electron correlation using the modified coupled-pair functional method. While the bending is predominantly electrostatic, the binding energies are significantly increased by electron correlation, because the donation from the metal d orbitals to the benzene pi* orbitals is not well described at the self-consistent-field level. The uncertainties in the computed binding energies are estimated to be about 5 kcal/mol. Although the calculated and experimental binding energies generally agree to within their combined uncertainties, it is likely that the true binding energies lie in the lower portion of the experimental range. This is supported by the very good agreement between the theoretical and recent experimental binding energies for AgC6H6(+).

Factors governing the substitution of La3+ for Ca2+ and Mg2+ in metalloproteins: a DFT/CDM study.

PubMed

Dudev, Todor; Chang, Li-Ying; Lim, Carmay

2005-03-23

Trivalent lanthanide cations are extensively being used in biochemical experiments to probe various dication-binding sites in proteins; however, the factors governing the binding specificity of lanthanide cations for these binding sites remain unclear. Hence, we have performed systematic studies to evaluate the interactions between La3+ and model Ca2+ - and Mg2+ -binding sites using density functional theory combined with continuum dielectric methods. The calculations reveal the key factors and corresponding physical bases favoring the substitution of trivalent lanthanides for divalent Ca2+ and Mg2+ in holoproteins. Replacing Ca2+ or Mg2+ with La3+ is facilitated by (1) minimizing the solvent exposure and the flexibility of the metal-binding cavity, (2) freeing both carboxylate oxygen atoms of Asp/Glu side chains in the metal-binding site so that they could bind bidentately to La3+, (3) maximizing the number of metal-bound carboxylate groups in buried sites, but minimizing the number of metal-bound carboxylate groups in solvent-exposed sites, and (4) including an Asn/Gln side chain for sites lined with four Asp/Glu side chains. In proteins bound to both Mg2+ and Ca2+, La3+ would prefer to replace Ca2+, as compared to Mg2+. A second Mg2+-binding site with a net positive charge would hamper the Mg2+ --> La3+ exchange, as compared to the respective mononuclear site, although the La3+ substitution of the first native metal is more favorable than the second one. The findings of this work are in accord with available experimental data.

Expression of simian virus 40 T antigen in Escherichia coli: localization of T-antigen origin DNA-binding domain to within 129 amino acids.

PubMed Central

Arthur, A K; Höss, A; Fanning, E

1988-01-01

The genomic coding sequence of the large T antigen of simian virus 40 (SV40) was cloned into an Escherichia coli expression vector by joining new restriction sites, BglII and BamHI, introduced at the intron boundaries of the gene. Full-length large T antigen, as well as deletion and amino acid substitution mutants, were inducibly expressed from the lac promoter of pUC9, albeit with different efficiencies and protein stabilities. Specific interaction with SV40 origin DNA was detected for full-length T antigen and certain mutants. Deletion mutants lacking T-antigen residues 1 to 130 and 260 to 708 retained specific origin-binding activity, demonstrating that the region between residues 131 and 259 must carry the essential binding domain for DNA-binding sites I and II. A sequence between residues 302 and 320 homologous to a metal-binding "finger" motif is therefore not required for origin-specific binding. However, substitution of serine for either of two cysteine residues in this motif caused a dramatic decrease in origin DNA-binding activity. This region, as well as other regions of the full-length protein, may thus be involved in stabilizing the DNA-binding domain and altering its preference for binding to site I or site II DNA. Images PMID:2835505

Glutamate Ligation in the Ni(II)- and Co(II)-Responsive Escherichia coli Transcriptional Regulator, RcnR

DOE Office of Scientific and Technical Information (OSTI.GOV)

Carr, Carolyn E.; Musiani, Francesco; Huang, Hsin-Ting

Escherichia coli RcnR (resistance to cobalt and nickel regulator, EcRcnR) is a metal-responsive repressor of the genes encoding the Ni(II) and Co(II) exporter proteins RcnAB by binding to PRcnAB. The DNA binding affinity is weakened when the cognate ions Ni(II) and Co(II) bind to EcRcnR in a six-coordinate site that features a (N/O)5S ligand donor-atom set in distinct sites: while both metal ions are bound by the N terminus, Cys35, and His64, Co(II) is additionally bound by His3. On the other hand, the noncognate Zn(II) and Cu(I) ions feature a lower coordination number, have a solvent-accessible binding site, and coordinatemore » protein ligands that do not include the N-terminal amine. A molecular model of apo-EcRcnR suggested potential roles for Glu34 and Glu63 in binding Ni(II) and Co(II) to EcRcnR. The roles of Glu34 and Glu63 in metal binding, metal selectivity, and function were therefore investigated using a structure/function approach. X-ray absorption spectroscopy was used to assess the structural changes in the Ni(II), Co(II), and Zn(II) binding sites of Glu → Ala and Glu → Cys variants at both positions. The effect of these structural alterations on the regulation of PrcnA by EcRcnR in response to metal binding was explored using LacZ reporter assays. These combined studies indicate that while Glu63 is a ligand for both metal ions, Glu34 is a ligand for Co(II) but possibly not for Ni(II). The Glu34 variants affect the structure of the cognate metal sites, but they have no effect on the transcriptional response. In contrast, the Glu63 variants affect both the structure and transcriptional response, although they do not completely abolish the function of EcRcnR. The structure of the Zn(II) site is not significantly perturbed by any of the glutamic acid variations. The spectroscopic and functional data obtained on the mutants were used to calculate models of the metal-site structures of EcRcnR bound to Ni(II), Co(II), and Zn(II). The results are interpreted in terms of a switch mechanism, in which a subset of the metal-binding ligands is responsible for the allosteric response required for DNA release.« less

A study on the dynamics of the zraP gene expression profile and its application to the construction of zinc adsorption bacteria.

PubMed

Ravikumar, Sambandam; Yoo, Ik-keun; Lee, Sang Yup; Hong, Soon Ho

2011-11-01

Zinc ion plays essential roles in biological chemistry. Bacteria acquire Zn(2+) from the environment, and cellular concentration levels are controlled by zinc homeostasis systems. In comparison with other homeostatic systems, the ZraSR two-component system was found to be more efficient in responding to exogenous zinc concentrations. To understand the dynamic response of the bacterium ZraSR two-component system with respect to exogenous zinc concentrations, the genetic circuit of the ZraSR system was integrated with a reporter protein. This study was helpful in the construction of an E. coli system that can display selective metal binding peptides on the surface of the cell in response to exogenous zinc. The engineered bacterial system for monitoring exogenous zinc was successfully employed to detect levels of zinc as low as 0.001 mM, which directly activates the expression of chimeric ompC(t)--zinc binding peptide gene to remove zinc by adsorbing a maximum of 163.6 μmol of zinc per gram of dry cell weight. These results indicate that the engineered bacterial strain developed in the present study can sense the specific heavy metal and activates a cell surface display system that acts to remove the metal.

Unifying the 2e(-) and 4e(-) Reduction of Oxygen on Metal Surfaces.

PubMed

Viswanathan, Venkatasubramanian; Hansen, Heine Anton; Rossmeisl, Jan; Nørskov, Jens K

2012-10-18

Understanding trends in selectivity is of paramount importance for multi-electron electrochemical reactions. The goal of this work is to address the issue of 2e(-) versus 4e(-) reduction of oxygen on metal surfaces. Using a detailed thermodynamic analysis based on density functional theory calculations, we show that to a first approximation an activity descriptor, ΔGOH*, the free energy of adsorbed OH*, can be used to describe trends for the 2e(-) and 4e(-) reduction of oxygen. While the weak binding of OOH* on Au(111) makes it an unsuitable catalyst for the 4e(-) reduction, this weak binding is optimal for the 2e(-) reduction to H2O2. We find quite a remarkable agreement between the predictions of the model and experimental results spanning nearly 30 years.

A Review on Recent Patents and Applications of Inorganic Material Binding Peptides.

PubMed

Thota, Veeranjaneyulu; Perry, Carole C

2017-01-01

Although the popularity of using combinatorial display techniques for recognising unique peptides having high affinity for inorganic (nano) particles has grown rapidly, there are no systematic reviews showcasing current developments or patents on binding peptides specific to these materials. In this review, we summarize and discuss recent progress in patents on material binding peptides specifically exploring inorganic nano surfaces such as metals, metal oxides, minerals, carbonbased materials, polymer based materials, magnetic materials and semiconductors. We consider both the peptide display strategies used and the exploitation of the identified peptides in the generation of advanced nanomaterials. In order to get a clear picture on the number of patents and literature present to date relevant to inorganic material binding biomolecules and their applications, a thorough online search was conducted using national and worldwide databases. The literature search include standard bibliographic databases while patents included EPO Espacenet, WIPO patent scope, USPTO, Google patent search, Patent lens, etc. along with commercial databases such as Derwent and Patbase. Both English and American spellings were included in the searches. The initial number of patents found related to material binders were 981. After reading and excluding irrelevant patents such as organic binding peptides, works published before 2001, repeated patents, documents not in English etc., 51 highly relevant patents published from 2001 onwards were selected and analysed. These patents were further separated into six categories based on their target inorganic material and combinatorial library used. They include relevant patents on metal, metal oxide or combination binding peptides (19), magnetic and semiconductor binding peptides (8), carbon based (3), mineral (5), polymer (8) and other binders (9). Further, how these material specific binders have been used to synthesize simple to complex bio- or nano-materials, mediate the controlled biomineralization process, direct self-assembly and nanofabrication of ordered structures, facilitate the immobilization of functional biomolecules and construct inorganic-inorganic or organic-inorganic nano hybrids are concisely described. From analysis of recent literature and patents, we clearly show that biomimetic material binders are in the vanguard of new design approaches for novel nanomaterials with improved/ controlled physical and chemical properties that have no adverse effect on the structural or functional activities of the nanomaterials themselves. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

A spin transition mechanism for cooperative adsorption in metal-organic frameworks

NASA Astrophysics Data System (ADS)

Reed, Douglas A.; Keitz, Benjamin K.; Oktawiec, Julia; Mason, Jarad A.; Runčevski, Tomče; Xiao, Dianne J.; Darago, Lucy E.; Crocellà, Valentina; Bordiga, Silvia; Long, Jeffrey R.

2017-10-01

Cooperative binding, whereby an initial binding event facilitates the uptake of additional substrate molecules, is common in biological systems such as haemoglobin. It was recently shown that porous solids that exhibit cooperative binding have substantial energetic benefits over traditional adsorbents, but few guidelines currently exist for the design of such materials. In principle, metal-organic frameworks that contain coordinatively unsaturated metal centres could act as both selective and cooperative adsorbents if guest binding at one site were to trigger an electronic transformation that subsequently altered the binding properties at neighbouring metal sites. Here we illustrate this concept through the selective adsorption of carbon monoxide (CO) in a series of metal-organic frameworks featuring coordinatively unsaturated iron(II) sites. Functioning via a mechanism by which neighbouring iron(II) sites undergo a spin-state transition above a threshold CO pressure, these materials exhibit large CO separation capacities with only small changes in temperature. The very low regeneration energies that result may enable more efficient Fischer-Tropsch conversions and extraction of CO from industrial waste feeds, which currently underutilize this versatile carbon synthon. The electronic basis for the cooperative adsorption demonstrated here could provide a general strategy for designing efficient and selective adsorbents suitable for various separations.

Acid-base and copper-binding properties of three organic matter fractions isolated from a forest floor soil solution

NASA Astrophysics Data System (ADS)

van Schaik, Joris W. J.; Kleja, Dan B.; Gustafsson, Jon Petter

2010-02-01

Vast amounts of knowledge about the proton- and metal-binding properties of dissolved organic matter (DOM) in natural waters have been obtained in studies on isolated humic and fulvic (hydrophobic) acids. Although macromolecular hydrophilic acids normally make up about one-third of DOM, their proton- and metal-binding properties are poorly known. Here, we investigated the acid-base and Cu-binding properties of the hydrophobic (fulvic) acid fraction and two hydrophilic fractions isolated from a soil solution. Proton titrations revealed a higher total charge for the hydrophilic acid fractions than for the hydrophobic acid fraction. The most hydrophilic fraction appeared to be dominated by weak acid sites, as evidenced by increased slope of the curve of surface charge versus pH at pH values above 6. The titration curves were poorly predicted by both Stockholm Humic Model (SHM) and NICA-Donnan model calculations using generic parameter values, but could be modelled accurately after optimisation of the proton-binding parameters (pH ⩽ 9). Cu-binding isotherms for the three fractions were determined at pH values of 4, 6 and 9. With the optimised proton-binding parameters, the SHM model predictions for Cu binding improved, whereas the NICA-Donnan predictions deteriorated. After optimisation of Cu-binding parameters, both models described the experimental data satisfactorily. Iron(III) and aluminium competed strongly with Cu for binding sites at both pH 4 and pH 6. The SHM model predicted this competition reasonably well, but the NICA-Donnan model underestimated the effects significantly at pH 6. Overall, the Cu-binding behaviour of the two hydrophilic acid fractions was very similar to that of the hydrophobic acid fraction, despite the differences observed in proton-binding characteristics. These results show that for modelling purposes, it is essential to include the hydrophilic acid fraction in the pool of 'active' humic substances.

Cobalamin-Independent Methionine Synthase (MetE): A Face-to-Face Double Barrel that Evolved by Gene Duplication

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pejcha, Robert; Ludwig, Martha L.

2010-03-08

Cobalamin-independent methionine synthase (MetE) catalyzes the transfer of a methyl group from methyltetrahydrofolate to L-homocysteine (Hcy) without using an intermediate methyl carrier. Although MetE displays no detectable sequence homology with cobalamin-dependent methionine synthase (MetH), both enzymes require zinc for activation and binding of Hcy. Crystallographic analyses of MetE from T. maritima reveal an unusual dual-barrel structure in which the active site lies between the tops of the two ({beta}{alpha}){sub 8} barrels. The fold of the N-terminal barrel confirms that it has evolved from the C-terminal polypeptide by gene duplication; comparisons of the barrels provide an intriguing example of homologous domainmore » evolution in which binding sites are obliterated. The C-terminal barrel incorporates the zinc ion that binds and activates Hcy. The zinc-binding site in MetE is distinguished from the (Cys){sub 3}Zn site in the related enzymes, MetH and betaine-homocysteine methyltransferase, by its position in the barrel and by the metal ligands, which are histidine, cysteine, glutamate, and cysteine in the resting form of MetE. Hcy associates at the face of the metal opposite glutamate, which moves away from the zinc in the binary E {center_dot} Hcy complex. The folate substrate is not intimately associated with the N-terminal barrel; instead, elements from both barrels contribute binding determinants in a binary complex in which the folate substrate is incorrectly oriented for methyl transfer. Atypical locations of the Hcy and folate sites in the C-terminal barrel presumably permit direct interaction of the substrates in a ternary complex. Structures of the binary substrate complexes imply that rearrangement of folate, perhaps accompanied by domain rearrangement, must occur before formation of a ternary complex that is competent for methyl transfer.« less

Interaction with biomacromolecules and antiproliferative activities of Mn(II), Ni(II), Zn(II) complexes of demethylcantharate and 2,2'-bipyridine

NASA Astrophysics Data System (ADS)

Zhang, Fan; Lin, Qiu-Yue; Hu, Wan-Li; Song, Wen-Ji; Shen, Shu-Ting; Gui, Pan

2013-06-01

Three new transition metal complexes [Mn2(DCA)2(bipy)2]·5H2O (1), [M2(DCA)2(bipy)2(H2O)]·10H2O(M = Ni(II)(2);Zn(II)(3)), (DCA = demethylcantharate, 7-oxabicyclo[2,2,1]heptane-2,3-dicarboxylate, C8H8O5) were synthesized and characterized by elemental analysis, molar conductance, infrared spectra and X-ray diffraction techniques. Each metal ion was six-coordinated in complexes. Complex 1 has a Mn2O2 center. Complexes 2 and 3 have asymmetric binuclear structure. Great amount of intermolecular hydrogen-bonding and π-π* stacking interactions were formed in these complex structures. The DNA-binding properties of complexes were investigated by electronic absorption spectra and viscosity measurements. The DNA binding constants Kb/(L mol-1) were 1.71 × 104 (1), 2.62 × 104 (2) and 1.59 × 104 (3) at 298 K. The complexes could quench the intrinsic fluorescence of bovine serum albumin (BSA) strongly through static quenching. The protein binding constants Ka/(L mol-1) were 7.27 × 104 (1), 4.55 × 104 (2) and 7.87 × 104 L mol-1 (3) and binding site was one. The complexes bind more tightly with DNA and BSA than with ligands. Complexes 1 and 3 had stronger inhibition ratios than Na2(DCA) against human hepatoma cells (SMMC-7721) lines and human gastric cancer cells (MGC80-3) lines in vitro. Complex 3 showed the strongest antiproliferative activity against SMMC-7721 (IC50 = 29.46 ± 2.12 μmol L-1) and MGC80-3 (IC50 = 27.02 ± 2.38 μmol L-1), which shows potential in anti-cancer drug development.

Optimization of exopolysaccharide production from Pseudomonas stutzeri AS22 and examination of its metal-binding abilities.

PubMed

Maalej, H; Hmidet, N; Boisset, C; Buon, L; Heyraud, A; Nasri, M

2015-02-01

To investigate the effect of culture conditions and medium components on exopolysaccharide (EPS) production by Pseudomonas stutzeri AS22 and to access the EPS performance as a metal-binding exopolysaccharide. The EPS production conditions of Ps. stutzeri AS22 in submerged culture were optimized using two approaches for EPS quantification, and its metal-binding capacity was evaluated using both single and mixed metal ions systems. Maximum EPS level was achieved after 24 h of incubation at 30°C with an initial pH of 8.0, 250 rev min(-1) stirring level and 10% inoculum size. 50 g l(-1) starch, 5 g l(-1) yeast extract, 0.5 g l(-1) NaCl, 1.4 g l(-1) K2 HPO4, 0.4 g l(-1) MgSO4, 0.4 g l(-1) CaCl2 and 1 g l(-1) mannose were found to be the most suitable carbon, nitrogen, mineral and additional carbohydrate sources, respectively. From metal-binding experiments, the crude EPS showed interesting metal adsorption capacity adopting the order Pb > Co > Fe > Cu > Cd. Lead was preferentially biosorbed with a maximal uptake of 460 mg g(-1) crude EPS. Under the optimal culture requirements, EPS level reached 10.2 g l(-1) after 24 h of fermentation, seven times more than the production under initial conditions. According to the metal-binding assay, the crude EPS has potential to be used as a novel biosorbent in the treatment of heavy metals-contaminated water. Our results are interesting in terms of yield as well as efficiency for the potential use of the Ps. stutzeri exopolysaccharide as a metal-absorbent polymer in the bioremediation field. © 2014 The Society for Applied Microbiology.

Superparamagnetic nickel colloidal nanocrystal clusters with antibacterial activity and bacteria binding ability

NASA Astrophysics Data System (ADS)

Peng, Bo; Zhang, Xinglin; Aarts, Dirk G. A. L.; Dullens, Roel P. A.

2018-06-01

Recent progress in synthetic nanotechnology and the ancient use of metals in food preservation and the antibacterial treatment of wounds have prompted the development of nanometallic materials for antimicrobial applications1-4. However, the materials designed so far do not simultaneously display antimicrobial activity and the capability of binding and capturing bacteria and spores. Here, we develop a one-step pyrolysis procedure to synthesize monodisperse superparamagnetic nickel colloidal nanocrystal clusters (SNCNCs), which show both antibacterial activity and the ability to bind Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacteria, as well as bacterial spores. The SNCNCs are formed from a rapid burst of nickel nanoparticles, which self-assemble slowly into clusters. The clusters can magnetically extract 99.99% of bacteria and spores and provide a promising approach for the removal of microbes, including hard-to-treat microorganisms. We believe that our work illustrates the exciting opportunities that nanotechnology offers for alternative antimicrobial strategies and other applications in microbiology.

Location of alkali metal binding sites in endothelin A selective receptor antagonists, cyclo(D-Trp-D-Asp-Pro-D-Val-Leu) and cyclo(D-Trp-D-Asp-Pro-D-Ile-Leu), from multistep collisionally activated decompositions.

PubMed

Ngoka, L C; Gross, M L

2000-02-01

We previously showed by using mass spectrometry that endothelin A selective receptor antagonists BQ123 and JKC301 form novel coordination compounds with sodium ions. This property may underlie the ability of an ET(A) antagonist to induce net tubular sodium reabsorption in the proximal tubule cells and reverse acute renal failure induced by severe ischemia. We have now defined the metal binding sites on BQ123 and JKC301 by subjecting the metal-containing peptides to multiple stages of collisionally activated decomposition (CAD) in an ion trap mass spectrometer. When submitted to low-energy CAD, the ring opens at the Asp-Pro amide bond. The metal ion, which bonds, inter alia, to the carbonyl oxygen of the proline residue, acts as a fixed charge site, and directs a charge-remote, sequence-specific fragmentation of the ring-opened peptide. Amino acid residues are sequentially cleaved from the C-terminal end, and the terminal aziridinone structure moves one step toward the N-terminus with each C-terminal amino acid residue removed. These observations are the basis of a new method to sequence cyclic peptides. Amino acid residues are observed as sets of three ions, a*(n)PD, b*(n)PD and c*(n)PD where n is the number of amino acid residues in the peptide. Copyright 2000 John Wiley & Sons, Ltd.

Trivalent metal ions based on inorganic compounds with in vitro inhibitory activity of matrix metalloproteinase 13.

PubMed

Wen, Hanyu; Qin, Yuan; Zhong, Weilong; Li, Cong; Liu, Xiang; Shen, Yehua

2016-10-01

Collagenase-3 (MMP-13) inhibitors have attracted considerable attention in recent years and have been developed as a therapeutic target for a variety of diseases, including cancer. Matrix metalloproteinases (MMPs) can be inhibited by a multitude of compounds, including hydroxamic acids. Studies have shown that materials and compounds containing trivalent metal ions, particularly potassium hexacyanoferrate (III) (K3[Fe(CN)6]), exhibit cdMMP-13 inhibitory potential with a half maximal inhibitory concentration (IC50) of 1.3μM. The target protein was obtained by refolding the recombinant histidine-tagged cdMMP-13 using size exclusion chromatography (SEC). The secondary structures of the refolded cdMMP-13 with or without metal ions were further analyzed via circular dichroism and the results indicate that upon binding with metal ions, an altered structure with increased domain stability was obtained. Furthermore, isothermal titration calorimetry (ITC) experiments demonstrated that K3[Fe(CN)6]is able to bind to MMP-13 and endothelial cell tube formation tests provide further evidence for this interaction to exhibit anti-angiogenesis potential. To the best of our knowledge, no previous report of an inorganic compound featuring a MMP-13 inhibitory activity has ever been reported in the literature. Our results demonstrate that K3[Fe(CN)6] is useful as a new effective and specific inhibitor for cdMMP-13 which may be of great potential for future drug screening applications. Copyright © 2016. Published by Elsevier Inc.

The Structure of the Metal Transporter Tp34 and its Affinity for Divalent Metal Ions

NASA Astrophysics Data System (ADS)

Knutsen, Gregory; Deka, Ranjit; Brautigam, Chad; Tomchick, Diana; Machius, Mischa; Norgard, Michael

2007-10-01

Tp34 is periplasmic membrane protein of the nonculitvatable spirochete Treponema pallidum, the pathogen of syphillis. It was proposed that Tp34 is a divalent metal transporter, but the identity of the preferred metal ion(s) was unclear. In this study we investigated the ability of divalent metal ions to induce rTp34 dimerization using hydrodynamic techniques and determine the crystal structure of metal bound forms. Using analytical ultracentrifugation sedimentation velocity experiments, we determined that cobalt is superior to nickel at inducing the dimerization of rTp34. rTp34 was crystallized and selected crystals were incubated at a pH 7.5 with CuSO4 and NiSO4. Diffraction experiments were conducted and the processed electron density maps showed that copper was bound to the major metal binding site as well as to three additional minor binding sites. By contrast nickel was only bound to the major metal binding site in one monomer and to three additional minor sites. These results along with previous findings support evidence of Tp34 being involved with metal transport and/or iron utilization.

Metal Ion Interactions with Immunoglobulin G (IgG). 1. Preliminary Studies with Electron Paramagnetic Resonance (EPR) Spectroscopy and Ultrafiltration

DTIC Science & Technology

1978-12-12

EPR and ultrafiltration studies are recommceided to conduct luture metal ion- IgG binding research. Using Scatchard plots, bind.ng levels can be...of the binding sites can be best pursued by EPR and ultrafiltration using the fragments of IgG . This report noted some difference in the binding...immunoelectrophoresis, ultrafiltration, UV spectroscopy, atomic absorption spectroscopy, and electron paramagnetic resonance (EPR). IgG used ,- ,is non

Multiheteromacrocycles that Complex Metal Ions. Sixth Progress Report, 1 May 1979-30 April 1980

DOE R&D Accomplishments Database

Cram, D. J.

