Surfactant biocatalyst for remediation of recalcitrant organics and heavy metals

DOEpatents

Brigmon, Robin L [North Augusta, SC; Story, Sandra [Greenville, SC; Altman, Denis J [Evans, GA; Berry, Christopher J [Aiken, SC

2011-05-03

Novel strains of isolated and purified bacteria have been identified which have the ability to degrade petroleum hydrocarbons including a variety of PAHs. Several isolates also exhibit the ability to produce a biosurfactant. The combination of the biosurfactant-producing ability along with the ability to degrade PAHs enhances the efficiency with which PAHs may be degraded. Additionally, the biosurfactant also provides an additional ability to bind heavy metal ions for removal from a soil or aquatic environment.

Surfactant biocatalyst for remediation of recalcitrant organics and heavy metals

DOEpatents

Brigmon, Robin L [North Augusta, SC; Story, Sandra [Greenville, SC; Altman,; Denis, J [Evans, GA; Berry, Christopher J [Aiken, SC

2011-03-29

Novel strains of isolated and purified bacteria have been identified which have the ability to degrade petroleum hydrocarbons including a variety of PAHs. Several isolates also exhibit the ability to produce a biosurfactant. The combination of the biosurfactant-producing ability along with the ability to degrade PAHs enhances the efficiency with which PAHs may be degraded. Additionally, the biosurfactant also provides an additional ability to bind heavy metal ions for removal from a soil or aquatic environment.

Combined copper/zinc attachment to prion protein

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Bernholc, Jerry

2013-03-01

Misfolding of prion protein (PrP) is responsible for diseases such as ``mad-cow disease'' in cattle and Creutzfeldt-Jacob in humans. Extensive experimental investigation has established that this protein strongly interacts with copper ions, and this ability has been linked to its still unknown function. Attachment of other metal ions (zinc, iron, manganese) have been demonstrated as well, but none of them could outcompete copper. Recent finding, however, indicates that at intermediate concentrations both copper and zinc ions can attach to the PrP at the octarepeat region, which contains high affinity metal binding sites. Based on this evidence, we have performed density functional theory simulations to investigate the combined Cu/Zn attachment. We consider all previously reported binding modes of copper at the octarepeat region and examine a possibility simultaneous Cu/Zn attachment. We find that this can indeed occur for only one of the known binding sites, when copper changes its coordination mode to allow for attachment of zinc ion. The implications of the simultaneous attachment on neural function remain to be explored.

Interactions between Hofmeister anions and the binding pocket of a protein.

PubMed

Fox, Jerome M; Kang, Kyungtae; Sherman, Woody; Héroux, Annie; Sastry, G Madhavi; Baghbanzadeh, Mostafa; Lockett, Matthew R; Whitesides, George M

2015-03-25

This paper uses the binding pocket of human carbonic anhydrase II (HCAII, EC 4.2.1.1) as a tool to examine the properties of Hofmeister anions that determine (i) where, and how strongly, they associate with concavities on the surfaces of proteins and (ii) how, upon binding, they alter the structure of water within those concavities. Results from X-ray crystallography and isothermal titration calorimetry show that most anions associate with the binding pocket of HCAII by forming inner-sphere ion pairs with the Zn(2+) cofactor. In these ion pairs, the free energy of anion-Zn(2+) association is inversely proportional to the free energetic cost of anion dehydration; this relationship is consistent with the mechanism of ion pair formation suggested by the "law of matching water affinities". Iodide and bromide anions also associate with a hydrophobic declivity in the wall of the binding pocket. Molecular dynamics simulations suggest that anions, upon associating with Zn(2+), trigger rearrangements of water that extend up to 8 Å away from their surfaces. These findings expand the range of interactions previously thought to occur between ions and proteins by suggesting that (i) weakly hydrated anions can bind complementarily shaped hydrophobic declivities, and that (ii) ion-induced rearrangements of water within protein concavities can (in contrast with similar rearrangements in bulk water) extend well beyond the first hydration shells of the ions that trigger them. This study paints a picture of Hofmeister anions as a set of structurally varied ligands that differ in size, shape, and affinity for water and, thus, in their ability to bind to—and to alter the charge and hydration structure of—polar, nonpolar, and topographically complex concavities on the surfaces of proteins.

Divalent ions are potential permeating blockers of the non-selective NaK ion channel: combined QM and MD based investigations.