1980-01-15

Objective is to design synthesize, and evaluate cyclic and polycyclic host organic compounds for their abilities to complex and lipophilize guest metal ions, their complexes, and their clusters. Host organic compounds consist of strategically placed solvating, coordinating, and ion-pairing sites tied together by covalent bonds through hydrocarbon units around cavities shaped to be occupied by guest metal ions or by metal ions plus their ligands. Specificity in complexation is sought by matching the following properties of host and guest: cavity and metal ion sizes; geometric arrangements of binding sites; number of binding sites; character of binding sites; and valences. During this period, hemispherands based on an aryloxy or cyclic urea unit, spherands based on aryloxyl units only, and their complexes with alkali metals and alkaline earths were investigated. An attempt to separate {sup 6}Li and {sup 7}Li by gel permeation chromatography of lithiospherium chloride failed. (DLC)

Studies with an immobilized metal affinity chromatography cassette system involving binuclear triazacyclononane-derived ligands: automation of batch adsorption measurements with tagged recombinant proteins.

PubMed

Petzold, Martin; Coghlan, Campbell J; Hearn, Milton T W

2014-07-18

This study describes the determination of the adsorption isotherms and binding kinetics of tagged recombinant proteins using a recently developed IMAC cassette system and employing automated robotic liquid handling procedures for IMAC resin screening. These results confirm that these new IMAC resins, generated from a variety of different metal-charged binuclear 1,4,7-triaza-cyclononane (tacn) ligands, interact with recombinant proteins containing a novel N-terminal metal binding tag, NT1A, with static binding capacities similar to those obtained with conventional hexa-His tagged proteins, but with significantly increased association constants. In addition, higher kinetic binding rates were observed with these new IMAC systems, an attribute that can be positively exploited to increase process productivity. The results from this investigation demonstrate that enhancements in binding capacities and affinities were achieved with these new IMAC resins and chosen NT1A tagged protein. Further, differences in the binding performances of the bis(tacn) xylenyl-bridged ligands were consistent with the distance between the metal binding centres of the two tacn moieties, the flexibility of the ligand and the potential contribution from the aromatic ring of the xylenyl group to undergo π/π stacking interactions with the tagged proteins. Copyright © 2014 Elsevier B.V. All rights reserved.

Structural characterization of metal binding to a cold-adapted frataxin.

PubMed

Noguera, Martín E; Roman, Ernesto A; Rigal, Juan B; Cousido-Siah, Alexandra; Mitschler, André; Podjarny, Alberto; Santos, Javier

2015-06-01

Frataxin is an evolutionary conserved protein that participates in iron metabolism. Deficiency of this small protein in humans causes a severe neurodegenerative disease known as Friedreich's ataxia. A number of studies indicate that frataxin binds iron and regulates Fe-S cluster biosynthesis. Previous structural studies showed that metal binding occurs mainly in a region of high density of negative charge. However, a comprehensive characterization of the binding sites is required to gain further insights into the mechanistic details of frataxin function. In this work, we have solved the X-ray crystal structures of a cold-adapted frataxin from a psychrophilic bacterium in the presence of cobalt or europium ions. We have identified a number of metal-binding sites, mainly solvent exposed, several of which had not been observed in previous studies on mesophilic homologues. No major structural changes were detected upon metal binding, although the structures exhibit significant changes in crystallographic B-factors. The analysis of these B-factors, in combination with crystal packing and RMSD among structures, suggests the existence of localized changes in the internal motions. Based on these results, we propose that bacterial frataxins possess binding sites of moderate affinity for a quick capture and transfer of iron to other proteins and for the regulation of Fe-S cluster biosynthesis, modulating interactions with partner proteins.

Computational Chemistry-Based Identification of Ultra-Low Temperature Water-Gas-Shift Catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Manos Mavrikakis

2008-08-31

The current work seeks to identify novel, catalytically-active, stable, poison-resistant LWGS catalysts that retain the superior activity typical of conventional Cu catalysts but can be operated at similar or lower temperatures. A database for the Binding Energies (BEs) of the LWGS relevant species, namely CO, O and OH on the most-stable, close-packed facets of a set of 17 catalytically relevant transition metals was established. This BE data and a database of previously established segregation energies was utilized to predict the stability of bimetallic NSAs that could be synthesized by combinations of the 17 parent transition metals. NSAs that were potentiallymore » stable both in vacuo and under the influence of strong-binding WGS intermediates were then selected for adsorption studies. A set of 40 NSAs were identified that satisfied all three screener criteria and the binding energies of CO, O and OH were calculated on a set of 66, 43 and 79 NSA candidates respectively. Several NSAs were found that bound intermediates weaker than the monometallic catalysts and were thus potentially poison-resistant. Finally, kinetic studies were performed and resulted in the discovery of a specific NSA-based bimetallic catalyst Cu/Pt that is potentially a promising LWGS catalyst. This stable Cu/Pt subsurface alloy is expected to provide facile H{sub 2}O activation and remain relatively resistant from the poisoning by CO, S and formate intermediates.« less

Computational design and elaboration of a de novo heterotetrameric alpha-helical protein that selectively binds an emissive abiological (porphinato)zinc chromophore.

PubMed

Fry, H Christopher; Lehmann, Andreas; Saven, Jeffery G; DeGrado, William F; Therien, Michael J

2010-03-24

The first example of a computationally de novo designed protein that binds an emissive abiological chromophore is presented, in which a sophisticated level of cofactor discrimination is pre-engineered. This heterotetrameric, C(2)-symmetric bundle, A(His):B(Thr), uniquely binds (5,15-di[(4-carboxymethyleneoxy)phenyl]porphinato)zinc [(DPP)Zn] via histidine coordination and complementary noncovalent interactions. The A(2)B(2) heterotetrameric protein reflects ligand-directed elements of both positive and negative design, including hydrogen bonds to second-shell ligands. Experimental support for the appropriate formulation of [(DPP)Zn:A(His):B(Thr)](2) is provided by UV/visible and circular dichroism spectroscopies, size exclusion chromatography, and analytical ultracentrifugation. Time-resolved transient absorption and fluorescence spectroscopic data reveal classic excited-state singlet and triplet PZn photophysics for the A(His):B(Thr):(DPP)Zn protein (k(fluorescence) = 4 x 10(8) s(-1); tau(triplet) = 5 ms). The A(2)B(2) apoprotein has immeasurably low binding affinities for related [porphinato]metal chromophores that include a (DPP)Fe(III) cofactor and the zinc metal ion hemin derivative [(PPIX)Zn], underscoring the exquisite active-site binding discrimination realized in this computationally designed protein. Importantly, elements of design in the A(His):B(Thr) protein ensure that interactions within the tetra-alpha-helical bundle are such that only the heterotetramer is stable in solution; corresponding homomeric bundles present unfavorable ligand-binding environments and thus preclude protein structural rearrangements that could lead to binding of (porphinato)iron cofactors.

Evidence for Posttranslational Protein Flavinylation in the Syphilis Spirochete Treponema pallidum: Structural and Biochemical Insights from the Catalytic Core of a Periplasmic Flavin-Trafficking Protein

PubMed Central

Deka, Ranjit K.; Brautigam, Chad A.; Liu, Wei Z.

2015-01-01

ABSTRACT The syphilis spirochete Treponema pallidum is an important human pathogen but a highly enigmatic bacterium that cannot be cultivated in vitro. T. pallidum lacks many biosynthetic pathways and therefore has evolved the capability to exploit host-derived metabolites via its periplasmic lipoprotein repertoire. We recently reported a flavin-trafficking protein in T. pallidum (Ftp_Tp; TP0796) as the first bacterial metal-dependent flavin adenine dinucleotide (FAD) pyrophosphatase that hydrolyzes FAD into AMP and flavin mononucleotide (FMN) in the spirochete’s periplasm. However, orthologs of Ftp_Tp from other bacteria appear to lack this hydrolytic activity; rather, they bind and flavinylate subunits of a cytoplasmic membrane redox system (Nqr/Rnf). To further explore this dichotomy, biochemical analyses, protein crystallography, and structure-based mutagenesis were used to show that a single amino acid change (N55Y) in Ftp_Tp converts it from an Mg2+-dependent FAD pyrophosphatase to an FAD-binding protein. We also demonstrated that Ftp_Tp has a second enzymatic activity (Mg2+-FMN transferase); it flavinylates protein(s) covalently with FMN on a threonine side chain of an appropriate sequence motif using FAD as the substrate. Moreover, mutation of a metal-binding residue (D284A) eliminates Ftp_Tp’s dual activities, thereby underscoring the role of Mg2+ in the enzyme-catalyzed reactions. The posttranslational flavinylation activity that can target a periplasmic lipoprotein (TP0171) has not previously been described. The observed activities reveal the catalytic flexibility of a treponemal protein to perform multiple functions. Together, these findings imply mechanisms by which a dynamic pool of flavin cofactor is maintained and how flavoproteins are generated by Ftp_Tp locally in the T. pallidum periplasm. PMID:25944861

First-second shell interactions in metal binding sites in proteins: a PDB survey and DFT/CDM calculations.

PubMed

Dudev, Todor; Lin, Yen-lin; Dudev, Minko; Lim, Carmay

2003-03-12

The role of the second shell in the process of metal binding and selectivity in metalloproteins has been elucidated by combining Protein Data Bank (PDB) surveys of Mg, Mn, Ca, and Zn binding sites with density functional theory/continuum dielectric methods (DFT/CDM). Peptide backbone groups were found to be the most common second-shell ligand in Mg, Mn, Ca, and Zn binding sites, followed (in decreasing order) by Asp/Glu, Lys/Arg, Asn/Gln, and Ser/Thr side chains. Aromatic oxygen- or nitrogen-containing side chains (Tyr, His, and Trp) and sulfur-containing side chains (Cys and Met) are seldom found in the second coordination layer. The backbone and Asn/Gln side chain are ubiquitous in the metal second coordination layer as their carbonyl oxygen and amide hydrogen can act as a hydrogen-bond acceptor and donor, respectively, and can therefore partner practically every first-shell ligand. The second most common outer-shell ligand, Asp/Glu, predominantly hydrogen bonds to a metal-bound water or Zn-bound histidine and polarizes the H-O or H-N bond. In certain cases, a second-shell Asp/Glu could affect the protonation state of the metal ligand. It could also energetically stabilize a positively charged metal complex more than a neutral ligand such as the backbone and Asn/Gln side chain. As for the first shell, the second shell is predicted to contribute to the metal selectivity of the binding site by discriminating between metal cations of different ionic radii and coordination geometries. The first-shell-second-shell interaction energies decay rapidly with increasing solvent exposure of the metal binding site. They are less favorable but are of the same order of magnitude as compared to the respective metal-first-shell interaction energies. Altogether, the results indicate that the structure and properties of the second shell are dictated by those of the first layer. The outer shell is apparently designed to stabilize/protect the inner-shell and complement/enhance its properties.

Copper speciation and binding by organic matter in copper-contaminated streamwater

USGS Publications Warehouse

Breault, R.F.; Colman, J.A.; Aiken, G.R.; McKnight, D.

1996-01-01

Fulvic acid binding sites (1.3-70 ??M) and EDTA (0.0017-0.18 ??M) accounted for organically bound Cu in seven stream samples measured by potentiometric titration. Cu was 84-99% organically bound in filtrates with 200 nM total Cu. Binding of Cu by EDTA was limited by competition from other trace metals. Water hardness was inversely related to properties of dissolved organic carbon (DOC) that enhance fulvic acid binding: DOC concentration, percentage of DOC that is fulvic acid, and binding sites per fulvic acid carbon. Dissolved trace metals, stabilized by organic binding, occurred at increased concentration in soft water as compared to hard water.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang Huayan; Yu Junping; Fu Guo

The interaction between integrin macrophage differentiation antigen associated with complement three receptor function (Mac-1) and intercellular adhesion molecule-1 (ICAM-1), which is controlled tightly by the ligand-binding activity of Mac-1, is central to the regulation of neutrophil adhesion in host defense. Several 'inside-out' signals and extracellular metal ions or antibodies have been found to activate Mac-1, resulting in an increased adhesiveness of Mac-1 to its ligands. However, the molecular basis for Mac-1 activation is not well understood yet. In this work, we have carried out a single-molecule study of Mac-1/ICAM-1 interaction force in living cells by atomic force microscopy (AFM). Ourmore » results showed that the binding probability and adhesion force of Mac-1 with ICAM-1 increased upon Mac-1 activation. Moreover, by comparing the dynamic force spectra of different Mac-1 mutants, we expected that Mac-1 activation is governed by the downward movement of its {alpha}7 helix.« less

Phosphorus vacancy cluster model for phosphorus diffusion gettering of metals in Si

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Renyu; Trzynadlowski, Bart; Dunham, Scott T.

2014-02-07

In this work, we develop models for the gettering of metals in silicon by high phosphorus concentration. We first performed ab initio calculations to determine favorable configurations of complexes involving phosphorus and transition metals (Fe, Cu, Cr, Ni, Ti, Mo, and W). Our ab initio calculations found that the P{sub 4}V cluster, a vacancy surrounded by 4 nearest-neighbor phosphorus atoms, which is the most favorable inactive P species in heavily doped Si, strongly binds metals such as Cu, Cr, Ni, and Fe. Based on the calculated binding energies, we build continuum models to describe the P deactivation and Fe getteringmore » processes with model parameters calibrated against experimental data. In contrast to previous models assuming metal-P{sub 1}V or metal-P{sub 2}V as the gettered species, the binding of metals to P{sub 4}V satisfactorily explains the experimentally observed strong gettering behavior at high phosphorus concentrations.« less

Characterization of a multi-metal binding biosorbent: Chemical modification and desorption studies.

PubMed

Abdolali, Atefeh; Ngo, Huu Hao; Guo, Wenshan; Zhou, John L; Du, Bin; Wei, Qin; Wang, Xiaochang C; Nguyen, Phuoc Dan

2015-10-01

This work attends to preparation and characterization of a novel multi-metal binding biosorbent after chemical modification and desorption studies. Biomass is a combination of tea waste, maple leaves and mandarin peels with a certain proportion to adsorb cadmium, copper, lead and zinc ions from aqueous solutions. The mechanism involved in metal removal was investigated by SEM, SEM/EDS and FTIR. SEM/EDS showed the presence of different chemicals and adsorbed heavy metal ions on the surface of biosorbent. FTIR of both unmodified and modified biosorbents revealed the important role of carboxylate groups in heavy metal biosorption. Desorption using different eluents and 0.1 M HCl showed the best desorption performance. The effectiveness of regeneration step by 1 M CaCl2 on five successive cycles of sorption and desorption displays this multi-metal binding biosorbent (MMBB) can effectively be utilized as an adsorbent to remove heavy metal ions from aqueous solutions in five cycles of sorption/desorption/regeneration. Copyright © 2015 Elsevier Ltd. All rights reserved.

Application of SEC-ICP-MS for comparative analyses of metal-containing species in cancerous and healthy human thyroid samples.

PubMed

Boulyga, Sergei F; Loreti, Valeria; Bettmer, Jörg; Heumann, Klaus G

2004-09-01

Size exclusion chromatography (SEC) was coupled on-line to inductively coupled plasma mass spectrometry (ICP-MS) for speciation study of trace metals in cancerous thyroid tissues in comparison to healthy thyroids aimed to estimation of changes in metalloprotein speciation in pathological tissue. The study showed a presence of species binding Cu, Zn, Cd and Pb in healthy thyroid tissue with a good reproducibility of chromatographic results, whereas the same species could not be detected in cancerous tissues. Thus, remarkable differences with respect to metal-binding species were revealed between healthy and pathological thyroid samples, pointing out a completely different distribution of trace metals in cancerous tissues. The metal-binding species could not be identified in the frame of this work because of a lack of appropriate standards. Nevertheless, the results obtained confirm the suitability of SEC-ICP-MS for monitoring of changes in trace metal distribution in cancerous tissue and will help to better understand the role of metal-containing species in thyroid pathology.

Insights into the Phosphoryl Transfer Catalyzed by cAMP-Dependent Protein Kinase: An X-ray Crystallographic Study of Complexes with Various Metals and Peptide Substrate SP20

PubMed Central

2013-01-01

X-ray structures of several ternary substrate and product complexes of the catalytic subunit of cAMP-dependent protein kinase (PKAc) have been determined with different bound metal ions. In the PKAc complexes, Mg2+, Ca2+, Sr2+, and Ba2+ metal ions could bind to the active site and facilitate the phosphoryl transfer reaction. ATP and a substrate peptide (SP20) were modified, and the reaction products ADP and the phosphorylated peptide were found trapped in the enzyme active site. Finally, we determined the structure of a pseudo-Michaelis complex containing Mg2+, nonhydrolyzable AMP-PCP (β,γ-methyleneadenosine 5′-triphosphate) and SP20. The product structures together with the pseudo-Michaelis complex provide snapshots of different stages of the phosphorylation reaction. Comparison of these structures reveals conformational, coordination, and hydrogen bonding changes that might occur during the reaction and shed new light on its mechanism, roles of metals, and active site residues. PMID:23672593

Gene expression differences between Noccaea caerulescens ecotypes help to identify candidate genes for metal phytoremediation.

PubMed

Halimaa, Pauliina; Lin, Ya-Fen; Ahonen, Viivi H; Blande, Daniel; Clemens, Stephan; Gyenesei, Attila; Häikiö, Elina; Kärenlampi, Sirpa O; Laiho, Asta; Aarts, Mark G M; Pursiheimo, Juha-Pekka; Schat, Henk; Schmidt, Holger; Tuomainen, Marjo H; Tervahauta, Arja I

2014-03-18

Populations of Noccaea caerulescens show tremendous differences in their capacity to hyperaccumulate and hypertolerate metals. To explore the differences that could contribute to these traits, we undertook SOLiD high-throughput sequencing of the root transcriptomes of three phenotypically well-characterized N. caerulescens accessions, i.e., Ganges, La Calamine, and Monte Prinzera. Genes with possible contribution to zinc, cadmium, and nickel hyperaccumulation and hypertolerance were predicted. The most significant differences between the accessions were related to metal ion (di-, trivalent inorganic cation) transmembrane transporter activity, iron and calcium ion binding, (inorganic) anion transmembrane transporter activity, and antioxidant activity. Analysis of correlation between the expression profile of each gene and the metal-related characteristics of the accessions disclosed both previously characterized (HMA4, HMA3) and new candidate genes (e.g., for nickel IRT1, ZIP10, and PDF2.3) as possible contributors to the hyperaccumulation/tolerance phenotype. A number of unknown Noccaea-specific transcripts also showed correlation with Zn(2+), Cd(2+), or Ni(2+) hyperaccumulation/tolerance. This study shows that N. caerulescens populations have evolved great diversity in the expression of metal-related genes, facilitating adaptation to various metalliferous soils. The information will be helpful in the development of improved plants for metal phytoremediation.

Magnesium-dependent RNA binding to the PA endonuclease domain of the avian influenza polymerase.

PubMed

Xiao, Shiyan; Klein, Michael L; LeBard, David N; Levine, Benjamin G; Liang, Haojun; MacDermaid, Christopher M; Alfonso-Prieto, Mercedes

2014-01-30

Influenza A viruses are highly pathogenic and pose an unpredictable public health danger to humans. An attractive target for developing new antiviral drugs is the PA N-terminal domain (PAN) of influenza polymerase, which is responsible for the endonuclease activity and essential for viral replication. Recently, the crystal structures of the holo form of PAN as well as PAN bound to different inhibitors have been reported, but the potency and selectivity of these inhibitors still need to be improved. New drug design can be guided by a better understanding of the endonuclease activity of PAN. However, this requires the structure of PAN in complex with the host mRNA, which has not been determined yet. In particular, divalent metal ions are known to be essential for RNA cleavage, but it is not clear whether there is either one or two Mg ions in the PAN active site. In the present work, we have modeled the complex of the PAN endonuclease domain with the host mRNA in the presence of either one or two Mg(2+) by using all-atom molecular dynamics. These simulations identify crucial interactions between the enzyme and the nucleic acid. Moreover, they validate a previous hypothesis that a second metal ion binds in the presence of the RNA substrate and therefore support a two-metal ion mechanism, in which K134 decreases the pKa of the nucleophilic water. Nevertheless, at low Mg concentrations an alternative, one-metal ion mechanism is possible, with K137 as the catalytic lysine and H41 as the general base, rationalizing previous unexpected mutagenesis results. The RNA-enzyme interactions determined here could likely be used to design more specific endonuclease inhibitors to fight influenza viral infections.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chakraborty, Saumen; Touw, Debra S.; Peacock, Anna F.A.

Over the past two decades, designed metallopeptides have held the promise for understanding a variety of fundamental questions in metallobiochemistry; however, these dreams have not yet been realized because of a lack of structural data to elaborate the protein scaffolds before metal complexation and the resultant metalated structures which ultimately exist. This is because there are few reports of structural characterization of such systems either in their metalated or nonmetalated forms and no examples where an apo structure and the corresponding metalated peptide assembly have both been defined by X-ray crystallography. Herein we present X-ray structures of two de novomore » designed parallel three-stranded coiled coils (designed using the heptad repeat (a {yields} g)) CSL9C (CS = Coil Ser) and CSL19C in their nonmetalated forms, determined to 1.36 and 2.15 {angstrom} resolutions, respectively. Leucines from either position 9 (a site) or 19 (d site) are replaced by cysteine to generate the constructs CSL9C and CSL19C, respectively, yielding thiol-rich pockets at the hydrophobic interior of these peptides, suitable to bind heavy metals such as As(III), Hg(II), Cd(II), and Pb(II). We use these structures to understand the inherent structural differences between a and d sites to clarify the basis of the observed differential spectroscopic behavior of metal binding in these types of peptides. Cys side chains of (CSL9C){sub 3} show alternate conformations and are partially preorganized for metal binding, whereas cysteines in (CSL19C){sub 3} are present as a single conformer. Zn(II) ions, which do not coordinate or influence Cys residues at the designed metal sites but are essential for forming X-ray quality crystals, are bound to His and Glu residues at the crystal packing interfaces of both structures. These 'apo' structures are used to clarify the changes in metal site organization between metalated As(CSL9C){sub 3} and to speculate on the differential basis of Hg(II) binding in a versus d peptides. Thus, for the first time, one can establish general rules for heavy metal binding to Cys-rich sites in designed proteins which may provide insight for understanding how heavy metals bind to metallochaperones or metalloregulatory proteins.« less

A Study on Spectro-Analytical Aspects, DNA - Interaction, Photo-Cleavage, Radical Scavenging, Cytotoxic Activities, Antibacterial and Docking Properties of 3 - (1 - (6 - methoxybenzo [d] thiazol - 2 - ylimino) ethyl) - 6 - methyl - 3H - pyran - 2, 4 - dione and its Metal Complexes.

PubMed

Ravi, Mudavath; Chennam, Kishan Prasad; Ushaiah, B; Eslavath, Ravi Kumar; Perugu, Shyam; Ajumeera, Rajanna; Devi, Ch Sarala

2015-09-01

The focus of the present work is on the design, synthesis, characterization, DNA-interaction, photo-cleavage, radical scavenging, in-vitro cytotoxicity, antimicrobial, docking and kinetic studies of Cu (II), Cd (II), Ce (IV) and Zr (IV) metal complexes of an imine derivative, 3 - (1 - (6 - methoxybenzo [d] thiazol - 2 - ylimino) ethyl) - 6 - methyl - 3H - pyran - 2, 4 - dione. The investigation of metal ligand interactions for the determination of composition of metal complexes, corresponding kinetic studies and antioxidant activity in solution was carried out by spectrophotometric methods. The synthesized metal complexes were characterized by EDX analysis, Mass, IR, (1)H-NMR, (13)C-NMR and UV-Visible spectra. DNA binding studies of metal complexes with Calf thymus (CT) DNA were carried out at room temperature by employing UV-Vis electron absorption, fluorescence emission and viscosity measurement techniques. The results revealed that these complexes interact with DNA through intercalation. The results of in vitro antibacterial studies showed the enhanced activity of chelating agent in metal chelated form and thus inferring scope for further development of new therapeutic drugs. Cell viability experiments indicated that all complexes showed significant dose dependent cytotoxicity in selected cell lines. The molecular modeling and docking studies were carried out with energy minimized structures of metal complexes to identify the receptor to metal interactions.

Crystal structure of glucose isomerase in complex with xylitol inhibitor in one metal binding mode.

PubMed

Bae, Ji-Eun; Kim, In Jung; Nam, Ki Hyun

2017-11-04

Glucose isomerase (GI) is an intramolecular oxidoreductase that interconverts aldoses and ketoses. These characteristics are widely used in the food, detergent, and pharmaceutical industries. In order to obtain an efficient GI, identification of novel GI genes and substrate binding/inhibition have been studied. Xylitol is a well-known inhibitor of GI. In Streptomyces rubiginosus, two crystal structures have been reported for GI in complex with xylitol inhibitor. However, a structural comparison showed that xylitol can have variable conformation at the substrate binding site, e.g., a nonspecific binding mode. In this study, we report the crystal structure of S. rubiginosus GI in a complex with xylitol and glycerol. Our crystal structure showed one metal binding mode in GI, which we presumed to represent the inactive form of the GI. The metal ion was found only at the M1 site, which was involved in substrate binding, and was not present at the M2 site, which was involved in catalytic function. The O 2 and O 4 atoms of xylitol molecules contributed to the stable octahedral coordination of the metal in M1. Although there was no metal at the M2 site, no large conformational change was observed for the conserved residues coordinating M2. Our structural analysis showed that the metal at the M2 site was not important when a xylitol inhibitor was bound to the M1 site in GI. Thus, these findings provided important information for elucidation or engineering of GI functions. Copyright © 2017 Elsevier Inc. All rights reserved.

Non-Native Metal Ion Reveals the Role of Electrostatics in Synaptotagmin 1-Membrane Interactions.

PubMed

Katti, Sachin; Nyenhuis, Sarah B; Her, Bin; Srivastava, Atul K; Taylor, Alexander B; Hart, P John; Cafiso, David S; Igumenova, Tatyana I

2017-06-27

C2 domains are independently folded modules that often target their host proteins to anionic membranes in a Ca 2+ -dependent manner. In these cases, membrane association is triggered by Ca 2+ binding to the negatively charged loop region of the C2 domain. Here, we used a non-native metal ion, Cd 2+ , in lieu of Ca 2+ to gain insight into the contributions made by long-range Coulombic interactions and direct metal ion-lipid bridging to membrane binding. Using X-ray crystallography, NMR, Förster resonance energy transfer, and vesicle cosedimentation assays, we demonstrate that, although Cd 2+ binds to the loop region of C2A/B domains of synaptotagmin 1 with high affinity, long-range Coulombic interactions are too weak to support membrane binding of individual domains. We attribute this behavior to two factors: the stoichiometry of Cd 2+ binding to the loop regions of the C2A and C2B domains and the impaired ability of Cd 2+ to directly coordinate the lipids. In contrast, electron paramagnetic resonance experiments revealed that Cd 2+ does support membrane binding of the C2 domains in full-length synaptotagmin 1, where the high local lipid concentrations that result from membrane tethering can partially compensate for lack of a full complement of divalent metal ions and specific lipid coordination in Cd 2+ -complexed C2A/B domains. Our data suggest that long-range Coulombic interactions alone can drive the initial association of C2A/B with anionic membranes and that Ca 2+ further augments membrane binding by the formation of metal ion-lipid coordination bonds and additional Ca 2+ ion binding to the C2 domain loop regions.

Au-Interaction of Slp1 Polymers and Monolayer from Lysinibacillus sphaericus JG-B53 - QCM-D, ICP-MS and AFM as Tools for Biomolecule-metal Studies

PubMed Central

Suhr, Matthias; Raff, Johannes; Pollmann, Katrin

2016-01-01

In this publication the gold sorption behavior of surface layer (S-layer) proteins (Slp1) of Lysinibacillus sphaericus JG-B53 is described. These biomolecules arrange in paracrystalline two-dimensional arrays on surfaces, bind metals, and are thus interesting for several biotechnical applications, such as biosorptive materials for the removal or recovery of different elements from the environment and industrial processes. The deposition of Au(0) nanoparticles on S-layers, either by S-layer directed synthesis 1 or adsorption of nanoparticles, opens new possibilities for diverse sensory applications. Although numerous studies have described the biosorptive properties of S-layers 2-5, a deeper understanding of protein-protein and protein-metal interaction still remains challenging. In the following study, inductively coupled mass spectrometry (ICP-MS) was used for the detection of metal sorption by suspended S-layers. This was correlated to measurements of quartz crystal microbalance with dissipation monitoring (QCM-D), which allows the online detection of proteinaceous monolayer formation and metal deposition, and thus, a more detailed understanding on metal binding. The ICP-MS results indicated that the binding of Au(III) to the suspended S-layer polymers is pH dependent. The maximum binding of Au(III) was obtained at pH 4.0. The QCM-D investigations enabled the detection of Au(III) sorption as well as the deposition of Au(0)-NPs in real-time during the in situ experiments. Further, this method allowed studying the influence of metal binding on the protein lattice stability of Slp1. Structural properties and protein layer stability could be visualized directly after QCM-D experiment using atomic force microscopy (AFM). In conclusion, the combination of these different methods provides a deeper understanding of metal binding by bacterial S-layer proteins in suspension or as monolayers on either bacterial cells or recrystallized surfaces. PMID:26863150

Chemistry of Marine Ligands and Siderophores

PubMed Central

Vraspir, Julia M.; Butler, Alison

2011-01-01

Marine microorganisms are presented with unique challenges to obtain essential metal ions required to survive and thrive in the ocean. The production of organic ligands to complex transition metal ions is one strategy to both facilitate uptake of specific metals, such as iron, and to mitigate the potential toxic effects of other metal ions, such as copper. A number of important trace metal ions are complexed by organic ligands in seawater, including iron, cobalt, nickel, copper, zinc, and cadmium, thus defining the speciation of these metal ions in the ocean. In the case of iron, siderophores have been identified and structurally characterized. Siderophores are low molecular weight iron-binding ligands produced by marine bacteria. Although progress has been made toward the identity of in situ iron-binding ligands, few compounds have been identified that coordinate the other trace metals. Deciphering the chemical structures and production stimuli of naturally produced organic ligands and the organisms they come from is fundamental to understanding metal speciation and bioavailability. The current evidence for marine ligands, with an emphasis on siderophores, and discussion of the importance and implications of metal-binding ligands in controlling metal speciation and cycling within the world’s oceans are presented. PMID:21141029

Catalytic "active-metal" template synthesis of [2]rotaxanes, [3]rotaxanes, and molecular shuttles, and some observations on the mechanism of the cu(i)-catalyzed azide-alkyne 1,3-cycloaddition.

PubMed

Aucagne, Vincent; Berna, José; Crowley, James D; Goldup, Stephen M; Hänni, Kevin D; Leigh, David A; Lusby, Paul J; Ronaldson, Vicki E; Slawin, Alexandra M Z; Viterisi, Aurélien; Walker, D Barney

2007-10-03

A synthetic approach to rotaxane architectures is described in which metal atoms catalyze covalent bond formation while simultaneously acting as the template for the assembly of the mechanically interlocked structure. This "active-metal" template strategy is exemplified using the Huisgen-Meldal-Fokin Cu(I)-catalyzed 1,3-cycloaddition of azides with terminal alkynes (the CuAAC "click" reaction). Coordination of Cu(I) to an endotopic pyridine-containing macrocycle allows the alkyne and azide to bind to metal atoms in such a way that the metal-mediated bond-forming reaction takes place through the cavity of the macrocycle--or macrocycles--forming a rotaxane. A variety of mono- and bidentate macrocyclic ligands are demonstrated to form [2]rotaxanes in this way, and by adding pyridine, the metal can turn over during the reaction, giving a catalytic active-metal template assembly process. Both the stoichiometric and catalytic versions of the reaction were also used to synthesize more complex two-station molecular shuttles. The dynamics of the translocation of the macrocycle by ligand exchange in these two-station shuttles could be controlled by coordination to different metal ions (rapid shuttling is observed with Cu(I), slow shuttling with Pd(II)). Under active-metal template reaction conditions that feature a high macrocycle:copper ratio, [3]rotaxanes (two macrocycles on a thread containing a single triazole ring) are also produced during the reaction. The latter observation shows that under these conditions the mechanism of the Cu(I)-catalyzed terminal alkyne-azide cycloaddition involves a reactive intermediate that features at least two metal ions.

MetalS2: a tool for the structural alignment of minimal functional sites in metal-binding proteins and nucleic acids.

PubMed

Andreini, Claudia; Cavallaro, Gabriele; Rosato, Antonio; Valasatava, Yana

2013-11-25

We developed a new software tool, MetalS(2), for the structural alignment of Minimal Functional Sites (MFSs) in metal-binding biological macromolecules. MFSs are 3D templates that describe the local environment around the metal(s) independently of the larger context of the macromolecular structure. Such local environment has a determinant role in tuning the chemical reactivity of the metal, ultimately contributing to the functional properties of the whole system. On our example data sets, MetalS(2) unveiled structural similarities that other programs for protein structure comparison do not consistently point out and overall identified a larger number of structurally similar MFSs. MetalS(2) supports the comparison of MFSs harboring different metals and/or with different nuclearity and is available both as a stand-alone program and a Web tool ( http://metalweb.cerm.unifi.it/tools/metals2/).

Catalysis by Atomic-Sized Centers: Methane Activation for Partial Oxidation and Combustion

DTIC Science & Technology

2015-07-21

example, H adsorbed alone on an oxide surface will bind to oxygen to form a hydroxide . However, if a Lewis base (e.g. any electron donor) is...that on a gold surface, which is not surprising considering the bonding character between the cluster and metal surfaces. The high mobility verifies

Iron loading site on the Fe-S cluster assembly scaffold protein is distinct from the active site.

PubMed

Rodrigues, Andria V; Kandegedara, Ashoka; Rotondo, John A; Dancis, Andrew; Stemmler, Timothy L

2015-06-01

Iron-sulfur (Fe-S) cluster containing proteins are utilized in almost every biochemical pathway. The unique redox and coordination chemistry associated with the cofactor allows these proteins to participate in a diverse set of reactions, including electron transfer, enzyme catalysis, DNA synthesis and signaling within several pathways. Due to the high reactivity of the metal, it is not surprising that biological Fe-S cluster assembly is tightly regulated within cells. In yeast, the major assembly pathway for Fe-S clusters is the mitochondrial ISC pathway. Yeast Fe-S cluster assembly is accomplished using the scaffold protein (Isu1) as the molecular foundation, with assistance from the cysteine desulfurase (Nfs1) to provide sulfur, the accessory protein (Isd11) to regulate Nfs1 activity, the yeast frataxin homologue (Yfh1) to regulate Nfs1 activity and participate in Isu1 Fe loading possibly as a chaperone, and the ferredoxin (Yah1) to provide reducing equivalents for assembly. In this report, we utilize calorimetric and spectroscopic methods to provide molecular insight into how wt-Isu1 from S. cerevisiae becomes loaded with iron. Isothermal titration calorimetry and an iron competition binding assay were developed to characterize the energetics of protein Fe(II) binding. Differential scanning calorimetry was used to identify thermodynamic characteristics of the protein in the apo state or under iron loaded conditions. Finally, X-ray absorption spectroscopy was used to characterize the electronic and structural properties of Fe(II) bound to Isu1. Current data are compared to our previous characterization of the D37A Isu1 mutant, and these suggest that when Isu1 binds Fe(II) in a manner not perturbed by the D37A substitution, and that metal binding occurs at a site distinct from the cysteine rich active site in the protein.

Effect of Zn2+ binding and enzyme active site on the transition state for RNA 2'-O-transphosphorylation interpreted through kinetic isotope effects.

PubMed

Chen, Haoyuan; Piccirilli, Joseph A; Harris, Michael E; York, Darrin M

2015-11-01

Divalent metal ions, due to their ability to stabilize high concentrations of negative charge, are important for RNA folding and catalysis. Detailed models derived from the structures and kinetics of enzymes and from computational simulations have been developed. However, in most cases the specific catalytic modes involving metal ions and their mechanistic roles and effects on transition state structures remain controversial. Valuable information about the nature of the transition state is provided by measurement of kinetic isotope effects (KIEs). However, KIEs reflect changes in all bond vibrational modes that differ between the ground state and transition state. QM calculations are therefore essential for developing structural models of the transition state and evaluating mechanistic alternatives. Herein, we present computational models for Zn2+ binding to RNA 2'O-transphosphorylation reaction models that aid in the interpretation of KIE experiments. Different Zn2+ binding modes produce distinct KIE signatures, and one binding mode involving two zinc ions is in close agreement with KIEs measured for non-enzymatic catalysis by Zn2+ aquo ions alone. Interestingly, the KIE signatures in this specific model are also very close to those in RNase A catalysis. These results allow a quantitative connection to be made between experimental KIE measurements and transition state structure and bonding, and provide insight into RNA 2'O-ransphosphorylation reactions catalyzed by metal ions and enzymes. This article is part of a Special Issue entitled: Enzyme Transition States from Theory and Experiment. Copyright © 2015. Published by Elsevier B.V.

Binding energies of benzene on coinage metal surfaces: Equal stability on different metals

NASA Astrophysics Data System (ADS)

Maaß, Friedrich; Jiang, Yingda; Liu, Wei; Tkatchenko, Alexandre; Tegeder, Petra

2018-06-01

Interfaces between organic molecules and inorganic solids adapt a prominent role in fundamental science, catalysis, molecular sensors, and molecular electronics. The molecular adsorption geometry, which is dictated by the strength of lateral and vertical interactions, determines the electronic structure of the molecule/substrate system. In this study, we investigate the binding properties of benzene on the noble metal surfaces Au(111), Ag(111), and Cu(111), respectively, using temperature-programmed desorption and first-principles calculations that account for non-locality of both electronic exchange and correlation effects. In the monolayer regime, we observed for all three systems a decrease of the binding energy with increasing coverage due to repulsive adsorbate/adsorbate interactions. Although the electronic properties of the noble metal surfaces are rather different, the binding strength of benzene on these surfaces is equal within the experimental error (accuracy of 0.05 eV), in excellent agreement with our calculations. This points toward the existence of a universal trend for the binding energy of aromatic molecules resulting from a subtle balance between Pauli repulsion and many-body van der Waals attraction.

Universal binding energy relations in metallic adhesion

NASA Technical Reports Server (NTRS)

Ferrante, J.; Smith, J. R.; Rose, J. J.

1984-01-01

Rose, Smith, and Ferrante have discovered scaling relations which map the adhesive binding energy calculated by Ferrante and Smith onto a single universal binding energy curve. These binding energies are calculated for all combinations of Al(111), Zn(0001), Mg(0001), and Na(110) in contact. The scaling involves normalizing the energy by the maximum binding energy and normalizing distances by a suitable combination of Thomas-Fermi screening lengths. Rose et al. have also found that the calculated cohesive energies of K, Ba, Cu, Mo, and Sm scale by similar simple relations, suggesting the universal relation may be more general than for the simple free electron metals for which it was derived. In addition, the scaling length was defined more generally in order to relate it to measurable physical properties. Further this universality can be extended to chemisorption. A simple and yet quite accurate prediction of a zero temperature equation of state (volume as a function of pressure for metals and alloys) is presented. Thermal expansion coefficients and melting temperatures are predicted by simple, analytic expressions, and results compare favorably with experiment for a broad range of metals.

Understanding the Role of Metal Ions in RNA Folding and Function: Lessons from RNase P, a Ribonucleoprotein Enzyme

NASA Astrophysics Data System (ADS)

Harris, Michael E.; Christian, Eric L.

There is a large and rapidly growing literature relating RNA function to metal ion identity and concentration; however, due to the complexity and large number of interactions it remains a significant experimental challenge to tie the interactions of individual ions to specific aspects of RNA function. Investigation of the ribonculeopro-tein enzyme RNase P function has assisted in defining characteristics of RNA—metal ion interactions and provided a useful model system for understanding RNA catalysis and ribonucleoprotein assembly. The goal of this chapter is to review progress in understanding the physical basis of functional metal ion interactions with P RNA and relate this progress to the development of our understanding of RNA metal ion interactions in general. The research results reviewed here encompass: (1) Determination of the contribution of divalent metal ion binding to specific aspects of enzyme function, (2) Identification of individual metal ion binding sites in P RNA and their contribution to function, and (3) The effect of protein binding on RNA—metal ion affinity.

Insight into the heavy metal binding potential of dissolved organic matter in MSW leachate using EEM quenching combined with PARAFAC analysis.

PubMed

Wu, Jun; Zhang, Hua; He, Pin-Jing; Shao, Li-Ming

2011-02-01

Dissolved organic matter (DOM) plays an important role in heavy metal migration from municipal solid waste (MSW) to aquatic environments via the leachate pathway. In this study, fluorescence excitation-emission matrix (EEM) quenching combined with parallel factor (PARAFAC) analysis was adopted to characterize the binding properties of four heavy metals (Cu, Pb, Zn and Cd) and DOM in MSW leachate. Nine leachate samples were collected from various stages of MSW management, including collection, transportation, incineration, landfill and subsequent leachate treatment. Three humic-like components and one protein-like component were identified in the MSW-derived DOM by PARAFAC. Significant differences in quenching effects were observed between components and metal ions, and a relatively consistent trend in metal quenching curves was observed among various leachate samples. Among the four heavy metals, Cu(II) titration led to fluorescence quenching of all four PARAFAC-derived components. Additionally, strong quenching effects were only observed in protein-like and fulvic acid (FA)-like components with the addition of Pb(II), which suggested that these fractions are mainly responsible for Pb(II) binding in MSW-derived DOM. Moreover, the significant quenching effects of the FA-like component by the four heavy metals revealed that the FA-like fraction in MSW-derived DOM plays an important role in heavy metal speciation; therefore, it may be useful as an indicator to assess the potential ability of heavy metal binding and migration. © 2010 Elsevier Ltd. All rights reserved.

Tiron Inhibits UVB-Induced AP-1 Binding Sites Transcriptional Activation on MMP-1 and MMP-3 Promoters by MAPK Signaling Pathway in Human Dermal Fibroblasts

PubMed Central

Zhang, Chao; Zhao, Mei; Zhang, Quan-Wu; Gao, Feng-Hou

2016-01-01

Recent research found that Tiron was an effective antioxidant that could act as the intracellular reactive oxygen species (ROS) scavenger or alleviate the acute toxic metal overload in vivo. In this study, we investigated the inhibitory effect of Tiron on matrix metalloproteinase (MMP)-1 and MMP-3 expression in human dermal fibroblast cells. Western blot and ELISA analysis revealed that Tiron inhibited ultraviolet B (UVB)-induced protein expression of MMP-1 and MMP-3. Real-time quantitative PCR confirmed that Tiron could inhibit UVB-induced mRNA expression of MMP-1 and MMP-3. Furthermore, Tiron significantly blocked UVB-induced activation of the MAPK signaling pathway and activator protein (AP)-1 in the downstream of this transduction pathway in fibroblasts. Through the AP-1 binding site mutation, it was found that Tiron could inhibit AP-1-induced upregulation of MMP-1 and MMP-3 expression through blocking AP-1 binding to the AP-1 binding sites in the MMP-1 and MMP-3 promoter region. In conclusion, Tiron may be a novel antioxidant for preventing and treating skin photoaging UV-induced. PMID:27486852

Effects of polyamines on the DNA-reactive properties of dimeric mithramycin complexed with cobalt(II): implications for anticancer therapy.

PubMed

Hou, Ming-Hon; Lu, Wen-Je; Huang, Chun-Yu; Fan, Ruey-Jane; Yuann, Jeu-Ming P

2009-06-09

Few studies have examined the effects of polyamines on the action of DNA-binding anticancer drugs. Here, a Co(II)-mediated dimeric mithramycin (Mith) complex, (Mith)(2)-Co(II), was shown to be resistant to polyamine competition toward the divalent metal ion when compared to the Fe(II)-mediated drug complexes. Surface plasmon resonance experiments demonstrated that polyamines interfered with the binding capacity and association rates of (Mith)(2)-Co(II) binding to DNA duplexes, while the dissociation rates were not affected. Although (Mith)(2)-Co(II) exhibited the highest oxidative activity under physiological conditions (pH 7.3 and 37 degrees C), polyamines (spermine in particular) inhibited the DNA cleavage activity of the (Mith)(2)-Co(II) in a concentration-dependent manner. Depletion of intracellular polyamines by methylglyoxal bis(guanylhydrazone) (MGBG) enhanced the sensitivity of A549 lung cancer cells to (Mith)(2)-Co(II), most likely due to the decreased intracellular effect of polyamines on the action of (Mith)(2)-Co(II). Our study suggests a novel method for enhancing the anticancer activity of DNA-binding metalloantibiotics through polyamine depletion.

Cadmium is a potent inhibitor of PPM phosphatases and targets the M1 binding site

PubMed Central

Pan, Chang; Liu, Hong-Da; Gong, Zheng; Yu, Xiao; Hou, Xu-Ben; Xie, Di-Dong; Zhu, Xi-Bin; Li, Hao-Wen; Tang, Jun-Yi; Xu, Yun-Fei; Yu, Jia-Qi; Zhang, Lian-Ying; Fang, Hao; Xiao, Kun-Hong; Chen, Yu-Guo; Wang, Jiang-Yun; Pang, Qi; Chen, Wei; Sun, Jin-Peng

2013-01-01

The heavy metal cadmium is a non-degradable pollutant. By screening the effects of a panel of metal ions on the phosphatase activity, we unexpectedly identified cadmium as a potent inhibitor of PPM1A and PPM1G. In contrast, low micromolar concentrations of cadmium did not inhibit PP1 or tyrosine phosphatases. Kinetic studies revealed that cadmium inhibits PPM phosphatases through the M1 metal ion binding site. In particular, the negative charged D441 in PPM1G specific recognized cadmium. Our results suggest that cadmium is likely a potent inhibitor of most PPM family members except for PHLPPs. Furthermore, we demonstrated that cadmium inhibits PPM1A-regulated MAPK signaling and PPM1G-regulated AKT signaling potently in vivo. Cadmium reversed PPM1A-induced cell cycle arrest and cadmium insensitive PPM1A mutant rescued cadmium induced cell death. Taken together, these findings provide a better understanding of the effects of the toxicity of cadmium in the contexts of human physiology and pathology. PMID:23903585

A Quantitative Raman Spectroscopic Signal for Metal–Phosphodiester Interactions in Solution†

PubMed Central

Christian, Eric L.; Anderson, Vernon E.; Carey, Paul R.; Harris, Michael E.

2011-01-01

Accurate identification and quantification of metal ion–phosphodiester interactions are essential for understanding the role of metal ions as determinants of three-dimensional folding of large RNAs and as cofactors in the active sites of both RNA and protein phosphodiesterases. Accomplishing this goal is difficult due to the dynamic and complex mixture of direct and indirect interactions formed with nucleic acids and other phosphodiesters in solution. To address this issue, Raman spectroscopy has been used to measure changes in bond vibrational energies due to metal interactions. However, the contributions of inner-sphere, H-bonding, and electrostatic interactions to the Raman spectrum of phosphoryl oxygens have not been analyzed quantitatively. Here, we report that all three forms of metal ion interaction result in attenuation of the Raman signal for the symmetric vibration of the nonbridging phosphate oxygens (νsPO2−), while only inner-sphere coordination gives rise to an apparent shift of νsPO2− to higher wavenumbers (νsPO2−M) in solution. Formation of νsPO2−M is shown to be both dependent on metal ion identity and an accurate measure of site-specific metal ion binding. In addition, the spectroscopic parameter reflecting the energetic difference between νsPO2− and νsPO2−M (ΔνM) is largely insensitive to changes in phosphodiester structure but strongly dependent on the absolute electronegativity and hardness of the interacting metal ion. Together, these studies provide strong experimental support for the use of νsPO2−M and ΔνM as general spectroscopic features for the quantitative analysis of metal binding affinity and the identification of metal ions associated with phosphodiesters in solution. PMID:20180599

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wilde, E.W.; Radway, J.C.; Santo Domingo, J.

The major objective of this project was to examine the potential of a novel hydrophilic polyurethane foam as an immobilization medium for algal, bacteria, and other types of biomass, and to test the resulting foam/biomass aggregates for their use in cleaning up waters contaminated with heavy metals, radionuclides and toxic organic compounds. Initial investigations focused on the bioremoval of heavy metals from wastewaters at SRS using immobilized algal biomass. This effort met with limited success for reasons which included interference in the binding of biomass and target metals by various non-target constituents in the wastewater, lack of an appropriate wastewatermore » at SRS for testing, and the unavailability of bioreactor systems capable of optimizing contact of target pollutants with sufficient biomass binding sites. Subsequent studies comparing algal, bacterial, fungal, and higher plant biomass demonstrated that other biomass sources were also ineffective for metal bioremoval under the test conditions. Radionuclide bioremoval using a Tc-99 source provided more promising results than the metal removal studies with the various types of biomass, and indicated that the alga Cyanidium was the best of the tested sources of biomass for this application. However, all of the biomass/foam aggregates tested were substantially inferior to a TEVA resin for removing Tc-99 in comparative testing. The authors also explored the use of hydrophilic polyurethane foam to embed Burkholderia cepacia, which is an efficient degrader of trichloroethylene (TCE), a contaminant of considerable concern at SRS and elsewhere. The embedded population proved to be incapable of growth on nutrient media, but retained respiratory activity. Lastly, the degradative capabilities of embedded G4 were examined. Phenol- or benzene-induced bacteria retained the ability to degrade TCE and benzene. The authors were successful in inducing enzyme activity after the organisms had already been embedded.« less

Heterogeneous Metal-Free Hydrogenation over Defect-Laden Hexagonal Boron Nitride

DOE PAGES

Nash, David J.; Restrepo, David T.; Parra, Natalia S.; ...

2016-12-21

Catalytic hydrogenation is an important process used for the production of everything from foods to fuels. Current heterogeneous implementations of this process utilize metals as the active species. Until recently, catalytic heterogeneous hydrogenation over a metal-free solid was unknown; implementation of such a system would eliminate the health, environmental, and economic concerns associated with metal-based catalysts. We report good hydrogenation rates and yields for a metal-free heterogeneous hydrogenation catalyst as well as its unique hydrogenation mechanism. We achieved catalytic hydrogenation of olefins over defect-laden h-BN (dh-BN) in a reactor designed to maximize the defects in h-BN sheets. Good yields (>90%)more » and turnover frequencies (6 × 10 –5–4 × 10 –3) were obtained for the hydrogenation of propene, cyclohexene, 1,1-diphenylethene, (E)- and (Z)-1,2-diphenylethene, octadecene, and benzylideneacetophenone. Temperature-programmed desorption of ethene over processed h-BN indicates the formation of a highly defective structure. Solid-state NMR (SSNMR) measurements of dh-BN with high and low propene surface coverages show four different binding modes. The introduction of defects into h-BN creates regions of electronic deficiency and excess. Density functional theory calculations show that both the alkene and hydrogen-bond order are reduced over four specific defects: boron substitution for nitrogen (B N), vacancies (V B and V N), and Stone–Wales defects. SSNMR and binding-energy calculations show that V N are most likely the catalytically active sites. Our work shows that catalytic sites can be introduced into a material previously thought to be catalytically inactive through the production of defects.« less

Heavy-metal-induced reactive oxygen species: phytotoxicity and physicochemical changes in plants.

PubMed

Shahid, Muhammad; Pourrut, Bertrand; Dumat, Camille; Nadeem, Muhammad; Aslam, Muhammad; Pinelli, Eric

2014-01-01

As a result of the industrial revolution, anthropogenic activities have enhanced there distribution of many toxic heavy metals from the earth's crust to different environmental compartments. Environmental pollution by toxic heavy metals is increasing worldwide, and poses a rising threat to both the environment and to human health.Plants are exposed to heavy metals from various sources: mining and refining of ores, fertilizer and pesticide applications, battery chemicals, disposal of solid wastes(including sewage sludge), irrigation with wastewater, vehicular exhaust emissions and adjacent industrial activity.Heavy metals induce various morphological, physiological, and biochemical dysfunctions in plants, either directly or indirectly, and cause various damaging effects. The most frequently documented and earliest consequence of heavy metal toxicity in plants cells is the overproduction of ROS. Unlike redox-active metals such as iron and copper, heavy metals (e.g, Pb, Cd, Ni, AI, Mn and Zn) cannot generate ROS directly by participating in biological redox reactions such as Haber Weiss/Fenton reactions. However, these metals induce ROS generation via different indirect mechanisms, such as stimulating the activity of NADPH oxidases, displacing essential cations from specific binding sites of enzymes and inhibiting enzymatic activities from their affinity for -SH groups on the enzyme.Under normal conditions, ROS play several essential roles in regulating the expression of different genes. Reactive oxygen species control numerous processes like the cell cycle, plant growth, abiotic stress responses, systemic signalling, programmed cell death, pathogen defence and development. Enhanced generation of these species from heavy metal toxicity deteriorates the intrinsic antioxidant defense system of cells, and causes oxidative stress. Cells with oxidative stress display various chemical,biological and physiological toxic symptoms as a result of the interaction between ROS and biomolecules. Heavy-metal-induced ROS cause lipid peroxidation, membrane dismantling and damage to DNA, protein and carbohydrates. Plants have very well-organized defense systems, consisting of enzymatic and non-enzymatic antioxidation processes. The primary defense mechanism for heavy metal detoxification is the reduced absorption of these metals into plants or their sequestration in root cells.Secondary heavy metal tolerance mechanisms include activation of antioxidant enzymes and the binding of heavy metals by phytochelatins, glutathione and amino acids. These defense systems work in combination to manage the cascades of oxidative stress and to defend plant cells from the toxic effects of ROS.In this review, we summarized the biochemiCal processes involved in the over production of ROS as an aftermath to heavy metal exposure. We also described the ROS scavenging process that is associated with the antioxidant defense machinery.Despite considerable progress in understanding the biochemistry of ROS overproduction and scavenging, we still lack in-depth studies on the parameters associated with heavy metal exclusion and tolerance capacity of plants. For example, data about the role of glutathione-glutaredoxin-thioredoxin system in ROS detoxification in plant cells are scarce. Moreover, how ROS mediate glutathionylation (redox signalling)is still not completely understood. Similarly, induction of glutathione and phytochelatins under oxidative stress is very well reported, but it is still unexplained that some studied compounds are not involved in the detoxification mechanisms. Moreover,although the role of metal transporters and gene expression is well established for a few metals and plants, much more research is needed. Eventually, when results for more metals and plants are available, the mechanism of the biochemical and genetic basis of heavy metal detoxification in plants will be better understood. Moreover, by using recently developed genetic and biotechnological tools it may be possible to produce plants that have traits desirable for imparting heavy metal tolerance.

Adsorption of acid-extractable organics from oil sands process-affected water onto biomass-based biochar: Metal content matters.

PubMed

Bhuiyan, Tazul I; Tak, Jin K; Sessarego, Sebastian; Harfield, Don; Hill, Josephine M

2017-02-01

The impact of biochar properties on acid-extractable organics (AEO) adsorption from oil sands process-affected water (OSPW) was studied. Biochar from wheat straw with the highest ash content (14%) had the highest adsorption capacity (0.59 mg/g) followed by biochar from pulp mill sludge, switchgrass, mountain pine, hemp shives, and aspen wood. The adsorption capacity had no obvious trend with surface area, total pore volume, bulk polarity and aromaticity. The large impact of metal content was consistent with the carboxylates (i.e., naphthenate species) in the OSPW binding to the metals (mainly Al and Fe) on the carbon substrate. Although the capacity of biochar is still approximately two orders of magnitude lower than that of a commercial activated carbon, confirming the property (i.e., metal content) that most influenced AEO adsorption, may allow biochar to become competitive with activated carbon after normalizing for cost, especially if this cost includes environmental impacts. Copyright © 2016 Elsevier Ltd. All rights reserved.

Adhesion of a bimetallic interface. Ph.D. Thesis - Case Western Reserve Univ.; [for Al, Mg, and Zn

NASA Technical Reports Server (NTRS)

Ferrante, J.

1978-01-01

The Hohenberg-Kohn and Kohn-Sham formalisms are used to examine binding (binding energy as a function of separation) for combinations of the simple metals Al(111), Zn(0001), Mg(0001), and Na(110) in contact. Similar metal contacts between Al, Zn, Mg, and Na are examined self-consistently in an ab initio calculation using the Kohn-Sham formalism. Crystallinity is included using the Aschroft pseudopotential via first order perturbation theory for the electron-ion interaction; and the ion-ion interaction is included exactly via a lattice sum. Binding energy was determined both in the local-density approximation and including gradient corrections to the exchange and correlation energy. Binding was found in all cases. In dissimilar metal contacts, interfacial bonding was greater than that in the weaker material predicting the possibility of metallic transfer. The nonzero position of the energy minimum in like metal contacts is explained in terms of consistency between the Ashcroft pseudopotential and the bulk charge density. Good agreement with experimental surface energies is obtained in the self-consistent calculation when nonlocal terms are included.

Solid-binding peptides: smart tools for nanobiotechnology.

PubMed

Care, Andrew; Bergquist, Peter L; Sunna, Anwar

2015-05-01

Over the past decade, solid-binding peptides (SBPs) have been used increasingly as molecular building blocks in nanobiotechnology. These peptides show selectivity and bind with high affinity to the surfaces of a diverse range of solid materials including metals, metal oxides, metal compounds, magnetic materials, semiconductors, carbon materials, polymers, and minerals. They can direct the assembly and functionalisation of materials, and have the ability to mediate the synthesis and construction of nanoparticles and complex nanostructures. As the availability of newly synthesised nanomaterials expands rapidly, so too do the potential applications for SBPs. Copyright © 2015 Elsevier Ltd. All rights reserved.

Evidence for involvement of the C-terminal domain in the dimerization of the CopY repressor protein from Enterococcus hirae

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pazehoski, Kristina O., E-mail: pazehosk@pitt.edu; Cobine, Paul A., E-mail: pac0006@auburn.edu; Winzor, Donald J.

2011-03-11

Research highlights: {yields} A metal-binding protein domain is directly involved in protein dimerization. {yields} Fusing the metal-binding domain to a monomeric protein induces dimerization. {yields} Frontal size-exclusion chromatography measures the strength of dimer interaction. {yields} Ultracentrifugation studies confirm the influence of metal binding on dimerization. -- Abstract: Metal binding to the C-terminal region of the copper-responsive repressor protein CopY is responsible for homodimerization and the regulation of the copper homeostasis pathway in Enterococcus hirae. Specific involvement of the 38 C-terminal residues of CopY in dimerization is indicated by zonal and frontal (large zone) size-exclusion chromatography studies. The studies demonstrate thatmore » the attachment of these CopY residues to the immunoglobulin-binding domain of streptococcal protein G (GB1) promotes dimerization of the monomeric protein. Although sensitivity of dimerization to removal of metal from the fusion protein is smaller than that found for CopY (as measured by ultracentrifugation studies), the demonstration that an unrelated protein (GB1) can be induced to dimerize by extending its sequence with the C-terminal portion of CopY confirms the involvement of this region in CopY homodimerization.« less

Interfacial metal and antibody recognition.

PubMed

Zhou, Tongqing; Hamer, Dean H; Hendrickson, Wayne A; Sattentau, Quentin J; Kwong, Peter D

2005-10-11

The unique ligation properties of metal ions are widely exploited by proteins, with approximately one-third of all proteins estimated to be metalloproteins. Although antibodies use various mechanisms for recognition, to our knowledge, none has ever been characterized that uses an interfacial metal. We previously described a family of CD4-reactive antibodies, the archetype being Q425. CD4:Q425 engagement does not interfere with CD4:HIV-1 gp120 envelope glycoprotein binding, but it blocks subsequent steps required for viral entry. Here, we use surface-plasmon resonance to show that Q425 requires calcium for recognition of CD4. Specifically, Q425 binding of calcium resulted in a 55,000-fold enhancement in affinity for CD4. X-ray crystallographic analyses of Q425 in the presence of Ca(2+), Ba(2+), or EDTA revealed an exposed metal-binding site, partially coordinated by five atoms contributed from four antibody complementarity-determining regions. The results suggest that Q425 recognition of CD4 involves direct ligation of antigen by the Q425-held calcium, with calcium binding each ligating atom of CD4 with approximately 1.5 kcal/mol of binding energy. This energetic contribution, which is greater than that from a typical protein atom, demonstrates how interfacial metal ligation can play a unique role in antigen recognition.

Interfacial metal and antibody recognition

PubMed Central

Zhou, Tongqing; Hamer, Dean H.; Hendrickson, Wayne A.; Sattentau, Quentin J.; Kwong, Peter D.

2005-01-01

The unique ligation properties of metal ions are widely exploited by proteins, with approximately one-third of all proteins estimated to be metalloproteins. Although antibodies use various mechanisms for recognition, to our knowledge, none has ever been characterized that uses an interfacial metal. We previously described a family of CD4-reactive antibodies, the archetype being Q425. CD4:Q425 engagement does not interfere with CD4:HIV-1 gp120 envelope glycoprotein binding, but it blocks subsequent steps required for viral entry. Here, we use surface-plasmon resonance to show that Q425 requires calcium for recognition of CD4. Specifically, Q425 binding of calcium resulted in a 55,000-fold enhancement in affinity for CD4. X-ray crystallographic analyses of Q425 in the presence of Ca2+, Ba2+, or EDTA revealed an exposed metal-binding site, partially coordinated by five atoms contributed from four antibody complementarity-determining regions. The results suggest that Q425 recognition of CD4 involves direct ligation of antigen by the Q425-held calcium, with calcium binding each ligating atom of CD4 with ≈1.5 kcal/mol of binding energy. This energetic contribution, which is greater than that from a typical protein atom, demonstrates how interfacial metal ligation can play a unique role in antigen recognition. PMID:16195378

DOE Office of Scientific and Technical Information (OSTI.GOV)

Myneni, Satish C. B.; Fein, Jeremy; Mishra, Bhoopesh

Bacteria are ubiquitous in a wide-range of low temperature aqueous systems, and can strongly affect the distribution and transport of metals and radionuclides in the environment. However, the role of metal adsorption onto bacteria, via the reactive cell wall functional groups, has been largely overlooked. Previous macroscale metal sorption, and XAS studies have shown that carboxyl and phosphoryl functional groups to be the important metal binding groups on bacterial cell walls and the sulfhydryl groups were not considered. The goal of our investigation was to evaluate the density of the sulfhydryl sites on different bacterial cell membranes that are commonmore » to soil systems, the binding affinities of these reactive groups towards Hg, and how this binding modifies the speciation of Hg in the natural waters.« less

DNA binding of supramolecular mixed-metal complexes

NASA Astrophysics Data System (ADS)

Swavey, Shawn; Williams, Rodd L.; Fang, Zhenglai; Milkevitch, Matthew; Brewer, Karen J.

2001-10-01

The high binding affinity of cisplatin toward DNA has led to its popularity as an anticancer agent. Due to cumulative drug resistance and toxic side effects, researchers are exploring related metallodrugs. Our approach involves the use of supramolecular complexes. These mixed-metal complexes incorporate a reactive platinum moiety bridged by a polyazine ligand to a light absorbing metal-based chromophore. The presence of the light absorber allows excitation of these systems, opening up the possibility of photoactivation. The use of a supramolecular design allows components of the assembly to be varied to enhance device function and light absorbing properties. Aspects of our molecular design process and results on the DNA binding properties for a number of these mixed-metal complexes will be discussed.

Supramolecular control of transition metal complexes in water by a hydrophobic cavity: a bio-inspired strategy.

PubMed

Bistri, Olivia; Reinaud, Olivia

2015-03-14

Supramolecular chemistry in water is a very challenging research area. In biology, water is the universal solvent where transition metal ions play major roles in molecular recognition and catalysis. In enzymes, it participates in substrate binding and/or activation in the heart of a pocket defined by the folded protein. The association of a hydrophobic cavity with a transition metal ion is thus a very appealing strategy for controlling the metal ion properties in the very competitive water solvent. Various systems based on intrinsically water-soluble macrocyclic structures such as cyclodextrins, cucurbituryls, and metallo-cages have been reported. Others use calixarenes and resorcinarenes functionalized with hydrophilic substituents. One approach for connecting a metal complex to these cavities is to graft a ligand for metal ion binding at their edge. Early work with cyclodextrins has shown Michaelis-Menten like catalysis displaying enhanced kinetics and substrate-selectivity. Remarkable examples of regio- and stereo-selective transformation of substrates have been reported as well. Dynamic two-phase systems for transition metal catalysis have also been developed. They rely on either water-transfer of the metal complex through ligand embedment or synergistic coordination of a metal ion and substrate hosting. Another strategy consists in using metallo-cages, which provide a well-defined hydrophobic space, to stabilize metal complexes in water. When the cages can host simultaneously a substrate and a reactive metal complex, size- and regio-selective catalysis was obtained. Finally, construction of a polydentate coordination site closely interlocked with a calixarene or resorcinarene macrocycle has been shown to be a very fruitful strategy for obtaining metal complexes with remarkable hosting properties. For each of these systems, the synergism resulting from the biomimetic association of a hydrophobic cavity and a metal ion is discussed within the objective of developing new tools for either selective molecular recognition (with analytical perspectives) or performant catalysis, in water.

Quercetin-Iron Complex: Synthesis, Characterization, Antioxidant, DNA Binding, DNA Cleavage, and Antibacterial Activity Studies.

PubMed

Raza, Aun; Xu, Xiuquan; Xia, Li; Xia, Changkun; Tang, Jian; Ouyang, Zhen

2016-11-01

Quercetin-iron (II) complex was synthesized and characterized by elemental analysis, ultraviolet-visible spectrophotometry, fourier transform infrared spectroscopy, mass spectrometry, proton nuclear magnetic resonance spectroscopy, thermogravimetry and differential scanning calorimetry, scanning electron micrography and molar conductivity. The low molar conductivity value investigates the non-electrolyte nature of the complex. The elemental analysis and other physical and spectroscopic methods reveal the 1:2 stoichiometric ratio (metal:ligand) of the complex. Antioxidant study of the quercetin and its metal complex against 2, 2-di-phenyl-1-picryl hydrazyl radical showed that the complex has much more radical scavenging activity than free quercetin. The interaction of quercetin-iron (II) complex with DNA was determined using ultraviolet visible spectra, fluorescence spectra and agarose gel electrophoresis. The results showed that quercetin-iron (II) complex can intercalate moderately with DNA, quench a strong intercalator ethidium bromide and compete for the intercalative binding sites. The complex showed significant cleavage of pBR 322 DNA from supercoiled form to nicked circular form and these cleavage effects were dose-dependent. Moreover, the mechanism of DNA cleavage indicated that it was an oxidative cleavage pathway. These results revealed the potential nuclease activity of complex to cleave DNA. In addition, antibacterial activity of complex on E.coli and S. aureus was also investigated. The results showed that complex has higher antibacterial activity than ligand.

Structural Analysis of the Hg(II)-Regulatory Protein Tn501 MerR from Pseudomonas aeruginosa

NASA Astrophysics Data System (ADS)

Wang, Dan; Huang, Shanqing; Liu, Pingying; Liu, Xichun; He, Yafeng; Chen, Weizhong; Hu, Qingyuan; Wei, Tianbiao; Gan, Jianhua; Ma, Jing; Chen, Hao

2016-09-01

The metalloprotein MerR is a mercury(II)-dependent transcriptional repressor-activator that responds to mercury(II) with extraordinary sensitivity and selectivity. It’s widely distributed in both Gram-negative and Gram-positive bacteria but with barely detectable sequence identities between the two sources. To provide structural basis for the considerable biochemical and biophysical experiments previously performed on Tn501 and Tn21 MerR from Gram-negative bacteria, we analyzed the crystal structure of mercury(II)-bound Tn501 MerR. The structure in the metal-binding domain provides Tn501 MerR with a high affinity for mercury(II) and the ability to distinguish mercury(II) from other metals with its unique planar trigonal coordination geometry, which is adopted by both Gram-negative and Gram-positive bacteria. The mercury(II) coordination state in the C-terminal metal-binding domain is transmitted through the allosteric network across the dimer interface to the N-terminal DNA-binding domain. Together with the previous mutagenesis analyses, the present data indicate that the residues in the allosteric pathway have a central role in maintaining the functions of Tn501 MerR. In addition, the complex structure exhibits significant differences in tertiary and quaternary structural arrangements compared to those of Bacillus MerR from Gram-positive bacteria, which probably enable them to function with specific promoter DNA with different spacers between -35 and -10 elements.

Inhibition of ATPase activity in rat synaptic plasma membranes by simultaneous exposure to metals.

PubMed

Carfagna, M A; Ponsler, G D; Muhoberac, B B

1996-03-08

Inhibition of Na+/K+-ATPase and Mg2+-ATPase activities by in vitro exposure to Cd2+, Pb2+ and Mn2+ was investigated in rat brain synaptic plasma membranes (SPMs). Cd2+ and Pb2+ produced a larger maximal inhibition of Na+/K+-ATPase than of Mg2+-ATPase activity. Metal concentrations causing 50% inhibition of Na+/K+-ATPase activity (IC50 values) were Cd2+ (0.6 microM) < Pb2+ (2.1 microM) < Mn2+ (approximately 3 mM), and the former two metals were substantially more potent in inhibiting SPM versus synaptosomal Na+/K+-ATPase. Dixon plots of SPM data indicated that equilibrium binding of metals occurs at sites causing enzyme inhibition. In addition, IC50 values for SPM K+-dependent p-nitrophenylphosphatase inhibition followed the same order and were Cd2+ (0.4 microM) < Pb2+ (1.2 microM) < Mn2+ (300 microM). Simultaneous exposure to the combinations Cd2+/Mn2+ or Pb2+/Mn2+ inhibited SPM Na+/K+-ATPase activity synergistically (i.e., greater than the sum of the metal-induced inhibitions assayed separately), while Cd2+/Pb2+ caused additive inhibition. Simultaneous exposure to Cd2+/Pb2+ antagonistically inhibited Mg2+-ATPase activity while Cd2+/Mn2+ or Pb2+/Mn2+ additively inhibited Mg2+-ATPase activity at low Mn2+ concentrations, but inhibited antagonistically at higher concentrations. The similar IC50 values for Cd2+ and Pb2+ versus Mn2+ inhibition of Na+/K+-ATPase and the pattern of inhibition/activation upon exposure to two metals simultaneously support similar modes of interaction of Cd2+ and Pb2+ with this enzyme, in agreement with their chemical reactivities.

Supramolecular regulation of bioorthogonal catalysis in cells using nanoparticle-embedded transition metal catalysts

NASA Astrophysics Data System (ADS)

Tonga, Gulen Yesilbag; Jeong, Youngdo; Duncan, Bradley; Mizuhara, Tsukasa; Mout, Rubul; Das, Riddha; Kim, Sung Tae; Yeh, Yi-Cheun; Yan, Bo; Hou, Singyuk; Rotello, Vincent M.

2015-07-01

Bioorthogonal catalysis broadens the functional possibilities of intracellular chemistry. Effective delivery and regulation of synthetic catalytic systems in cells are challenging due to the complex intracellular environment and catalyst instability. Here, we report the fabrication of protein-sized bioorthogonal nanozymes through the encapsulation of hydrophobic transition metal catalysts into the monolayer of water-soluble gold nanoparticles. The activity of these catalysts can be reversibly controlled by binding a supramolecular cucurbit[7]uril ‘gate-keeper’ onto the monolayer surface, providing a biomimetic control mechanism that mimics the allosteric regulation of enzymes. The potential of this gated nanozyme for use in imaging and therapeutic applications was demonstrated through triggered cleavage of allylcarbamates for pro-fluorophore activation and propargyl groups for prodrug activation inside living cells.

Rational design of a conformation-switchable Ca2+- and Tb3+-binding protein without the use of multiple coupled metal-binding sites.

PubMed

Li, Shunyi; Yang, Wei; Maniccia, Anna W; Barrow, Doyle; Tjong, Harianto; Zhou, Huan-Xiang; Yang, Jenny J

2008-10-01

Ca2+, as a messenger of signal transduction, regulates numerous target molecules via Ca2+-induced conformational changes. Investigation into the determinants for Ca2+-induced conformational change is often impeded by cooperativity between multiple metal-binding sites or protein oligomerization in naturally occurring proteins. To dissect the relative contributions of key determinants for Ca2+-dependent conformational changes, we report the design of a single-site Ca2+-binding protein (CD2.trigger) created by altering charged residues at an electrostatically sensitive location on the surface of the host protein rat Cluster of Differentiation 2 (CD2).CD2.trigger binds to Tb3+ and Ca2+ with dissociation constants of 0.3 +/- 0.1 and 90 +/- 25 microM, respectively. This protein is largely unfolded in the absence of metal ions at physiological pH, but Tb3+ or Ca2+ binding results in folding of the native-like conformation. Neutralization of the charged coordination residues, either by mutation or protonation, similarly induces folding of the protein. The control of a major conformational change by a single Ca2+ ion, achieved on a protein designed without reliance on sequence similarity to known Ca2+-dependent proteins and coupled metal-binding sites, represents an important step in the design of trigger proteins.

Synthesis, spectroscopic characterization, DNA interaction and biological activities of Mn(II), Co(II), Ni(II) and Cu(II) complexes with [(1H-1,2,4-triazole-3-ylimino)methyl]naphthalene-2-ol

NASA Astrophysics Data System (ADS)

Gaber, Mohamed; El-Wakiel, Nadia A.; El-Ghamry, Hoda; Fathalla, Shaimaa K.

2014-11-01

Manganese(II), cobalt(II), nickel(II) and copper(II) complexes of [(1H-1,2,4-triazole-3-ylimino)methyl]naphthalene-2-ol have been synthesized. The structure of complexes have been characterized by elemental analysis, molar conductance, magnetic moment measurements and spectral (IR, 1H NMR, EI-mass, UV-Vis and ESR), and thermal studies. The results showed that the chloro and nitrato Cu(II) complexes have octahedral geometry while Ni(II), Co(II) and Mn(II) complexes in addition to acetato Cu(II) complex have tetrahedral geometry. The possible structures of the metal complexes have been computed using the molecular mechanic calculations using the hyper chem. 8.03 molecular modeling program to confirm the proposed structures. The kinetic and thermodynamic parameters of the thermal decomposition steps were calculated from the TG curves. The binding modes of the complexes with DNA have been investigated by UV-Vis absorption titration. The results showed that the mode of binding of the complexes to DNA is intercalative or non-intercalative binding modes. Schiff base and its metal complexes have been screened for their in vitro antimicrobial activities against Gram positive bacteria (Staphylococcus aureus), Gram negative bacteria (Escherichia coli and Pesudomonas aeruginosa), fungi (Asperigllus flavus and Mucer) and yeast (Candida albicans and Malassezia furfur).

Synthesis, characterization of 1,2,4-triazole Schiff base derived 3d-metal complexes: Induces cytotoxicity in HepG2, MCF-7 cell line, BSA binding fluorescence and DFT study

NASA Astrophysics Data System (ADS)

Tyagi, Prateek; Tyagi, Monika; Agrawal, Swati; Chandra, Sulekh; Ojha, Himanshu; Pathak, Mallika

2017-01-01

Two novel Schiff base ligands H2L1 and H2L2 have been synthesized by condensation reaction of amine derivative of 1,2,4-triazole moiety with 2-hydroxy-4-methoxybenzaldehyde. Co(II), Ni(II), Cu(II) and Zn(II) of the synthesized Schiff bases were prepared by using a molar ratio of ligand:metal as 1:1. The structure of the Schiff bases and synthesized metal complexes were established by 1H NMR, UV-Vis, IR, Mass spectrometry and molar conductivity. The thermal stability of the complexes was study by TGA. Fluorescence quenching mechanism of metal complexes 1-4 show that Zn(II) and Cu(II) complex binds more strongly to BSA. In DFT studies the geometries of Schiff bases and metal complexes were fully optimized with respect to the energy using the 6-31 + g(d,p) basis set. The spectral data shows that the ligands behaves as binegative tridentate. On the basis of the spectral studies, TGA and DFT data an octahedral geometry has been assigned for Co(II), Ni(II), square planar for Cu(II) and tetrahedral for Zn(II) complexes. The anticancer activity were screened against human breast cancer cell line (MCF-7) and human hepatocellular liver carcinoma cell line (Hep-G2). Result indicates that metal complexes shows increase cytotoxicity in proliferation to cell lines as compared to free ligand.

Multinuclear metal-binding ability of a carotene

PubMed Central

Horiuchi, Shinnosuke; Tachibana, Yuki; Yamashita, Mitsuki; Yamamoto, Koji; Masai, Kohei; Takase, Kohei; Matsutani, Teruo; Kawamata, Shiori; Kurashige, Yuki; Yanai, Takeshi; Murahashi, Tetsuro

2015-01-01

Carotenes are naturally abundant unsaturated hydrocarbon pigments, and their fascinating physical and chemical properties have been studied intensively not only for better understanding of the roles in biological processes but also for the use in artificial chemical systems. However, their metal-binding ability has been virtually unexplored. Here we report that β-carotene has the ability to assemble and align ten metal atoms to afford decanuclear homo- and heterometal chain complexes. The metallo–carotenoid framework shows reversible metalation–demetalation reactivity with multiple metals, which allows us to control the size of metal chains as well as the heterobimetallic composition and arrangement of the carotene-supported metal chains. PMID:25857402

Trace Metal-Humic Complexes in Natural Waters: Insights From Speciation Experiments

NASA Astrophysics Data System (ADS)

Stern, J. C.; Salters, V.; Sonke, J.

2006-12-01

The DOM cycle is intimately linked to the cycling and bioavailability of trace metals in aqueous environments. The presence or absence of DOM in the water column can determined whether trace elements will be present in limited quantities as a nutrient, or in surplus quantities as a toxicant. Humic substances (HS), which represent the refractory products of DOM degradation, strongly affect the speciation of trace metals in natural waters. To simulate metal-HS interactions in nature, experiments must be carried out using trace metal concentrations. Sensitive detection systems such as ICP-MS make working with small (nanomolar) concentrations possible. Capillary electrophoresis coupled with ICP-MS (CE-ICP-MS) has recently been identified as a rapid and accurate method to separate metal species and calculate conditional binding constants (log K_c) of metal-humic complexes. CE-ICP-MS was used to measure partitioning of metals between humic substances and a competing ligand (EDTA) and calculate binding constants of rare earth element (REE) and Th, Hf, and Zr-humic complexes at pH 3.5-8 and ionic strength of 0.1. Equilibrium dialysis ligand exchange (EDLE) experiments to validate the CE-ICP-MS method were performed to separate the metal-HS and metal-EDTA species by partitioning due to size exclusion via diffusion through a 1000 Da membrane. CE-ICP-MS experiments were also conducted to compare binding constants of REE with humic substances of various origin, including soil, peat, and aquatic DOM. Results of our experiments show an increase in log K_c with decrease in ionic radius for REE-humic complexes (the lanthanide contraction effect). Conditional binding constants of tetravalent metal-humic complexes were found to be several orders of magnitude higher than REE-humic complexes, indicating that tetravalent metals have a very strong affinity for humic substances. Because thorium is often used as a proxy for the tetravalent actinides, Th-HS binding constants can allow us to assess the importance of tetravalent actinide-humic complexes in groundwater transport from nuclear repositories. Our results suggest that tetravalent actinide-humic complexes couild be more important to account for in predictive speciation models than previously thought.

Synthesis, Characterization and Biological Evaluation of Transition Metal Complexes Derived from N, S Bidentate Ligands

PubMed Central

Md Yusof, Enis Nadia; Ravoof, Thahira Begum S. A.; Tiekink, Edward R. T.; Veerakumarasivam, Abhimanyu; Crouse, Karen Anne; Mohamed Tahir, Mohamed Ibrahim; Ahmad, Haslina

2015-01-01

Two bidentate NS ligands were synthesized by the condensation reaction of S-2-methylbenzyldithiocarbazate (S2MBDTC) with 2-methoxybenzaldehyde (2MB) and 3-methoxybenzaldehyde (3MB). The ligands were reacted separately with acetates of Cu(II), Ni(II) and Zn(II) yielding 1:2 (metal:ligand) complexes. The metal complexes formed were expected to have a general formula of [M(NS)2] where M = Cu2+, Ni2+, and Zn2+. These compounds were characterized by elemental analysis, molar conductivity, magnetic susceptibility and various spectroscopic techniques. The magnetic susceptibility measurements and spectral results supported the predicted coordination geometry in which the Schiff bases behaved as bidentate NS donor ligands coordinating via the azomethine nitrogen and thiolate sulfur. The molecular structures of the isomeric S2M2MBH (1) and S2M3MBH (2) were established by X-ray crystallography to have very similar l-shaped structures. The Schiff bases and their metal complexes were evaluated for their biological activities against estrogen receptor-positive (MCF-7) and estrogen receptor-negative (MDA-MB-231) breast cancer cell lines. Only the Cu(II) complexes showed marked cytotoxicity against the cancer cell lines. Both Schiff bases and other metal complexes were found to be inactive. In concordance with the cytotoxicity studies, the DNA binding studies indicated that Cu(II) complexes have a strong DNA binding affinity. PMID:25988384

A DNAzyme requiring two different metal ions at two distinct sites

PubMed Central

Zhou, Wenhu; Zhang, Yupei; Huang, Po-Jung Jimmy; Ding, Jinsong; Liu, Juewen

2016-01-01

Most previously reported RNA-cleaving DNAzymes require only a single divalent metal ion for catalysis. We recently reported a general trivalent lanthanide-dependent DNAzyme named Ce13d. This work shows that Ce13d requires both Na+ and a trivalent lanthanide (e.g. Ce3+), simultaneously. This discovery is facilitated by the sequence similarity between Ce13d and a recently reported Na+-specific DNAzyme, NaA43. The Ce13d cleavage rate linearly depends on the concentration of both metal ions. Sensitized Tb3+ luminescence and DMS footprinting experiments indicate that the guanines in the enzyme loop are important for Na+-binding. The Na+ dissociation constants of Ce13d measured from the cleavage activity assay, Tb3+ luminescence and DMS footprinting are 24.6, 16.3 and 47 mM, respectively. Mutation studies indicate that the role of Ce3+ might be replaced by G23 in NaA43. Ce3+ functions by stabilizing the transition state phosphorane, thus promoting cleavage. G23 competes favorably with low concentration Ce3+ (below 1 μM). The G23-to-hypoxanthine mutation suggests the N1 position of the guanine as a hydrogen bond donor. Together, Ce13d has two distinct metal binding sites, each fulfilling a different role. DNAzymes can be quite sophisticated in utilizing metal ions for catalysis and molecular recognition, similar to protein metalloenzymes. PMID:26657636

Biostable L-DNAzyme for Sensing of Metal Ions in Biological Systems

PubMed Central

2015-01-01

DNAzymes, an important type of metal ion-dependent functional nucleic acid, are widely applied in bioanalysis and biomedicine. However, the use of DNAzymes in practical applications has been impeded by the intrinsic drawbacks of natural nucleic acids, such as interferences from nuclease digestion and protein binding, as well as undesired intermolecular interactions with other nucleic acids. On the basis of reciprocal chiral substrate specificity, the enantiomer of D-DNAzyme, L-DNAzyme, could initiate catalytic cleavage activity with the same achiral metal ion as a cofactor. Meanwhile, by using the advantage of nonbiological L-DNAzyme, which is not subject to the interferences of biological matrixes, as recognition units, a facile and stable L-DNAzyme sensor was proposed for sensing metal ions in complex biological samples and live cells. PMID:26691677

Remediation of metal-contaminated marine sediments using active capping with limestone, steel slag, and activated carbon: a laboratory experiment.

PubMed

Park, Seong-Jik; Kang, Ku; Lee, Chang-Gu; Choi, Jae-Woo

2018-05-18

The objectives of this study are to assess the effectiveness of limestone (LS), steel slag (SS), and activated carbon (AC) as capping materials to sequester trace metals including As, Cd, Cr, Cu, Ni, Pb, and Zn in heavily contaminated marine sediments and to minimize the release of these metals into the water column. A flat flow tank was filled with 10 mm of capping material, contaminated sediments, and seawater, and the metal concentrations were monitored over 32 d. After completion of the flow tank experiments, the sediments below the capping material were sampled and were sequentially extracted. SS effectively reduced the As, Cr, Cu, Ni, Pb, and particularly Cd elution from the contaminated sediments to the overlying seawater. Adsorption and surface precipitation were the key mechanisms for interrupting the release of cationic trace metals by SS. LS was appropriate for interrupting the release of only Cu and Pb with high hydrolysis reaction constants. AC capping could interrupt the release of Cr, Cu, Ni, and particularly Zn from the sediments by binding with the metals via electrostatic interaction. The results obtained from the sequential extraction revealed that LS capping is appropriate for stabilizing Zn, whereas AC is appropriate for Cd and Pb. LS, SS, and AC can be applied effectively for remediation of sediments contaminated by trace metals because it interrupts their release and stabilizes the trace metals in the sediments.

Disclosure of key stereoelectronic factors for efficient H2 binding and cleavage in the active site of [NiFe]-hydrogenases.

PubMed

Bruschi, Maurizio; Tiberti, Matteo; Guerra, Alessandro; De Gioia, Luca

2014-02-05

A comparative analysis of a series of DFT models of [NiFe]-hydrogenases, ranging from minimal NiFe clusters to very large systems including both the first and second coordination sphere of the bimetallic cofactor, was carried out with the aim of unraveling which stereoelectronic properties of the active site of [NiFe]-hydrogenases are crucial for efficient H2 binding and cleavage. H2 binding to the Ni-SIa redox state is energetically favored (by 4.0 kcal mol(-1)) only when H2 binds to Ni, the NiFe metal cluster is in a low spin state, and the Ni cysteine ligands have a peculiar seesaw coordination geometry, which in the enzyme is stabilized by the protein environment. The influence of the Ni coordination geometry on the H2 binding affinity was then quantitatively evaluated and rationalized analyzing frontier molecular orbitals and populations. Several plausible reaction pathways leading to H2 cleavage were also studied. It turned out that a two-step pathway, where H2 cleavage takes place on the Ni-SIa redox state of the enzyme, is characterized by very low reaction barriers and favorable reaction energies. More importantly, the seesaw coordination geometry of Ni was found to be a key feature for facile H2 cleavage. The discovery of the crucial influence of the Ni coordination geometry on H2 binding and activation in the active site of [NiFe]-hydrogenases could be exploited in the design of novel biomimetic synthetic catalysts.

Influence of Humic Acid Complexation with Metal Ions on Extracellular Electron Transfer Activity.

PubMed

Zhou, Shungui; Chen, Shanshan; Yuan, Yong; Lu, Qin

2015-11-23

Humic acids (HAs) can act as electron shuttles and mediate biogeochemical cycles, thereby influencing the transformation of nutrients and environmental pollutants. HAs commonly complex with metals in the environment, but few studies have focused on how these metals affect the roles of HAs in extracellular electron transfer (EET). In this study, HA-metal (HA-M) complexes (HA-Fe, HA-Cu, and HA-Al) were prepared and characterized. The electron shuttle capacities of HA-M complexes were experimentally evaluated through microbial Fe(III) reduction, biocurrent generation, and microbial azoreduction. The results show that the electron shuttle capacities of HAs were enhanced after complexation with Fe but were weakened when using Cu or Al. Density functional theory calculations were performed to explore the structural geometry of the HA-M complexes and revealed the best binding sites of the HAs to metals and the varied charge transfer rate constants (k). The EET activity of the HA-M complexes were in the order HA-Fe > HA-Cu > HA-Al. These findings have important implications for biogeochemical redox processes given the ubiquitous nature of both HAs and various metals in the environment.

Ca(2+) -complex stability of GAPAGPLIVPY peptide in gas and aqueous phase, investigated by affinity capillary electrophoresis and molecular dynamics simulations and compared to mass spectrometric results.

PubMed

Nachbar, Markus; El Deeb, Sami; Mozafari, Mona; Alhazmi, Hassan A; Preu, Lutz; Redweik, Sabine; Lehmann, Wolf Dieter; Wätzig, Hermann

2016-03-01

Strong, sequence-specific gas-phase bindings between proline-rich peptides and alkaline earth metal ions in nanoESI-MS experiments were reported by Lehmann et al. (Rapid Commun. Mass Spectrom. 2006, 20, 2404-2410), however its relevance for physiological-like aqueous phase is uncertain. Therefore, the complexes should also be studied in aqueous solution and the relevance of the MS method for binding studies be evaluated. A mobility shift ACE method was used for determining the binding between the small peptide GAPAGPLIVPY and various metal ions in aqueous solution. The findings were compared to the MS results and further explained using computational methods. While the MS data showed a strong alkaline earth ion binding, the ACE results showed nonsignificant binding. The proposed vacuum state complex also decomposed during a molecular dynamic simulation in aqueous solution. This study shows that the formed stable peptide-metal ion adducts in the gas phase by ESI-MS does not imply the existence of analogous adducts in the aqueous phase. Comparing peptide-metal ion interaction under the gaseous MS and aqueous ACE conditions showed huge difference in binding behavior. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Investigation of DOTA-Metal Chelation Effects on the Chemical Shift of 129 Xe

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jeong, Keunhong; Slack, Clancy C.; Vassiliou, Christophoros C.

2015-09-17

Recent work has shown that xenon chemical shifts in cryptophane-cage sensors are affected when tethered chelators bind to metals. Here in this paper, we explore the xenon shifts in response to a wide range of metal ions binding to diastereomeric forms of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) linked to cryptophane-A. The shifts induced by the binding of Ca 2+, Cu 2+, Ce 3+, Zn 2+, Cd 2+, Ni 2+, Co 2+, Cr 2+, Fe 3+, and Hg 2+ are distinct. In addition, the different responses of the diastereomers for the same metal ion indicate that shifts are affected by partial folding withmore » a correlation between the expected coordination number of the metal in the DOTA complex and the chemical shift of 129Xe. Lastly, these sensors may be used to detect and quantify many important metal ions, and a better understanding of the basis for the induced shifts could enhance future designs.« less

Copper metallothioneins.

PubMed

Calvo, Jenifer; Jung, Hunmin; Meloni, Gabriele

2017-04-01

Metallothioneins (MTs) are a class of low molecular weight and cysteine-rich metal binding proteins present in all the branches of the tree of life. MTs efficiently bind with high affinity several essential and toxic divalent and monovalent transition metals by forming characteristic polynuclear metal-thiolate clusters within their structure. MTs fulfil multiple biological functions related to their metal binding properties, with essential roles in both Zn(II) and Cu(I) homeostasis as well as metal detoxification. Depending on the organism considered, the primary sequence, and the specific physiological and metabolic status, Cu(I)-bound MT isoforms have been isolated, and their chemistry and biology characterized. Besides the recognized role in the biochemistry of divalent metals, it is becoming evident that unique biological functions in selectively controlling copper levels, its reactivity as well as copper-mediated biochemical processes have evolved in some members of the MT superfamily. Selected examples are reviewed to highlight the peculiar chemical properties and biological functions of copper MTs. © 2016 IUBMB Life, 69(4):236-245, 2017. © 2017 International Union of Biochemistry and Molecular Biology.

Active site remodeling during the catalytic cycle in metal-dependent fructose-1,6-bisphosphate aldolases.

PubMed

Jacques, Benoit; Coinçon, Mathieu; Sygusch, Jurgen

2018-03-28

Crystal structures of two bacterial metal (Zn) dependent D-fructose 1,6-bisphosphate (FBP) aldolases in complex with substrate, analogues, and triose-P reaction products were determined to 1.5-2.0 Å resolution. The ligand complexes cryotrapped in native or mutant H. pylori aldolase crystals enabled a novel mechanistic description of FBP C 3 -C 4 bond cleavage. The reaction mechanism uses active site remodelling during the catalytic cycle implicating relocation of the Zn cofactor that is mediated by conformational changes of active site loops. Substrate binding initiates conformational changes, triggered upon P 1 -phosphate binding, which liberates the Zn chelating His180, allowing it to act as a general base for the proton abstraction at the FBP C 4 -hydroxyl group. A second zinc chelating His83 hydrogen bonds the substrate C 4 - hydroxyl group and assists cleavage by stabilizing the developing negative charge during proton abstraction. Cleavage is concerted with relocation of the metal cofactor from an interior to a surface exposed site, thereby stabilizing the nascent enediolate form. Conserved residue Glu142 is essential for protonation of the enediolate form, prior to product release. A D-tagatose 1,6-bisphosphate enzymatic complex reveals how His180 mediated proton abstraction controls stereospecificity of the cleavage reaction. Recognition and discrimination of the reaction products, dihydroxyacetone-P and D-glyceraldehyde-3-P, occurs via charged hydrogen bonds between hydroxyl groups of the triose-Ps and conserved residues, Asp82 and Asp255, respectively, and are crucial aspects of the enzyme's role in gluconeogenesis. Conformational changes in mobile loops β5-α7 and β6-α8 (containing catalytic residues Glu142 and His180, respectively) drive active site remodelling enabling the relocation of the metal cofactor. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Single Pt Atoms Confined into a Metal-Organic Framework for Efficient Photocatalysis.

PubMed

Fang, Xinzuo; Shang, Qichao; Wang, Yu; Jiao, Long; Yao, Tao; Li, Yafei; Zhang, Qun; Luo, Yi; Jiang, Hai-Long

2018-02-01

It is highly desirable yet remains challenging to improve the dispersion and usage of noble metal cocatalysts, beneficial to charge transfer in photocatalysis. Herein, for the first time, single Pt atoms are successfully confined into a metal-organic framework (MOF), in which electrons transfer from the MOF photosensitizer to the Pt acceptor for hydrogen production by water splitting under visible-light irradiation. Remarkably, the single Pt atoms exhibit a superb activity, giving a turnover frequency of 35 h -1 , ≈30 times that of Pt nanoparticles stabilized by the same MOF. Ultrafast transient absorption spectroscopy further unveils that the single Pt atoms confined into the MOF provide highly efficient electron transfer channels and density functional theory calculations indicate that the introduction of single Pt atoms into the MOF improves the hydrogen binding energy, thus greatly boosting the photocatalytic H 2 production activity. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reactivity of food phenols with iron and copper ions: binding, dioxygen activation and oxidation mechanisms.

PubMed

Nkhili, Ezzohra; Loonis, Michèle; Mihai, Simona; El Hajji, Hakima; Dangles, Olivier

2014-06-01

In this work, the affinity of common dietary phenols (gallic acid, caffeic acid, catechin, and rutin) for iron and copper ions was quantitatively investigated in neutral phosphate buffer as well as the reactivity of the complexes toward dioxygen. Contrasting behaviors were observed: because of the competing phosphate ions, Fe(III) binding is much slower than Fe(II) binding, which is rapidly followed by autoxidation of Fe(II) into Fe(III). With both ions, O2 consumption and H2O2 production are modest and the phenolic ligands are only slowly oxidized. By contrast, metal-phenol binding is fast with both Cu(I) and Cu(II). With Cu(I), O2 consumption and H2O2 production are very significant and the phenolic ligands are rapidly oxidized into a complex mixture of oligomers. The corresponding mechanism with Cu(II) is hampered by the preliminary rate-determining step of Cu(II) reduction by the phenols. The consequences of these findings for the stability and antioxidant activity of plant phenols are discussed.

Quantitation and localization of intracellular redox active metals by X-ray fluorescence microscopy in cortical neurons derived from APP and APLP2 knockout tissue

DOE PAGES

Ciccotosto, Giuseppe D.; James, Simon A.; Altissimo, Matteo; ...

2014-10-01

The amyloid precursor protein (APP) gene family includes APP and the amyloid precursor-like proteins, APLP1 and APLP2. These proteins contain metal binding sites for copper, zinc and iron and are known to have physiological roles in modulating the metal homeostasis in brain cells. Here we report the application of X-ray fluorescence microscopy (XFM) to investigate the subcellular distribution patterns of the metal ions Cu, Zn, Fe, and Ca in individual neurons derived from APP and APLP2 knockout mice brains to further define their role in metal homeostasis. These studies add to the growing body of data that the APP familymore » of proteins are metalloproteins that have shared as well as distinct effects on metals. As we continue to delineate the cellular effects of the APP family of proteins it is important to consider how metals are involved in their actions.« less

Recognizing metal and acid radical ion-binding sites by integrating ab initio modeling with template-based transferals.

PubMed

Hu, Xiuzhen; Dong, Qiwen; Yang, Jianyi; Zhang, Yang

2016-11-01

More than half of proteins require binding of metal and acid radical ions for their structure and function. Identification of the ion-binding locations is important for understanding the biological functions of proteins. Due to the small size and high versatility of the metal and acid radical ions, however, computational prediction of their binding sites remains difficult. We proposed a new ligand-specific approach devoted to the binding site prediction of 13 metal ions (Zn 2+ , Cu 2+ , Fe 2+ , Fe 3+ , Ca 2+ , Mg 2+ , Mn 2+ , Na + , K + ) and acid radical ion ligands (CO3 2- , NO2 - , SO4 2- , PO4 3- ) that are most frequently seen in protein databases. A sequence-based ab initio model is first trained on sequence profiles, where a modified AdaBoost algorithm is extended to balance binding and non-binding residue samples. A composite method IonCom is then developed to combine the ab initio model with multiple threading alignments for further improving the robustness of the binding site predictions. The pipeline was tested using 5-fold cross validations on a comprehensive set of 2,100 non-redundant proteins bound with 3,075 small ion ligands. Significant advantage was demonstrated compared with the state of the art ligand-binding methods including COACH and TargetS for high-accuracy ion-binding site identification. Detailed data analyses show that the major advantage of IonCom lies at the integration of complementary ab initio and template-based components. Ion-specific feature design and binding library selection also contribute to the improvement of small ion ligand binding predictions. http://zhanglab.ccmb.med.umich.edu/IonCom CONTACT: hxz@imut.edu.cn or zhng@umich.eduSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Structural dynamics of F-actin: I. Changes in the C terminus.

PubMed

Orlova, A; Egelman, E H

1995-02-03

The biochemical properties of G-actin, and the kinetics of polymerization of G-actin into F-actin, are dependent upon whether Mg2+ or Ca2+ is bound at the high-affinity metal-binding site in actin. Three-dimensional reconstructions from electron micrographs show that a bridge of density, that we interpret as arising from a major shift of the C terminus, exists between the two strands of the filament in Ca(2+)-actin that is absent in Mg(2+)-actin. This bridge is also absent in models of F-actin built from an atomic structure of G-Ca(2+)-actin. The cleavage of the DNase I-binding loop in actin between residues 42 and 43, with the non-covalent association of the 42 cleaved residues with the remainder of the actin, induces an even larger bridge of density between the two strands. When the bridge is absent, the two C-terminal residues in F-actin are easily cleaved by trypsin, while these residues become increasingly resistant to tryptic cleavage as the bridge becomes more prominent. Conversely, cleavage of the two C-terminal residues leads to a conformational change in the DNase I-binding loop. Since both the DNase I-binding loop and the metal-binding site are quite distant from the C terminus, large allosteric effects must exist in F-actin. The conformational change in F-actin that results from the creation of this bridge may be induced by myosin binding, since this movement generates changes in actin's diffraction that are very similar to the changes in the muscle X-ray pattern during activation that are associated with the binding of myosin to the thin filament.

The Effects of Select Histidine to Cysteine Mutations on Transcriptional Regulation by E. coli RcnR‡

PubMed Central

Higgins, Khadine A.; Hu, Heidi Q.; Chivers, Peter T.; Maroney, Michael J.

2013-01-01

The RcnR metalloregulator represses the transcription of the Co(II) and Ni(II) exporter, RcnAB. Previous studies have shown that Co(II) and Ni(II) bind to RcnR in six-coordinate sites, resulting in de-repression. Here, the roles of His60, His64, and His67 in specific metal recognition are examined. His60 and His64 correspond to ligands that are important for Cu(I) binding in the homologous Cu(I)-responsive metalloregulator, CsoR. These residues are known to be functionally important in RcnR transcriptional regulation. XAS was used to examine the structure of bound cognate and non-cognate metal ions, and lacZ reporter assays were used to assess the transcription of rcnA in response to metal binding in the three His → Cys mutations, H60C, H64C and H67C. These studies confirm that both Ni(II) and Co(II) use His64 as a ligand. H64C-RcnR is also the only known mutation that retains a Co(II) response while eliminating the response to Ni(II) binding. XAS data indicate that His60 and His67 are potential Co(II) ligands. The effects of the mutations of His60, His64, and His67 residues on the structures of the non-cognate metal ions (Zn(II) and Cu(I)) reveals that these residues have distinctive roles in binding non-cognate metals. None of the His → Cys mutants in RcnR confer any response to Cu(I) binding, including H64C-RcnR, where the ligands involved in Cu(I) binding in CsoR are present. These data indicate that while the secondary, tertiary and quaternary structures of CsoR and RcnR are quite similar, small changes in primary sequence reveal that the specific mechanisms involved in metal recognition are quite different. PMID:23215580

X-ray Crystal Structures Elucidate the Nucleotidyl Transfer Reaction of Transcript Initiation Using Two Nucleotides

DOE Office of Scientific and Technical Information (OSTI.GOV)

M Gleghorn; E Davydova; R Basu

2011-12-31

We have determined the X-ray crystal structures of the pre- and postcatalytic forms of the initiation complex of bacteriophage N4 RNA polymerase that provide the complete set of atomic images depicting the process of transcript initiation by a single-subunit RNA polymerase. As observed during T7 RNA polymerase transcript elongation, substrate loading for the initiation process also drives a conformational change of the O helix, but only the correct base pairing between the +2 substrate and DNA base is able to complete the O-helix conformational transition. Substrate binding also facilitates catalytic metal binding that leads to alignment of the reactive groupsmore » of substrates for the nucleotidyl transfer reaction. Although all nucleic acid polymerases use two divalent metals for catalysis, they differ in the requirements and the timing of binding of each metal. In the case of bacteriophage RNA polymerase, we propose that catalytic metal binding is the last step before the nucleotidyl transfer reaction.« less

Methylocystis strain SB2 materials and methods

DOEpatents

Semrau, Jeremy D; Gallagher, Warren; Yoon, Sukhwan; Im, Jeongdae; DiSpririto, Alan A; Lee, Sung-Woo; Hartsel, Scott; McEllistrem, Marcus T

2014-01-14

The present disclosures provides isolated or purified compounds, each of which bind to a metal atom. Generally, the compounds are small in size (e.g., molecular weight of less than about 1 kDa) and peptidic in nature, inasmuch as the compounds comprise amino acids. In some embodiments, the compound comprises a structure of Formula I; M.sub.1-P.sub.1-M.sub.2-P.sub.2 wherein each of P.sub.1 and P.sub.2 is a peptide comprising at least two amino acids, M.sub.1 is a first metal binding moiety comprising a substituted imidazolone ring, M.sub.2 is a second metal binding moiety comprising a substituted oxazolone ring, and wherein M.sub.1 and M.sub.2 bind to a single metal atom. Also provided are related complexes, conjugates, cells which synthesize the compounds of the present disclosures, substantially homogenous cultures thereof, kits and compositions, and methods of making or using the materials of the present disclosures.

Inflammatory responses to the occupational inhalation of metal fume.

PubMed

Palmer, K T; McNeill Love, R M C; McNeill-Love, R; Poole, J R; Coggon, D; Frew, A J; Linaker, C H; Shute, J K

2006-02-01

Occupational exposure to metal fume promotes a reversible increase in the risk of pneumonia, but by mechanisms which are unclear. To investigate, the current authors measured various markers of host defence function in welders and nonwelders. Induced sputum and venous blood samples were collected from 27 welders with regular long-term exposure to ferrous metal fume and 31 unexposed matched controls. In sputum, the present authors measured cell counts, the soluble and cellular iron concentration, and levels of interleukin-8, tumour necrosis factor-alpha, myeloperoxidase, matrix metalloproteinase-9, immunoglobulin (Ig)A, alpha(2)-macroglobulin and unsaturated iron-binding capacity. Blood samples were assayed for evidence of neutrophil activation and pneumococcal IgG antibodies. Welders had significantly higher iron levels and a substantially lower unsaturated iron-binding capacity in their sputum, but, despite a high iron challenge, there was a noteworthy absence of an inflammatory response. Only blood counts of eosinophils and basophils were significantly related to the extent of welding. Weak nonsignificant trends were observed for several other measures, consistent with low-grade priming of neutrophils. In conclusion, these data suggest that chronic exposure to metal fume blunts responsiveness to inhaled particulate matter. However, the mechanism behind the lack of detectable local inflammatory response requires further investigation.

Contrasting effects of photochemical and microbial degradation on Cu(II) binding with fluorescent DOM from different origins.

PubMed

Xu, Huacheng; Guan, Dong-Xing; Zou, Li; Lin, Hui; Guo, Laodong

2018-08-01

Effects of photochemical and microbial degradation on variations in composition and molecular-size of dissolved organic matter (DOM) from different sources (algal and soil) and the subsequent influence on Cu(II) binding were investigated using UV-Vis, fluorescence excitation-emission matrices coupled with parallel factor analysis, flow field-flow fractionation (FlFFF), and metal titration. The degradation processes resulted in an initial rapid decline in the bulk dissolved organic carbon and chromophoric and fluorescent DOM components, followed by a small or little decrease. Specifically, photochemical reaction decreased the aromaticity, humification and apparent molecular weights of all DOM samples, whereas a reverse trend was observed during microbial degradation. The FlFFF fractograms revealed that coagulation of both protein- and humic-like DOM induced an increase in molecular weights for algal-DOM, while the molecular weight enhancement for allochthonous soil samples was mainly attributed to the self-assembly of humic-like components. The Cu(II) binding capacity of algal-derived humic-like and fulvic-like DOM consistently increased during photo- and bio-degradation, while the soil-derived DOM exhibited a slight decline in Cu(II) binding capacity during photo-degradation but a substantial increase during microbial degradation, indicating source- and degradation-dependent metal binding heterogeneities. Pearson correlation analysis demonstrated that the Cu(II) binding potential was mostly related with aromaticity and molecular size for allochthonous soil-derived DOM, but was regulated by both DOM properties and specific degradation processes for autochthonous algal-derived DOM. This study highlighted the coupling role of inherent DOM properties and external environmental processes in regulating metal binding, and provided new insights into metal-DOM interactions and the behavior and fate of DOM-bound metals in aquatic environments. Copyright © 2018 Elsevier Ltd. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nash, David J.; Restrepo, David T.; Parra, Natalia S.

Catalytic hydrogenation is an important process used for the production of everything from foods to fuels. Current heterogeneous implementations of this process utilize metals as the active species. Until recently, catalytic heterogeneous hydrogenation over a metal-free solid was unknown; implementation of such a system would eliminate the health, environmental, and economic concerns associated with metal-based catalysts. We report good hydrogenation rates and yields for a metal-free heterogeneous hydrogenation catalyst as well as its unique hydrogenation mechanism. We achieved catalytic hydrogenation of olefins over defect-laden h-BN (dh-BN) in a reactor designed to maximize the defects in h-BN sheets. Good yields (>90%)more » and turnover frequencies (6 × 10 –5–4 × 10 –3) were obtained for the hydrogenation of propene, cyclohexene, 1,1-diphenylethene, (E)- and (Z)-1,2-diphenylethene, octadecene, and benzylideneacetophenone. Temperature-programmed desorption of ethene over processed h-BN indicates the formation of a highly defective structure. Solid-state NMR (SSNMR) measurements of dh-BN with high and low propene surface coverages show four different binding modes. The introduction of defects into h-BN creates regions of electronic deficiency and excess. Density functional theory calculations show that both the alkene and hydrogen-bond order are reduced over four specific defects: boron substitution for nitrogen (B N), vacancies (V B and V N), and Stone–Wales defects. SSNMR and binding-energy calculations show that V N are most likely the catalytically active sites. Our work shows that catalytic sites can be introduced into a material previously thought to be catalytically inactive through the production of defects.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gavva, S.R.; Harris, B.G.; Cook, P.F.

A thiol group at the malate-binding site of the NAD-malic enzyme from Ascaris suum has been modified to thiocyanate. The modified enzyme generally exhibits slight increases in K{sub NAD} and K{sub i metal} and decreases in V{sub max} as the metal size increases from Mg{sup 2+} to Mn{sup 2+} to Cd{sup 2+}, indicative of crowding in the site. The K{sub malate} value increases 10- to 30-fold, suggesting that malate does not bind optimally to the modified enzyme. Deuterium isotope effects on V and V/K{sub malate} increase with all three metal ions compared to the native enzyme concomitant with a decreasemore » in the {sup 13}C isotope effect, suggesting a switch in the rate limitation of the hydride transfer and decarboxylation steps with hydride transfer becoming more rate limiting. The {sup 13}C effect decreases only slightly when obtained with deuterated malate, suggestive of the presence of a secondary {sup 13}C effect in the hydride transfer step, similar to data obtained with non-nicotinamide-containing dinucleotide substrates for the native enzyme (see the preceding paper in this issue). The native enzyme is inactivated in a time-dependent manner by Cd{sup 2+}. This inactivation occurs whether the enzyme alone is present or whether the enzyme is turning over with Cd{sup 2+} as the divalent metal activator. Upon inactivation, only Cd{sup 2+} ions are bound at high stoichiometry to the enzyme, which eventually becomes denatured. Conversion of the active-site thiol to thiocyanate makes it more difficult to inactivate the enzyme by treatment with Cd{sup 2+}.« less

Characterization of metal binding sites onto biochar using rare earth elements as a fingerprint.

PubMed

Pourret, Olivier; Houben, David

2018-02-01

The ability of biochar to immobilize metals relies on the amount of functional groups at its surface but the contribution of each functional groups (e.g. carboxylic, phenolic) to metal bonding is poorly known. Using a new approach based on previous works on rare earth element (REE) interactions with humic substances, we aim at elucidating the relative contribution of these binding sites to metal sorption under various conditions (i.e. pH and ionic strengths, IS). Using batch experiments, REE sorption onto biochar was analyzed from pH 3 to 9 and IS 10 -1 mol/L to 10 -3 mol/L. Rare earth element patterns show a Middle REE (MREE) downward concavity at acidic pH and low ionic strength. These patterns are in good agreement with existing datasets quantifying REE binding with humic substances. Indeed, the MREE downward concavity displayed by REE-biochar complexation pattern compares well with REE patterns with various organic compounds. This similarity in the REE complexation pattern shapes suggests that carboxylic groups are the main binding sites of REE in biochar. Overall, our results indicate that the strength of the metal bonding with biochar increases when pH and IS increase, suggesting that biochar is more efficient for long-term metal immobilization at near neutral pH and high ionic strength.

Hydrogen adsorption in HKUST-1: a combined inelastic neutron scattering and first-principles study.

PubMed

Brown, Craig M; Liu, Yun; Yildirim, Taner; Peterson, Vanessa K; Kepert, Cameron J

2009-05-20

Hydrogen adsorption in high surface area nanoporous coordination polymers has attracted a great deal of interest in recent years due to the potential applications in energy storage. Here we present combined inelastic neutron scattering measurements and detailed first-principles calculations aimed at unraveling the nature of hydrogen adsorption in HKUST-1 (Cu3(1,3,5-benzenetricarboxylate)2), a metal-organic framework (MOF) with unsaturated metal centers. We reveal that, in this system, the major contribution to the overall binding comes from the classical Coulomb interaction which is not screened due to the open metal site; this explains the relatively high binding energies and short H2-metal distances observed in MOFs with exposed metal sites as compared to traditional ones. Despite the short distances, there is no indication of an elongation of the H-H bond for the bound H2 molecule at the metal site. We find that both the phonon and rotational energy levels of the hydrogen molecule are closely similar, making the interpretation of the inelastic neutron scattering data difficult. Finally, we show that the orientation of H2 has a surprisingly large effect on the binding potential, reducing the classical binding energy by almost 30%. The implication of these results for the development of MOF materials for better hydrogen storage is discussed.

Hydrogen adsorption in HKUST-1: a combined inelastic neutron scattering and first-principles study

NASA Astrophysics Data System (ADS)

Brown, Craig M.; Liu, Yun; Yildirim, Taner; Peterson, Vanessa K.; Kepert, Cameron J.

2009-05-01

Hydrogen adsorption in high surface area nanoporous coordination polymers has attracted a great deal of interest in recent years due to the potential applications in energy storage. Here we present combined inelastic neutron scattering measurements and detailed first-principles calculations aimed at unraveling the nature of hydrogen adsorption in HKUST-1 (Cu3(1,3,5-benzenetricarboxylate)2), a metal-organic framework (MOF) with unsaturated metal centers. We reveal that, in this system, the major contribution to the overall binding comes from the classical Coulomb interaction which is not screened due to the open metal site; this explains the relatively high binding energies and short H2-metal distances observed in MOFs with exposed metal sites as compared to traditional ones. Despite the short distances, there is no indication of an elongation of the H-H bond for the bound H2 molecule at the metal site. We find that both the phonon and rotational energy levels of the hydrogen molecule are closely similar, making the interpretation of the inelastic neutron scattering data difficult. Finally, we show that the orientation of H2 has a surprisingly large effect on the binding potential, reducing the classical binding energy by almost 30%. The implication of these results for the development of MOF materials for better hydrogen storage is discussed.

Binding of naringin and naringenin with hen egg white lysozyme: A spectroscopic investigation and molecular docking study

NASA Astrophysics Data System (ADS)

Das, Sourav; Ghosh, Pooja; Koley, Sudipta; Singha Roy, Atanu

2018-03-01

The interactions of naringenin (NG) and naringin (NR) with Hen Egg White Lysozyme (HEWL) in aqueous medium have been investigated using UV-vis spectroscopy, steady-state fluorescence, circular dichroism (CD), Fourier Transform infrared spectroscopy (FT-IR) and molecular docking analyses. Both NG and NR can quench the intrinsic fluorescence of HEWL via static quenching mechanism. At 300 K, the value of binding constant (Kb) of HEWL-NG complex (5.596 ± 0.063 × 104 M- 1) was found to be greater than that of HEWL-NR complex (3.404 ± 0.407 × 104 M- 1). The negative ΔG° values in cases of both the complexes specify the spontaneous binding. The binding distance between the donor (HEWL) and acceptor (NG/NR) was estimated using the Försters theory and the possibility of non-radiative energy transfer from HEWL to NG/NR was observed. The presence of metal ions (Ca2 +, Cu2 + and Fe2 +) decreased the binding affinity of NG/NR towards HEWL. Synchronous fluorescence studies indicate the change in Trp micro-environment due to the incorporation of NG/NR into HEWL. CD and FT-IR studies indicated that the α-helicity of the HEWL was slightly enhanced due to ligand binding. NG and NR inhibited the enzymatic activity of HEWL and exhibited their affinity for the active site of HEWL. Molecular docking studies revealed that both NG and NR bind in the close vicinity of Trp 62 and Trp 63 residues which is vital for the catalytic activity.

Stereochemical control over Mn(II)-Thio versus Mn(II)-Oxy coordination in adenosine 5 prime -O-(1-thiodiphosphate) complexes at the active site of creatine kinase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smithers, G.W.; Sammons, R.D.; Goodhart, P.J.

1989-02-21

The stereochemical configurations of the Mn(II) complexes with the resolved epimers of adenosine 5{prime}-O-(1-thiodiphosphate) (ADP{alpha}S), bound at the active site of creatine kinase, have been determined in order to assess the relative strengths of enzymic stereoselectivity versus Lewis acid/base preferences in metal-ligand binding. Electron paramagnetic resonance (EPR) data have been obtained for Mn(II) in anion-stabilized, dead-end (transition-state analogue) complexes, in ternary enzyme-Mn{sup II}ADP{alpha}S complexes, and in the central complexes of the equilibrium mixture. The modes of coordination of Mn(II) at P{sub alpha} in the nitrate-stabilized, dead-end complexes with each epimer of ADP{alpha}S were ascertained by EPR measurements with (R{sub p})-({alpha}-{supmore » 17}O)ADP{alpha}S and (S{sub p})-({alpha}-{sup 17}O)ADP{alpha}S. A reduction in the magnitude of the {sup 55}Mn hyperfine coupling constant in the spectrum for the complex containing (S{sub p})-ADP{alpha}S is indicative of Mn(II)-thio coordination at P{sub alpha}. The results indicate that a strict discrimination for a unique configuration of the metal-nucleotide substrate is expressed upon binding of all of the substrates to form the active complex (or an analogue thereof). This enzymic stereoselectivity provides sufficient binding energy to overcome an intrinsic preference for the hard Lewis acid Mn(II) to coordinate to the hard Lewis base oxygen.« less

Effects of UV-B and heavy metals on nitrogen and phosphorus metabolism in three cyanobacteria.

PubMed

Yadav, Shivam; Prajapati, Rajesh; Atri, Neelam

2016-01-01

Cyanobacteria sp. (diazotrophic and planktonic) hold a major position in ecosystem, former one due to their intrinsic capability of N2-fixation and later because of mineralization of organic matter. Unfortunately, their exposure to variety of abiotic stresses is unavoidable. Comparative analysis of interactive effect of UV-B and heavy metals (Cd/Zn) on nitrogen and phosphorus metabolism of three cyanobacteria (Anabaena, Microcystis, Nostoc) revealed additive inhibition (χ(2) significant p < 0.05) of NH4(+) and PO4(3-) uptake whereas increase in nitrate uptake upon UV-B + heavy metal exposure. Glutamine synthetase and Alkaline phosphatase activity was reduced after all treatments whereas Nitrate reductase activity showed slight stimulation in UV-B and UV-B + heavy metals treatment. Combination of UV-B and metals seems more detrimental to the NH4(+), PO4(3-) uptake, GS and APA activity. A significant stimulation in NO3(-) uptake and NR activity following exposure to UV-B in all the three cyanobacteria suggests UV-B-induced structural change(s) in the enzyme/carriers. Metals seem to compete for the binding sites of the enzymes and carriers; as noticed for Anabaena and Microcystis showing change in Km while no change in the Km value of Nostoc suggests non-competitive nutrient uptake. Higher accumulation and more adverse effect on Na(+) and K(+) efflux proposes Cd as more toxic compared to Zn. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Metal imaging in neurodegenerative diseases

PubMed Central

Bourassa, Megan W.

2014-01-01

Metal ions are known to play an important role in many neurodegenerative diseases including Alzheimer’s disease (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), and prion diseases. In these diseases, aberrant metal binding or improper regulation of redox active metal ions can induce oxidative stress by producing cytotoxic reactive oxygen species (ROS). Altered metal homeostasis is also frequently seen in the diseased state. As a result, the imaging of metals in intact biological cells and tissues has been very important for understanding the role of metals in neurodegenerative diseases. A wide range of imaging techniques have been utilized, including X-ray fluorescence microscopy (XFM), particle induced X-ray emission (PIXE), energy dispersive X-ray spectroscopy (EDS), laser ablation inductively coupled mass spectrometry (LA-ICP-MS), and secondary ion mass spectrometry (SIMS), all of which allow for the imaging of metals in biological specimens with high spatial resolution and detection sensitivity. These techniques represent unique tools for advancing the understanding of the disease mechanisms and for identifying possible targets for developing treatments. In this review, we will highlight the advances in neurodegenerative disease research facilitated by metal imaging techniques. PMID:22797194

Theoretical Investigations of the Electrochemical Reduction of CO on Single Metal Atoms Embedded in Graphene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kirk, Charlotte; Chen, Leanne D.; Siahrostami, Samira

Single transition metal atoms embedded at single vacancies of graphene provide a unique paradigm for catalytic reactions. We present a density functional theory study of such systems for the electrochemical reduction of CO. Theoretical investigations of CO electrochemical reduction are particularly challenging in that electrochemical activation energies are a necessary descriptor of activity. We determined the electrochemical barriers for key proton–electron transfer steps using a state-of-the-art, fully explicit solvent model of the electrochemical interface. The accuracy of GGA-level functionals in describing these systems was also benchmarked against hybrid methods. We find the first proton transfer to form CHO from COmore » to be a critical step in C 1 product formation. On these single atom sites, the corresponding barrier scales more favorably with the CO binding energy than for 211 and 111 transition metal surfaces, in the direction of improved activity. Intermediates and transition states for the hydrogen evolution reaction were found to be less stable than those on transition metals, suggesting a higher selectivity for CO reduction. We present a rate volcano for the production of methane from CO. We identify promising candidates with high activity, stability, and selectivity for the reduction of CO. As a result, this work highlights the potential of these systems as improved electrocatalysts over pure transition metals for CO reduction.« less

Theoretical Investigations of the Electrochemical Reduction of CO on Single Metal Atoms Embedded in Graphene

DOE PAGES

Kirk, Charlotte; Chen, Leanne D.; Siahrostami, Samira; ...

2017-12-18

Single transition metal atoms embedded at single vacancies of graphene provide a unique paradigm for catalytic reactions. We present a density functional theory study of such systems for the electrochemical reduction of CO. Theoretical investigations of CO electrochemical reduction are particularly challenging in that electrochemical activation energies are a necessary descriptor of activity. We determined the electrochemical barriers for key proton–electron transfer steps using a state-of-the-art, fully explicit solvent model of the electrochemical interface. The accuracy of GGA-level functionals in describing these systems was also benchmarked against hybrid methods. We find the first proton transfer to form CHO from COmore » to be a critical step in C 1 product formation. On these single atom sites, the corresponding barrier scales more favorably with the CO binding energy than for 211 and 111 transition metal surfaces, in the direction of improved activity. Intermediates and transition states for the hydrogen evolution reaction were found to be less stable than those on transition metals, suggesting a higher selectivity for CO reduction. We present a rate volcano for the production of methane from CO. We identify promising candidates with high activity, stability, and selectivity for the reduction of CO. As a result, this work highlights the potential of these systems as improved electrocatalysts over pure transition metals for CO reduction.« less

Peptide-templated noble metal catalysts: syntheses and applications

PubMed Central

Wang, Wei; Anderson, Caleb F.; Wang, Zongyuan; Wu, Wei

2017-01-01

Noble metal catalysts have been widely used in many applications because of their high activity and selectivity. However, a controllable preparation of noble metal catalysts still remains as a significant challenge. To overcome this challenge, peptide templates can play a critical role in the controllable syntheses of catalysts owing to their flexible binding with specific metallic surfaces and self-assembly characteristics. By employing peptide templates, the size, shape, facet, structure, and composition of obtained catalysts can all be specifically controlled under the mild synthesis conditions. In addition, catalysts with spherical, nanofiber, and nanofilm structures can all be produced by associating with the self-assembly characteristics of peptide templates. Furthermore, the peptide-templated noble metal catalysts also reveal significantly enhanced catalytic behaviours compared with conventional catalysts because the electron conductivity, metal dispersion, and reactive site exposure can all be improved. In this review, we summarize the research progresses in the syntheses of peptide-templated noble metal catalysts. The applications of the peptide-templated catalysts in organic reactions, photocatalysis, and electrocatalysis are discussed, and the relationship between structure and activity of these catalysts are addressed. Future opportunities, including new catalytic materials designed by using biological principles, are indicated to achieve selective, eco-friendly, and energy neutral synthesis approaches. PMID:28507701

Fatty Acid-Mediated Inhibition of Metal Binding to the Multi-Metal Site on Serum Albumin: Implications for Cardiovascular Disease.

PubMed

Blindauer, Claudia A; Khazaipoul, Siavash; Yu, Ruitao; Stewart, Alan J

2016-01-01

Human serum albumin (HSA) is the major protein in blood plasma and is responsible for circulatory transport of a range of small molecules including fatty acids, metal ions and drugs. We previously identified the major plasma Zn2+ transport site on HSA and revealed that fatty-acid binding (at a distinct site called the FA2 site) and Zn2+ binding are interdependent via an allosteric mechanism. Since binding affinities of long-chain fatty acids exceed those of plasma Zn2+, this means that under certain circumstances the binding of fatty acid molecules to HSA is likely to diminish HSA Zn2+-binding, and hence affects the control of circulatory and cellular Zn2+ dynamics. This relationship between circulatory fatty acid and Zn2+ dynamics is likely to have important physiological and pathological implications, especially since it has been recognised that Zn2+ acts as a signalling agent in many cell types. Fatty acid levels in the blood are dynamic, but most importantly, chronic elevation of plasma fatty acid levels is associated with some metabolic disorders and disease states - including myocardial infarction and other cardiovascular diseases. In this article, we briefly review the metal-binding properties of albumin and highlight the importance of their interplay with fatty acid binding. We also consider the impact of this dynamic link upon levels and speciation of plasma Zn2+, its effect upon cellular Zn2+ homeostasis and its relevance to cardiovascular and circulatory processes in health and disease.

Studies on the chitin/chitosan binding properties of six cuticular proteins analogous to peritrophin 3 from Bombyx mori.

PubMed

Qu, M; Ren, Y; Liu, Y; Yang, Q

2017-08-01

Chitin deacetylation is required to make the cuticle rigid and compact through chitin chain crosslinking. Thus it is presumed that specialized proteins are required to bind deacetylated chitin chains together. However, deacetylated-chitin binding proteins have not ever been reported. In a previous work, six cuticular proteins analogous to peritrophin 3 (CPAP3s) were found to be abundant in the moulting fluid of Bombyx mori. In this study, these BmCPAP3s (BmCPAP3-A1, BmCPAP3-A2, BmCPAP3-B, BmCPAP3-C, BmCPAP3-D1 and BmCPAP3-D2) were cloned and expressed in Escherichia coli and purified using metal-chelating affinity chromatography. Their binding activities demonstrated that although all of the BmCPAP3s showed similar binding abilities toward crystalline chitin and colloidal chitin, they differed in their affinities toward partially and fully deacetylated chitin. Amongst them, BmCPAP3-D1 exhibited the highest binding activity toward deacetylated chitin. The gene expression pattern of BmCPAP3-D1 was similar to BmCPAP3-A1 and BmCPAP3-C at most stages except that it was dramatically upregulated at the beginning of the pupa to adult transition stage. This work is the first report of a chitin-binding protein, BmCPAP3-D1, which exhibits high binding affinity to deacetylated chitin. © 2017 The Royal Entomological Society.

Macrocycles inserted in graphene: from coordination chemistry on graphene to graphitic carbon oxide.

NASA Astrophysics Data System (ADS)

Liu, Wei; Liu, Jingyao; Miao, Maosheng

Tuning the electronic structure and the chemical properties of graphene by binding with metals has become a focus in the area of two dimension materials. Despite many interesting results and promising potentials, the approach suffers from weak binding and the high reactivity of the metal atoms. On the other hand, many macrocyclic molecules such as crown ether show strong and selective binding with metal atoms. The alliance of the two substances will largely benefit the two parallel fields: it will provide a scaffold for coordination chemistry as well as a controllable method for tuning the electronic structure of graphene through strong binding with metals. Here, using crown ether as an example, we demonstrate by first principles calculations that the embedment of macrocyclic molecules into graphene honeycomb lattice can be very thermochemically favored. The embedment of crown ether on graphene can form a family of new two-dimensional materials that possess varying band gaps and band edges. The one with highest O composition (C2O), with similar structure features as graphilic C3N4, shows strong potentials for photolysis and as true two-dimensional superconductor while binding with alkali metals. Calculations are performed on NSF-funded XSEDE resources (TG-DMR130005). This research is also supported by National Natural Science Foundation of China (Grants No. 21373098) in China.

Green synthesis of nano sized transition metal complexes containing heterocyclic Schiff base: Structural and morphology characterization and bioactivity study

NASA Astrophysics Data System (ADS)

Jawoor, Shailaja S.; Patil, Sangamesh A.; Kumbar, Mahantesh; Ramawadgi, Prashant B.

2018-07-01

In the current involvement of our research work in coordination chemistry, novel transition metal complexes were synthesized from regular reflux method and hydrothermal method using Schiff base prepared via condensation of ethyl 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate with 8-carbaldehyde-7-hydroxy-4-methylcoumarin. All the synthesized compounds were interpreted using different analytical, physicochemical and spectral methods such as magnetic moment measurement, FT-IR, 1H and 13C NMR, GCMS/ESI-MS, UV/Vis spectroscopy and TGA. The size and morphology of the nano metal complexes were determined using atomic force microscope (AFM), field emission scanning electron spectroscopy (FE-SEM) and X-ray powder diffraction (PXRD). The non-electrolytic nature of the metal complexes was confirmed by molar conductance studies. The obtained FT-IR data supports the binding of metal ion to Schiff base. Elemental analysis study suggests [ML2(H2O)2] stoichiometry, here M = Co(II), Ni(II) and Cu(II), L = deprotonated ligand. Electronic spectral results reveal six-coordinated geometry for the synthesized metal complexes. All the tested compounds show good DNA cleavage (Calf Thymus DNA) and in vitro anticancer activity (PA-I cell line), the activity results for the tested compounds are prominent and compound 9 exhibited a little enhanced activity than the other tested compounds.

DNA as a Target for Anticancer Phen-Imidazole Pd(II) Complexes.

PubMed

Heydari, Maryam; Moghadam, Mahboube Eslami; Tarlani, AliAkbar; Farhangian, Hossein

2017-05-01

Imidazole ring is a known structure in many natural or synthetic drug molecules and its metal complexes can interact with DNA and do the cleavage. Hence, to study the influence of the structure and size of the ligand on biological behavior of metal complexes, two water-soluble Pd(II) complexes of phen and FIP ligands (where phen is 1,10-phenanthroline and FIP is 2-(Furan-2-yl)-1H-Imidazo[4,5-f][1, 10]phenanthroline) with the formula of [Pd(phen)(FIP)](NO 3 ) 2 and [Pd(FIP) 2 ]Cl 2 , that were activated against chronic myelogenous leukemia cell line, K562, were selected. Also, the interaction of these anticancer Pd(II) complexes with highly polymerized calf thymus DNA was extensively studied by means of electronic absorption, fluorescence, and circular dichroism in Tris-buffer. The results showed that the binding was positive cooperation and [Pd(phen)(FIP)](NO 3 ) 2 (K f  = 127 M -1 G = 1.2) exhibited higher binding constant and number of binding sites than [Pd(FIP) 2 ]Cl 2 (K f  = 13 M -1 G = 1.03) upon binding to DNA. The fluorescence data indicates that quenching effect for [Pd(phen)(FIP)](NO 3 ) 2 (K SV  = 58 mM -1 ) was higher than [Pd(FIP) 2 ]Cl 2 (K SV  = 12 mM -1 ). Also, [Pd(FIP) 2 ]Cl 2 interacts with ethidium bromide-DNA, as non-competitive inhibition, and can bind to DNA via groove binding and [Pd(phen)(FIP)](NO 3 ) 2 can intercalate in DNA. These results were confirmed by circular dichroism spectra. Docking data revealed that longer complexes have higher interaction energy and bind to DNA via groove binding. Graphical Abstract Two anticancer Pd(II) complexes of imidazole derivative have been synthesized and interacted with calf thymus DNA. Modes of binding have been studied by electronic absorption, fluorescence, and CD measurements. [Pd(FIP) 2 ]Cl 2 can bind to DNA via groove binding while intercalation mode of binding is observed for [Pd(phen)(FIP)](NO 3 ) 2 .

Probes of the catalytic site of cysteine dioxygenase.

PubMed

Chai, Sergio C; Bruyere, John R; Maroney, Michael J

2006-06-09

The first major step of cysteine catabolism, the oxidation of cysteine to cysteine sulfinic acid, is catalyzed by cysteine dioxygenase (CDO). In the present work, we utilize recombinant rat liver CDO and cysteine derivatives to elucidate structural parameters involved in substrate recognition and x-ray absorption spectroscopy to probe the interaction of the active site iron center with cysteine. Kinetic studies using cysteine structural analogs show that most are inhibitors and that a terminal functional group bearing a negative charge (e.g. a carboxylate) is required for binding. The substrate-binding site has no stringent restrictions with respect to the size of the amino acid. Lack of the amino or carboxyl groups at the alpha-carbon does not prevent the molecules from interacting with the active site. In fact, cysteamine is shown to be a potent activator of the enzyme without being a substrate. CDO was also rendered inactive upon complexation with the metal-binding inhibitors azide and cyanide. Unlike many non-heme iron dioxygenases that employ alpha-keto acids as cofactors, CDO was shown to be the only dioxygenase known to be inhibited by alpha-ketoglutarate.

Investigating the features in differential absorbance spectra of NOM associated with metal ion binding: A comparison of experimental data and TD-DFT calculations for model compounds.

PubMed

Yan, Mingquan; Han, Xuze; Zhang, Chenyang

2017-11-01

In this study, seven model compounds containing typical functional groups (phenolic and carboxylic groups) present in nature organic matter (NOM) were used to ascertain the nature of the characteristic bands in differential absorbance spectra (DAS) of NOM that are induced by metal ion binding. Some similarities were found between the DAS of the examined model compounds, caffeic acid, ferulic acid, sinapic acid, terephthalic acid, isophthalic acid, esculetin and myricetin and those of NOM. The binding of Cu(II) with carboxylic group might produce two peaks, A1 and A2, while the binding of Cu(II) with phenolic group might produce all four Gaussian peaks, from A1 to A4 displayed in the DAS of NOM. The UV-visible spectra predicted using time-dependent density functional theory (TD-DFT)-based methods met well with the experimental DAS of model compounds at different stages of Cu(II) binding. It demonstrates that the features in absorbance spectra are chiefly caused by HOMO (Highest Occupied Molecular Orbital) - LUMO (Lowest Unoccupied Molecular Orbital) transitions in the molecule and that the appearance of peaks in DAS reflects the changes of the molecular orbitals around reactive functional groups in a molecule before and after metal ion binding. The basis of the DAS features of NOM that are induced by metal ion binding could be identified primarily by the frontier molecular orbital theory. Copyright © 2017 Elsevier Ltd. All rights reserved.

DNA-Binding Interaction Studies of Microwave Assisted Synthesized Sulfonamide Substituted 8-Hydroxyquinoline Derivatives.

PubMed

Dixit, Ritu B; Patel, Tarosh S; Vanparia, Satish F; Kunjadiya, Anju P; Keharia, Harish R; Dixit, Bharat C

2011-01-01

Sulfonamide substituted 8-hydroxyquinoline derivatives were prepared using a microwave synthesizer. The interaction of sulfonamide substituted 8-hydroxyquinoline derivatives and their transition metal complexes with Plasmid (pUC 19) DNA and Calf Thymus DNA were investigated by UV spectroscopic studies and gel electrophoresis measurements. The interaction between ligand/metal complexes and DNA was carried out by increasing the concentration of DNA from 0 to 12 μl in UV spectroscopic study, while the concentration of DNA in gel electrophoresis remained constant at 10 μl. These studies supported the fact that, the complex binds to DNA by intercalation via ligand into the base pairs of DNA. The relative binding efficacy of the complexes to DNA was much higher than the binding efficacy of ligands, especially the complex of Cu-AHQMBSH had the highest binding ability to DNA. The mobility of the bands decreased as the concentration of the complex was increased, indicating that there was increase in the interaction between the metal ion and DNA. Complexes of AHQMBSH were excellent for DNA binding as compared to HQMABS.

The regulation of integrin function by divalent cations

PubMed Central

Zhang, Kun; Chen, JianFeng

2012-01-01

Integrins are a family of α/β heterodimeric adhesion metalloprotein receptors and their functions are highly dependent on and regulated by different divalent cations. Recently advanced studies have revolutionized our perception of integrin metal ion-binding sites and their specific functions. Ligand binding to integrins is bridged by a divalent cation bound at the MIDAS motif on top of either α I domain in I domain-containing integrins or β I domain in α I domain-less integrins. The MIDAS motif in β I domain is flanked by ADMIDAS and SyMBS, the other two crucial metal ion binding sites playing pivotal roles in the regulation of integrin affinity and bidirectional signaling across the plasma membrane. The β-propeller domain of α subunit contains three or four β-hairpin loop-like Ca2+-binding motifs that have essential roles in integrin biogenesis. The function of another Ca2+-binding motif located at the genu of α subunit remains elusive. Here, we provide an overview of the integrin metal ion-binding sites and discuss their roles in the regulation of integrin functions. PMID:22647937

Amides Do Not Always Work: Observation of Guest Binding in an Amide-Functionalized Porous Metal-Organic Framework.

PubMed

Benson, Oguarabau; da Silva, Ivan; Argent, Stephen P; Cabot, Rafel; Savage, Mathew; Godfrey, Harry G W; Yan, Yong; Parker, Stewart F; Manuel, Pascal; Lennox, Matthew J; Mitra, Tamoghna; Easun, Timothy L; Lewis, William; Blake, Alexander J; Besley, Elena; Yang, Sihai; Schröder, Martin

2016-11-16

An amide-functionalized metal organic framework (MOF) material, MFM-136, shows a high CO 2 uptake of 12.6 mmol g -1 at 20 bar and 298 K. MFM-136 is the first example of an acylamide pyrimidyl isophthalate MOF without open metal sites and, thus, provides a unique platform to study guest binding, particularly the role of free amides. Neutron diffraction reveals that, surprisingly, there is no direct binding between the adsorbed CO 2 /CH 4 molecules and the pendant amide group in the pore. This observation has been confirmed unambiguously by inelastic neutron spectroscopy. This suggests that introduction of functional groups solely may not necessarily induce specific guest-host binding in porous materials, but it is a combination of pore size, geometry, and functional group that leads to enhanced gas adsorption properties.

How Metal Substitution Affects the Enzymatic Activity of Catechol-O-Methyltransferase

PubMed Central

Sparta, Manuel; Alexandrova, Anastassia N.

2012-01-01

Catechol-O-methyltransferase (COMT) degrades catecholamines, such as dopamine and epinephrine, by methylating them in the presence of a divalent metal cation (usually Mg(II)), and S-adenosyl-L-methionine. The enzymatic activity of COMT is known to be vitally dependent on the nature of the bound metal: replacement of Mg(II) with Ca(II) leads to a complete deactivation of COMT; Fe(II) is slightly less than potent Mg(II), and Fe(III) is again an inhibitor. Considering the fairly modest role that the metal plays in the catalyzed reaction, this dependence is puzzling, and to date remains an enigma. Using a quantum mechanical / molecular mechanical dynamics method for extensive sampling of protein structure, and first principle quantum mechanical calculations for the subsequent mechanistic study, we explicate the effect of metal substitution on the rate determining step in the catalytic cycle of COMT, the methyl transfer. In full accord with experimental data, Mg(II) bound to COMT is the most potent of the studied cations and it is closely followed by Fe(II), whereas Fe(III) is unable to promote catalysis. In the case of Ca(II), a repacking of the protein binding site is observed, leading to a significant increase in the activation barrier and higher energy of reaction. Importantly, the origin of the effect of metal substitution is different for different metals: for Fe(III) it is the electronic effect, whereas in the case of Ca(II) it is instead the effect of suboptimal protein structure. PMID:23056605

Structure of the thermophilic l-Arabinose isomerase from Geobacillus kaustophilus reveals metal-mediated intersubunit interactions for activity and thermostability.

PubMed

Choi, Jin Myung; Lee, Yong-Jik; Cao, Thinh-Phat; Shin, Sun-Mi; Park, Min-Kyu; Lee, Han-Seung; di Luccio, Eric; Kim, Seong-Bo; Lee, Sang-Jae; Lee, Sang Jun; Lee, Sung Haeng; Lee, Dong-Woo

2016-04-15

Thermophilic l-arabinose isomerase (AI), which catalyzes the interconversion of l-arabinose and l-ribulose, can be used to produce d-tagatose, a sugar substitute, from d-galactose. Unlike mesophilic AIs, thermophilic AIs are highly dependent on divalent metal ions for their catalytic activity and thermostability at elevated temperatures. However, the molecular basis underlying the substrate preferences and metal requirements of multimeric AIs remains unclear. Here we report the first crystal structure of the apo and holo forms of thermophilic Geobacillus kaustophilus AI (GKAI) in hexamer form. The structures, including those of GKAI in complex with l-arabitol, and biochemical analyses revealed not only how the substrate-binding site of GKAI is formed through displacement of residues at the intersubunit interface when it is bound to Mn(2+), but also revealed the water-mediated H-bonding networks that contribute to the structural integrity of GKAI during catalysis. These observations suggest metal-mediated isomerization reactions brought about by intersubunit interactions at elevated temperatures are responsible for the distinct active site features that promote the substrate specificity and thermostability of thermophilic AIs. Copyright © 2016 Elsevier Inc. All rights reserved.

Surface enhanced Raman spectroscopy detection of biomolecules using EBL fabricated nanostructured substrates.

PubMed

Peters, Robert F; Gutierrez-Rivera, Luis; Dew, Steven K; Stepanova, Maria

2015-03-20

Fabrication and characterization of conjugate nano-biological systems interfacing metallic nanostructures on solid supports with immobilized biomolecules is reported. The entire sequence of relevant experimental steps is described, involving the fabrication of nanostructured substrates using electron beam lithography, immobilization of biomolecules on the substrates, and their characterization utilizing surface-enhanced Raman spectroscopy (SERS). Three different designs of nano-biological systems are employed, including protein A, glucose binding protein, and a dopamine binding DNA aptamer. In the latter two cases, the binding of respective ligands, D-glucose and dopamine, is also included. The three kinds of biomolecules are immobilized on nanostructured substrates by different methods, and the results of SERS imaging are reported. The capabilities of SERS to detect vibrational modes from surface-immobilized proteins, as well as to capture the protein-ligand and aptamer-ligand binding are demonstrated. The results also illustrate the influence of the surface nanostructure geometry, biomolecules immobilization strategy, Raman activity of the molecules and presence or absence of the ligand binding on the SERS spectra acquired.

Crystal structure of Yersinia pestis virulence factor YfeA reveals two polyspecific metal-binding sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Radka, Christopher D.; DeLucas, Lawrence J.; Wilson, Landon S.

2017-06-30

Gram-negative bacteria use siderophores, outer membrane receptors, inner membrane transporters and substrate-binding proteins (SBPs) to transport transition metals through the periplasm. The SBPs share a similar protein fold that has undergone significant structural evolution to communicate with a variety of differentially regulated transporters in the cell. InYersinia pestis, the causative agent of plague, YfeA (YPO2439, y1897), an SBP, is important for full virulence during mammalian infection. To better understand the role of YfeA in infection, crystal structures were determined under several environmental conditions with respect to transition-metal levels. Energy-dispersive X-ray spectroscopy and anomalous X-ray scattering data show that YfeA ismore » polyspecific and can alter its substrate specificity. In minimal-media experiments, YfeA crystals grown after iron supplementation showed a threefold increase in iron fluorescence emission over the iron fluorescence emission from YfeA crystals grown from nutrient-rich conditions, and YfeA crystals grown after manganese supplementation during overexpression showed a fivefold increase in manganese fluorescence emission over the manganese fluorescence emission from YfeA crystals grown from nutrient-rich conditions. In all experiments, the YfeA crystals produced the strongest fluorescence emission from zinc and could not be manipulated otherwise. Additionally, this report documents the discovery of a novel surface metal-binding site that prefers to chelate zinc but can also bind manganese. Flexibility across YfeA crystal forms in three loops and a helix near the buried metal-binding site suggest that a structural rearrangement is required for metal loading and unloading.« less

Proton and metal ion binding to natural organic polyelectrolytes-II. Preliminary investigation with a peat and a humic acid

USGS Publications Warehouse

Marinsky, J.A.; Reddy, M.M.

1984-01-01

We summarize here experimental studies of proton and metal ion binding to a peat and a humic acid. Data analysis is based on a unified physico-chemical model for reaction of simple ions with polyelectrolytes employing a modified Henderson-Hasselbalch equation. Peat exhibited an apparent intrinsic acid dissociation constant of 10-4.05, and an apparent intrinsic metal ion binding constant of: 400 for cadmium ion; 600 for zinc ion; 4000 for copper ion; 20000 for lead ion. A humic acid was found to have an apparent intrinsic proton binding constant of 10-2.6. Copper ion binding to this humic acid sample occurred at two types of sites. The first site exhibited reaction characteristics which were independent of solution pH and required the interaction of two ligands on the humic acid matrix to simultaneously complex with each copper ion. The second complex species is assumed to be a simple monodentate copper ion-carboxylate species with a stability constant of 18. ?? 1984.

Structures of rat cytosolic PEPCK: insight into the mechanism of phosphorylation and decarboxylation of oxaloacetic acid.

PubMed

Sullivan, Sarah M; Holyoak, Todd

2007-09-04

The structures of the rat cytosolic isoform of phosphoenolpyruvate carboxykinase (PEPCK) reported in the PEPCK-Mn2+, -Mn2+-oxaloacetic acid (OAA), -Mn2+-OAA-Mn2+-guanosine-5'-diphosphate (GDP), and -Mn2+-Mn2+-guanosine-5'-tri-phosphate (GTP) complexes provide insight into the mechanism of phosphoryl transfer and decarboxylation mediated by this enzyme. OAA is observed to bind in a number of different orientations coordinating directly to the active site metal. The Mn2+-OAA and Mn2+-OAA-Mn2+GDP structures illustrate inner-sphere coordination of OAA to the manganese ion through the displacement of two of the three water molecules coordinated to the metal in the holo-enzyme by the C3 and C4 carbonyl oxygens. In the PEPCK-Mn2+-OAA complex, an alternate bound conformation of OAA is present. In this conformation, in addition to the previous interactions, the C1 carboxylate is directly coordinated to the active site Mn2+, displacing all of the waters coordinated to the metal in the holo-enzyme. In the PEPCK-Mn2+-GTP structure, the same water molecule displaced by the C1 carboxylate of OAA is displaced by one of the gamma-phosphate oxygens of the triphosphate nucleotide. The structures are consistent with a mechanism of direct in-line phosphoryl transfer, supported by the observed stereochemistry of the reaction. In the catalytically competent binding mode, the C1 carboxylate of OAA is sandwiched between R87 and R405 in an environment that would serve to facilitate decarboxylation. In the reverse reaction, these two arginines would form the CO2 binding site. Comparison of the Mn2+-OAA-Mn2+GDP and Mn2+-Mn2+GTP structures illustrates a marked difference in the bound conformations of the nucleotide substrates in which the GTP nucleotide is bound in a high-energy state resulting from the eclipsing of all three of the phosphoryl groups along the triphosphate chain. This contrasts a previously determined structure of PEPCK in complex with a triphosphate nucleotide analogue in which the analogue mirrors the conformation of GDP as opposed to GTP. Last, the structures illustrate a correlation between conformational changes in the P-loop, the nucleotide binding site, and the active site lid that are important for catalysis.

Nanopore Device for Reversible Ion and Molecule Sensing or Migration

NASA Technical Reports Server (NTRS)

Seger, R. Adam (Inventor); Pourmand, Nader (Inventor); Actis, Paolo (Inventor); Singaram, Bakthan (Inventor); Vilozny, Boaz (Inventor)

2015-01-01

Disclosed are methods and devices for detection of ion migration and binding, utilizing a nanopipette adapted for use in an electrochemical sensing circuit. The nanopipette may be functionalized on its interior bore with metal chelators for binding and sensing metal ions or other specific binding molecules such as boronic acid for binding and sensing glucose. Such a functionalized nanopipette is comprised in an electrical sensor that detects when the nanopipette selectively and reversibly binds ions or small molecules. Also disclosed is a nanoreactor, comprising a nanopipette, for controlling precipitation in aqueous solutions by voltage-directed ion migration, wherein ions may be directed out of the interior bore by a repulsing charge in the bore.

Metal accumulation and differentially expressed proteins in gill of oyster (Crassostrea hongkongensis) exposed to long-term heavy metal-contaminated estuary.

PubMed

Luo, Lianzhong; Ke, Caihuan; Guo, Xiaoyu; Shi, Bo; Huang, Miaoqin

2014-06-01

Bio-accumulation and bio-transmission of toxic metals and the toxicological responses of organisms exposed to toxic metals have been focused, due to heavy metal contaminations have critically threatened the ecosystem and food security. However, still few investigations focused on the responses of certain organisms exposed to the long term and severe heavy metal contamination in specific environments. In present investigation, the Hong Kong oyster, Crassostrea hongkongensis were obtained from 3 sites which were contaminated by different concentrations of heavy metals (such as zinc, copper, manganese and lead etc.), respectively. Heavy metal concentrations in the sea water samples collected from the 3 sites and the dissected tissues of the oysters with blue visceral mass were determinated to estimate the metal contamination levels in environments and the bio-accumulation ratios of the heavy metals in the different tissues of oysters. Moreover, Proteomic methods were employed to analyze the differentially expressed proteins in the gills of oysters exposed to long-term heavy metal contaminations. Results indicated that the Jiulong River estuary has been severely contaminated by Cu, Zn and slightly with Cr, Ni, Mn, etc, moreover, Zn and Cu were the major metals accumulated by oysters to phenomenally high concentrations (more than 3.0% of Zn and about 2.0% of Cu against what the dry weight of tissues were accumulated), and Cr, Ni, Mn, etc were also significantly accumulated. The differentially expressed proteins in the gills of oysters exposed to heavy metals participate in several cell processes, such as metal binding, transporting and saving, oxidative-reduction balance maintaining, stress response, signal transduction, etc. Significantly up-regulated expression (about 10 folds) of an important metal binding protein, metallothionein (MT) and granular cells was observed in the gills of oysters exposed to long-term and severely heavy-metal-contaminated estuary, it suggested that binding toxic metals with metallothionein-like proteins (MTLP) and storing toxic metals in metal-rich granules (MRG) with insoluble forms were the important strategies of oyster to detoxify the toxic metals and adapt to the high level of metal-contaminated environment. Most of the stress and immunity responsive proteins, such as heat shock proteins (HSP), extracellular superoxide dismutase (ECSOD) and cavortin, and the cellular redox reaction relative proteins such as 20G-Fe (II) oxygenase family oxidoreductase, aldehyde dehydrogenase and retinal dehydrogenase 2, were detected significantly down-regulated in the gills of oysters exposed to long term heavy metal contaminated environments, it indicated that long term exposure different from emergent exposure to heavy metal contamination may significantly suppress the stress and immunity response system of oysters. Moreover, Formin homology 2 domain containing protein (FH2). The only protein domain to directly nucleate actin monomers into unbranched filament polymers, by which will subsequently control gene expression and chromatin remodelling complexes, was also detected greatly up-regulated in the gills of oysters exposed to long-term heavy metal contaminations. It indicated that nuclear activity regulation may also be important for oyster to adapt to the long-term heavy-metal-contaminated environment. Copyright © 2014 Elsevier Ltd. All rights reserved.

Specific binding of trivalent metal ions to λ-carrageenan.

PubMed

Cao, Yiping; Li, Shugang; Fang, Yapeng; Nishinari, Katsuyoshi; Phillips, Glyn O; Lerbret, Adrien; Assifaoui, Ali

2018-04-01

Carrageenans are a family of sulphated cell wall polysaccharides extracted from seaweeds and are widely used in different industrial sectors. Relative to κ-carrageenan (κ-car) and ι-carrageenan (ι-car), the ionic binding behavior of λ-carrageenan (λ-car) is far less studied. In this work, the interaction and binding behavior between λ-car and metal ions of different valency (Na + , K + , Mg 2+ , Ca 2+ , Fe 2+ , Fe 3+ , Al 3+ , Cr 3+ ) have been investigated. In contrast to the non-specific interaction of the monovalent and divalent cations, specific binding has been identified between λ-car and Fe 3+ /Al 3+ . The specific binding could lead to either precipitation or gelation of λ-car, depending on the way of introducing Fe 3+ /Al 3+ ions. Fe 3+ and Al 3+ exhibit the same binding stoichiometry of [M 3+ ]/[repeating unit] = 1.0, with the former having a relatively larger binding constant. Cr 3+ , though having very similar physical properties with Fe 3+ /Al 3+ , is incapable of binding specifically to Cr 3+ . The phenomena could not be interpreted in terms of counterion condensation, and are rather attributable to a mechanism in which hexa-coordination of Fe 3+ /Al 3+ and entropy-driven cation dehydration play crucial roles in driving the binding of the trivalent metal ions to λ-car. Copyright © 2017 Elsevier B.V. All rights reserved.

An unconventional origin of metal-ion rescue and inhibition in the Tetrahymena group I ribozyme reaction.

PubMed Central

Shan, S O; Herschlag, D

2000-01-01

The presence of catalytic metal ions in RNA active sites has often been inferred from metal-ion rescue of modified substrates and sometimes from inhibitory effects of alternative metal ions. Herein we report that, in the Tetrahymena group I ribozyme reaction, the deleterious effect of a thio substitution at the pro-Sp position of the reactive phosphoryl group is rescued by Mn2+. However, analysis of the reaction of this thio substrate and of substrates with other modifications strongly suggest that this rescue does not stem from a direct Mn2+ interaction with the Sp sulfur. Instead, the apparent rescue arises from a Mn2+ ion interacting with the residue immediately 3' of the cleavage site, A(+1), that stabilizes the tertiary interactions between the oligonucleotide substrate (S) and the active site. This metal site is referred to as site D herein. We also present evidence that a previously observed Ca2+ ion that inhibits the chemical step binds to metal site D. These and other observations suggest that, whereas the interactions of Mn2+ at site D are favorable for the chemical reaction, the Ca2+ at site D exerts its inhibitory effect by disrupting the alignment of the substrates within the active site. These results emphasize the vigilance necessary in the design and interpretation of metal-ion rescue and inhibition experiments. Conversely, in-depth mechanistic analysis of the effects of site-specific substrate modifications can allow the effects of specific metal ion-RNA interactions to be revealed and the properties of individual metal-ion sites to be probed, even within the sea of metal ions bound to RNA. PMID:10864040

Homogeneous and Supported Niobium Catalysts as Lewis Acid and Radical Catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wayne Tikkanen

2006-12-31

The synthesis of tetrachlorotetraphenylcyclopentadienyl group 5 metal complexes has been accomplished through two routes, one a salt metathesis with lithiumtetraphenylcyclopentadiende and the other, reaction with trimethyltintetraphenylcyclopentadiene. The reactants and products have been characterized by {sup 1}H and {sup 13}C({sup 1}H) NMR spectroscopy. The niobium complex promotes the silylcyanation of butyraldehyde. The grafting of metal complexes to silica gel surfaces has been accomplished using tetrakisdimethylamidozirconium as the metal precursor. The most homogeneous binding as determined by CP-MAS {sup 13}C NMR and infrared spectroscopy was obtained with drying at 500 C at 3 mtorr vacuum. The remaining amido groups can be replacedmore » by reaction with alcohols to generate surface bound metal alkoxides. These bound catalysts promote silylcyanation of aryl aldehydes and can be reused three times with no loss of activity.« less

Spectral and in vitro antimicrobial properties of 2-oxo-4-phenyl-6-styryl-1,2,3,4-tetrahydro-pyrimidine-5-carboxylic acid transition metal complexes

NASA Astrophysics Data System (ADS)

Dhankar, Raksha P.; Rahatgaonkar, Anjali M.; Chorghade, Mukund S.; Tiwari, Ashutosh

2-oxo-4-phenyl-6-styryl-1,2,3,4-tetrahydro-pyrimidine-5-carboxylic acid (ADP) was complexed with acetates of Mn(II), Ni(II), Cu(II) and Zn(II). The structures of the ligand and its metal complexes were characterized by microanalysis, IR, NMR, UV-vis spectroscopy, magnetic susceptibility and TGA-DTA analyses. Octahedral and square planar geometries were suggested for the complexes in which the central metal ion coordinated with sbnd O donors of ligand and acetate ions. Each ligand binds the metal using carboxylate oxygens. The ligand and complexes were evaluated for their antimicrobial activities against different species of pathogenic bacteria and fungi. The present novel pyrimidine containing complexes could constitute a new group of antibacterial and antifungal agents.

DNA Binding, Cleavage and Antibacterial Activity of Mononuclear Cu(II), Ni(II) and Co(II) Complexes Derived from Novel Benzothiazole Schiff Bases.

PubMed

Vamsikrishna, Narendrula; Kumar, Marri Pradeep; Tejaswi, Somapangu; Rambabu, Aveli; Shivaraj

2016-07-01

A series of novel bivalent metal complexes M(L1)2 and M(L2)2 where M = Cu(II), Ni(II), Co(II) and L1 = 2-((benzo [d] thiazol-6-ylimino)methyl)-4-bromophenol [BTEMBP], L2 = 1-((benzo [d] thiazol-6-ylimino)methyl) naphthalen-2-ol [BTEMNAPP] were synthesized. All the compounds have been characterized by elemental analysis, SEM, Mass, (1)H NMR, (13)C NMR, UV-Vis, IR, ESR, spectral data and magnetic susceptibility measurements. Based on the analytical and spectral data four-coordinated square planar geometry is assigned to all the complexes. DNA binding properties of these complexes have been investigated by electronic absorption spectroscopy, fluorescence and viscosity measurements. It is observed that these binary complexes strongly bind to calf thymus DNA by an intercalation mode. DNA cleavage efficacy of these complexes was tested in presence of H2O2 and UV light by gel electrophoresis and found that all the complexes showed better nuclease activity. Finally the compounds were screened for antibacterial activity against few pathogens and found that the complexes have potent biocidal activity than their free ligands.

Transition-metal prion protein attachment: Competition with copper

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Bernholc, Jerry

2012-02-01

Prion protein, PrP, is a protein capable of binding copper ions in multiple modes depending on their concentration. Misfolded PrP is implicated in a group of neurodegenerative diseases, which include ``mad cow disease'' and its human form, variant Creutzfeld-Jacob disease. An increasing amount of evidence suggests that attachment of non-copper metal ions to PrP triggers transformations to abnormal forms similar to those observed in prion diseases. In this work, we use hybrid Kohn-Sham/orbital-free density functional theory simulations to investigate copper replacement by other transition metals that bind to PrP, including zinc, iron and manganese. We consider all known copper binding modes in the N-terminal domain of PrP. Our calculations identify modes most susceptible to copper replacement and reveal metals that can successfully compete with copper for attachment to PrP.

Binding of uridine 5'-diphosphate in the "basic patch" of the zinc deacetylase LpxC and implications for substrate binding.

PubMed

Gennadios, Heather A; Christianson, David W

2006-12-26

LpxC is a zinc metalloenzyme that catalyzes the first committed step in the biosynthesis of lipid A, a vital component of the outer membrane of Gram-negative bacteria. Accordingly, the inhibition of LpxC is an attractive strategy for the treatment of Gram-negative bacterial infections. Here, we report the 2.7 A resolution X-ray crystal structure of LpxC from Aquifex aeolicus complexed with uridine 5'-diphosphate (UDP), and the 3.1 A resolution structure of LpxC complexed with pyrophosphate. The X-ray crystal structure of the LpxC-UDP complex provides the first view of interactions likely to be exploited by the substrate UDP group in the "basic patch" of the active site. The diphosphate group of UDP makes hydrogen bond interactions with strictly conserved residue K239 as well as solvent molecules. The ribose moiety of UDP interacts with partially conserved residue E197. The UDP uracil group hydrogen bonds with both the backbone NH group and the backbone carbonyl group of E160, and with the backbone NH group of K162 through an intervening water molecule. Finally, the alpha-phosphate and uracil groups of UDP interact with R143 and R262 through intervening water molecules. The structure of LpxC complexed with pyrophosphate reveals generally similar intermolecular interactions in the basic patch. Unexpectedly, diphosphate binding in both complexes is accompanied by coordination to an additional zinc ion, resulting in the identification of a new metal-binding site termed the E-site. The structures of the LpxC-UDP and LpxC-pyrophosphate complexes provide new insights with regard to substrate recognition in the basic patch and metal ion coordination in the active site of LpxC.

Vanadium-Binding Ability of Nucleoside Diphosphate Kinase from the Vanadium-Rich Fan Worm, Pseudopotamilla occelata.

PubMed

Yamaguchi, Nobuo; Yoshinaga, Masafumi; Kamino, Kei; Ueki, Tatsuya

2016-06-01

Polychaete fan worms and ascidians accumulate high levels of vanadium ions. Several vanadiumbinding proteins, known as vanabins, have been found in ascidians. However, no vanadium-binding factors have been isolated from the fan worm. In the present study, we sought to identify vanadiumbinding proteins in the branchial crown of the fan worm using immobilized metal ion affinity chromatography. A nucleoside diphosphate kinase (NDK) homolog was isolated and determined to be a vanadium-binding protein. Kinase activity of the NDK homologue, PoNDK, was suppressed by the addition of V(IV), but was unaffected by V(V). The effect of V(IV) on PoNDK precedes its activation by Mg(II). This is the first report to describe the relationship between NDK and V(IV). PoNDK is located in the epidermis of the branchial crown, and its distribution is very similar to that of vanadium. These results suggest that PoNDK is associated with vanadium accumulation and metabolism in P. occelata.

Ligand-receptor assay for evaluation of functional activity of human recombinant VEGF and VEGFR-1 extracellular fragment.

PubMed

Leopol'd, A V; Baklaushev, V P; Korchagina, A A; Shein, S A; Grinenko, N F; Pavlov, K A; Ryabukhin, I A; Chekhonin, V P

2012-04-01

cDNA encoding VEGF and Ig-like extracellular domains 2-4 of VEGFR-1 (sFlt-1(2-4)) were cloned into prokaryotic expression vectors pET32a and pQE60. Recombinant proteins were purified (metal affinity chromatography) and renatured. Chemiluminescent study for the interaction of recombinant VEGF and sFlt-1(2-4) showed that biotinylated VEGF specifically binds to the polystyrene-immobilized receptor extracellular fragment. Biotinylated recombinant sFlt-1 interacts with immobilized VEGF. Analysis of the interaction of immobilized recombinant VEGFR-1 and VEGF with C6 glioma cells labeled with CFDA-SE (vital fluorescent dye) showed that recombinant VEGFR-1 also binds to native membrane-associated VEGF. Recombinant VEGF was shown to bind to specific receptors expressed on the surface of C6 glioma cells. Functional activity of these proteins was confirmed by ligand-receptor assay for VEGF and VEGFR-1 (sFlt-1) and quantitative chemiluminescent detection.

Immobilized metal ion affinity electrophoresis. A study with several model proteins containing histidine.

PubMed

Goubran-Botros, H; Nanak, E; Abdul Nour, J; Birkenmeir, G; Vijayalakshmi, M A

1992-04-24

Immobilized metal ion affinity electrophoresis (IMA-Elec) is one among the many methods derived from the immobilized metal ion affinity chromatography. Two approaches for incorporating the metal ligand, were studied. One was in the form of insoluble particulate material based on Sepharose 6B and the other in the form of soluble polymer based on polyethylene glycol (PEG) 5000. Both the polymers coupled with iminodiacetate and metallized with copper or zinc were used as ligands, incorporated into soluble agarose as the electrophoretic gel. Several histidine-containing model proteins were studied with both the systems and their metal binding strengths were determined as the dissociation constants, Kd. The results clearly demonstrated that the mechanism of protein recognition by immobilized copper or zinc via the accessible histidyl residues was maintained in the IMA-Elec system. Proteins with increasing numbers of histidine residues showed increasing binding strength (lower Kd values). While this basic mechanism was conserved, the supporting polymers (Sepharose 6B and the PEG 5000) showed significant differences in the metal binding to the protein. The polysaccharide Sepharose 6B enhanced the binding strength compared with PEG 5000. The optimum electrophoretic parameters were determined to be current intensities up to 20 mA and pH ca. 7.0. At pH greater than 8.0, a significant decrease in the affinity was observed, this decrease being greater with PEG 5000 than Sepharose 6B as supporting material.

DJ-1 Is a Copper Chaperone Acting on SOD1 Activation*

PubMed Central

Girotto, Stefania; Cendron, Laura; Bisaglia, Marco; Tessari, Isabella; Mammi, Stefano; Zanotti, Giuseppe; Bubacco, Luigi

2014-01-01

Lack of oxidative stress control is a common and often prime feature observed in many neurodegenerative diseases. Both DJ-1 and SOD1, proteins involved in familial Parkinson disease and amyotrophic lateral sclerosis, respectively, play a protective role against oxidative stress. Impaired activity and modified expression of both proteins have been observed in different neurodegenerative diseases. A potential cooperative action of DJ-1 and SOD1 in the same oxidative stress response pathway may be suggested based on a copper-mediated interaction between the two proteins reported here. To investigate the mechanisms underlying the antioxidative function of DJ-1 in relation to SOD1 activity, we investigated the ability of DJ-1 to bind copper ions. We structurally characterized a novel copper binding site involving Cys-106, and we investigated, using different techniques, the kinetics of DJ-1 binding to copper ions. The copper transfer between the two proteins was also examined using both fluorescence spectroscopy and specific biochemical assays for SOD1 activity. The structural and functional analysis of the novel DJ-1 copper binding site led us to identify a putative role for DJ-1 as a copper chaperone. Alteration of the coordination geometry of the copper ion in DJ-1 may be correlated to the physiological role of the protein, to a potential failure in metal transfer to SOD1, and to successive implications in neurodegenerative etiopathogenesis. PMID:24567322

Allosteric modulation of ATP-gated P2X receptor channels

PubMed Central

Coddou, Claudio; Stojilkovic, Stanko S.; Huidobro-Toro, J. Pablo

2013-01-01

Seven mammalian purinergic receptor subunits, denoted P2X1 to P2X7, and several spliced forms of these subunits have been cloned. When heterologously expressed, these cDNAs encode ATP-gated non-selective cation channels organized as trimers. All activated receptors produce cell depolarization and promote Ca2+ influx through their pores and indirectly by activating voltage-gated calcium channels. However, the biophysical and pharmacological properties of these receptors differ considerably, and the majority of these subunits are also capable of forming heterotrimers with other members of the P2X receptor family, which confers further different properties. These channels have three ATP binding domains, presumably located between neighboring subunits, and occupancy of at least two binding sites is needed for their activation. In addition to the orthosteric binding sites for ATP, these receptors have additional allosteric sites that modulate the agonist action at receptors, including sites for trace metals, protons, neurosteroids, reactive oxygen species and phosphoinositides. The allosteric regulation of P2X receptors is frequently receptor-specific and could be a useful tool to identify P2X members in native tissues and their roles in signaling. The focus of this review is on common and receptor-specific allosteric modulation of P2X receptors and the molecular base accounting for allosteric binding sites. PMID:21639805

Mutation in the β-hairpin of the Bordetella pertussis adenylate cyclase toxin modulates N-lobe conformation in calmodulin.

PubMed

Springer, Tzvia I; Goebel, Erich; Hariraju, Dinesh; Finley, Natosha L

2014-10-10

Bordetella pertussis, causative agent of whooping cough, produces an adenylate cyclase toxin (CyaA) that is an important virulence factor. In the host cell, the adenylate cyclase domain of CyaA (CyaA-ACD) is activated upon association with calmodulin (CaM), an EF-hand protein comprised of N- and C-lobes (N-CaM and C-CaM, respectively) connected by a flexible tether. Maximal CyaA-ACD activation is achieved through its binding to both lobes of intact CaM, but the structural mechanisms remain unclear. No high-resolution structure of the intact CaM/CyaA-ACD complex is available, but crystal structures of isolated C-CaM bound to CyaA-ACD shed light on the molecular mechanism by which this lobe activates the toxin. Previous studies using molecular modeling, biochemical, and biophysical experiments demonstrate that CyaA-ACD's β-hairpin participates in site-specific interactions with N-CaM. In this study, we utilize nuclear magnetic resonance (NMR) spectroscopy to probe the molecular association between intact CaM and CyaA-ACD. Our results indicate binding of CyaA-ACD to CaM induces large conformational perturbations mapping to C-CaM, while substantially smaller structural changes are localized primarily to helices I, II, and IV, and the metal-binding sites in N-CaM. Site-specific mutations in CyaA-ACD's β-hairpin structurally modulate N-CaM, resulting in conformational perturbations in metal binding sites I and II, while no significant structural modifications are observed in C-CaM. Moreover, dynamic light scattering (DLS) analysis reveals that mutation of the β-hairpin results in a decreased hydrodynamic radius (Rh) and reduced thermal stability in the mutant complex. Taken together, our data provide new structural insights into the β-hairpin's role in stabilizing interactions between CyaA-ACD and N-CaM. Copyright © 2014 Elsevier Inc. All rights reserved.

First report of phytochelatins in a mushroom: induction of phytochelatins by metal exposure in Boletus edulis.

PubMed

Collin-Hansen, Christian; Pedersen, Sindre A; Andersen, Rolf A; Steinnes, Eiliv

2007-01-01

Some species of macromycetes (mushrooms) consistently are found to contain high concentrations of toxic metals such as cadmium (Cd) and mercury (Hg), and consumption of wild-growing mushrooms is acknowledged as a significant source for Cd and Hg in humans. Yet little is known about the speciation of Cd and Hg in mushroom tissues. Here we present the first evidence of peptides of the phytochelatin family being responsible for binding a large fraction of Cd in caps of the macromycete Boletus edulis exposed to excess metals. Concentrations of Cd, Zn, Cu and Hg, as well as cytosolic Cd-binding capacity (CCBC), glutathione (GSH) and free proline (Pro) were quantified in fruiting bodies of B. edulis differentially exposed to a wide range of metals. Metal distribution among cytosolic compounds were investigated by size exclusion chromatography (SEC), followed by metal determinations with atomic absorption chromatography (AAS) and HR-ICP-MS. Cd-binding compounds in SEC elutates were investigated further by high performance liquid chromatography-mass spectrometry (HPLC-MS). CCBC was >90 times higher in the exposed group relative to the reference group (Mann-Whitney's P < 0.001), whereas concentrations of free Pro were almost identical for the two groups. For the whole study selection, CCBC correlated positively with metal exposure (Spearman's P < 0.001 for all four metals), suggesting dose-dependent induction of Cd-binding compounds by exposure to these metals, possibly as a defense mechanism. The presence of phytochelatins (PCs), a family of cystein-rich oligopeptides, was confirmed in Cd-containing SEC fractions by HPLC-MS. The appearance of more complex PCs was coupled to declining concentrations of GSH. To our knowledge this is the first report demonstrating the presence of PCs in a macromycete.

Gastrin Receptor-Avid Peptide Conjugates

DOEpatents

Hoffman, Timothy J.; Volkert, Wynn A.; Li, Ning; Sieckman, Gary; Higginbotham, Chrys-Ann

2005-07-26

A compound for use as a therapeutic or diagnostic radiopharmaceutical includes a group capable of complexing a medically useful metal attached to a moiety which is capable of binding to a gastrin releasing peptide receptor. A method for treating a subject having a neoplastic disease includes administering to the subject an effective amount of a radiopharmaceutical having a metal chelated with a chelating group attached to a moiety capable of binding to a gastrin releasing peptide receptor expressed on tumor cells with subsequent internalization inside of the cell. A method of forming a therapeutic or diagnostic compound includes reacting a metal synthon with a chelating group covalently linked with a moiety capable of binding a gastrin releasing peptide receptor.

Gastrin receptor-avid peptide conjugates

DOEpatents

Hoffman, Timothy J.; Volkert, Wynn A.; Li, Ning; Sieckman, Gary; Higginbotham, C. A.

2001-01-01

A compound for use as a therapeutic or diagnostic radiopharmaceutical includes a group capable of complexing a medically useful metal attached to a moiety which is capable of binding to a gastrin releasing peptide receptor. A method for treating a subject having a neoplastic disease includes administering to the subject an effective amount of a radiopharmaceutical having a metal chelated with a chelating group attached to a moiety capable of binding to a gastrin releasing peptide receptor expressed on tumor cells with subsequent internalization inside of the cell. A method of forming a therapeutic or diagnostic compound includes reacting a metal synthon with a chelating group covalently linked with a moiety capable of binding a gastrin releasing peptide receptor.

Gastrin receptor-avid peptide conjugates

DOEpatents

Hoffman, Timothy J.; Volkert, Wynn A.; Sieckman, Gary; Smith, Charles J.; Gali, Hariprasad

2006-06-13

A compound for use as a therapeutic or diagnostic radiopharmaceutical includes a group capable of complexing a medically useful metal attached to a moiety which is capable of binding to a gastrin releasing peptide receptor. A method for treating a subject having a neoplastic disease includes administering to the subject an effective amount of a radiopharmaceutical having a metal chelated with a chelating group attached to a-moiety capable of binding to a gastrin releasing peptide receptor expressed on tumor cells with subsequent internalization inside of the cell. A method of forming a therapeutic or diagnostic compound includes reacting a metal synthon with a chelating group covalently linked with a moiety capable of binding a gastrin releasing peptide receptor.

Gastrin receptor-avid peptide conjugates

DOEpatents

Hoffman, Timothy J.; Volkert, Wynn A.; Li, Ning; Sieckman, Gary; Higginbotham, Chrys-Ann

2006-12-12

A compound for use as a therapeutic or diagnostic radiopharmaceutical includes a group capable of complexing a medically useful metal attached to a moiety which is capable of binding to a gastrin releasing peptide receptor. A method for treating a subject having a neoplastic disease includes administering to the subject an effective amount of a radiopharmaceutical having a metal chelated with a chelating group attached to a moiety capable of binding to a gastrin releasing peptide receptor expressed on tumor cells with subsequent internalization inside of the cell. A method of forming a therapeutic or diagnostic compound includes reacting a metal synthon with a chelating group covalently linked with a moiety capable of binding a gastrin releasing peptide receptor.