PubMed

Sadhu, Biswajit; Sundararajan, Mahesh; Bandyopadhyay, Tusar

2017-10-18

The bacterial NaK ion channel is distinctly different from other known ion channels due to its inherent non-selective feature. One of the unexplored and rather interesting features is its ability to permeate divalent metal ions (such as Ca 2+ and Ba 2+ ) and not monovalent alkali metal ions. Several intriguing questions about the energetics and structural aspects still remain unanswered. For instance, what causes Ca 2+ to permeate as well as block the selectivity filter (SF) of the NaK ion channel and act as a "permeating blocker"? How and at what energetic cost does another chemical congener, Sr 2+ , as well as Ba 2+ , a potent blocker of the K + ion channel, permeate through the SF of the NaK ion channel? Finally, how do their translocation energetics differ from those of monovalent ions such as K + ? Here, in an attempt to address these outstanding issues, we elucidate the structure, binding and selectivity of divalent ions (Ca 2+ , Sr 2+ and Ba 2+ ) as they permeate through the SF of the NaK ion channel using all-atom molecular dynamics simulations and density functional theory based calculations. We unveil mechanistic insight into this translocation event using well-tempered metadynamics simulations in a polarizable environment using the mean-field model of water and incorporating electronic continuum corrections for ions via charge rescaling. The results show that, akin to K + coordination, Sr 2+ and Ba 2+ bind at the SF in a very similar fashion and remain octa-coordinated at all sites. Interestingly, differing from its local hydration structure, Ca 2+ interacts with eight carbonyls to remain at the middle of the S3 site. Furthermore, the binding of divalent metals at SF binding sites is more favorable than the binding of K + . However, their permeation through the extracellular entrance faces a considerably higher energetic barrier compared to that for K + , which eventually manifests their inherent blocking feature.

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

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

Inert Reassessment Document for Ethylenediaminetetracetric acid (EDTA)

EPA Pesticide Factsheets

EDTA is a chelating agent. Its ability to bind heavy metal ions can be used to sequester these trace metals. However, trace amounts of various metals are necessary for the proper functioning of the body.

Hydrogen ion block of the sodium pore in squid giant axons

PubMed Central

1983-01-01

The block of squid axon sodium channels by H ions was studied using voltage-clamp and internal perfusion techniques. An increase in the concentration of internal permeant ions decreased the block produced by external H ions. The voltage dependence of the block was found to be nonmonotonic: it was reduced by both large positive and large negative potentials. The ability of internal ions to modify the block by external H+ is explained by a competition among these ions for a binding site within the pore. The nonmonotonic voltage dependence is consistent with this picture if the hydrogen ions are allowed to be permeant. PMID:6315859

Single-Molecule Patch-Clamp FRET Anisotropy Microscopy Studies of NMDA Receptor Ion Channel Activation and Deactivation under Agonist Ligand Binding in Living Cells.

PubMed

Sasmal, Dibyendu Kumar; Yadav, Rajeev; Lu, H Peter

2016-07-20

N-methyl-d-aspartate (NMDA) receptor ion channel is activated by the binding of two pairs of glycine and glutamate along with the application of action potential. Binding and unbinding of ligands changes its conformation that plays a critical role in the open-close activities of NMDA receptor. Conformation states and their dynamics due to ligand binding are extremely difficult to characterize either by conventional ensemble experiments or single-channel electrophysiology method. Here we report the development of a new correlated technical approach, single-molecule patch-clamp FRET anisotropy imaging and demonstrate by probing the dynamics of NMDA receptor ion channel and kinetics of glycine binding with its ligand binding domain. Experimentally determined kinetics of ligand binding with receptor is further verified by computational modeling. Single-channel patch-clamp and four-channel fluorescence measurement are recorded simultaneously to get correlation among electrical on and off states, optically determined conformational open and closed states by FRET, and binding-unbinding states of the glycine ligand by anisotropy measurement at the ligand binding domain of GluN1 subunit. This method has the ability to detect the intermediate states in addition to electrical on and off states. Based on our experimental results, we have proposed that NMDA receptor gating goes through at least one electrically intermediate off state, a desensitized state, when ligands remain bound at the ligand binding domain with the conformation similar to the fully open state.

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.

Identification of gold sensing peptide by integrative proteomics and a bacterial two-component system

NASA Astrophysics Data System (ADS)

Ng, I.-Son; Yu, You-Jin; Yi, Ying-Chen; Tan, Shih-I.; Huang, Bo-Chuan; Han, Yin-Lung

2017-12-01

The proteomics strategy was utilized to analyze and identify the gold adsorption proteins from Tepidimonas fonticaldi AT-A2, due to its outstanding performance in gold-binding and recovery. The results showed that three small proteins, including histidine biosynthesis protein (HisIE), iron donor protein (CyaY) and hypothetical protein_65aa, have a higher ability to adsorb gold ions because of the negatively charged domains or metal binding sites. On the other hand, the Salmonella PmrA/PmrB two-component system first replaces the iron (III)-binding motif using the peptide sequence from hypothetical protein_65aa, and this is then used to reveal the sensing and responsiveness to gold metal ions, which is totally different from the performance of traditional gold binding peptide (GBP) on the crystals on the surface of gold (111). We have successfully demonstrated an integrative proteomics and bacterial two-component system to explore the novel gold binding peptide. Finally, the heterologous over-expression of gold binding peptide by E. coli and the equilibrium of binding capacity for Au(III) have been conducted.

Occupancy of a C2-C2 type 'zinc-finger' protein domain by copper. Direct observation by electrospray ionization mass spectrometry.

PubMed

Hutchens, T W; Allen, M H; Li, C M; Yip, T T

1992-09-07

The metal ion specificity of most 'zinc-finger' metal binding domains is unknown. The human estrogen receptor protein contains two different C2-C2 type 'zinc-finger' sequences within its DNA-binding domain (ERDBD). Copper inhibits the function of this protein by mechanisms which remain unclear. We have used electrospray ionization mass spectrometry to evaluate directly the 71-residue ERDBD (K180-M250) in the absence and presence of Cu(II) ions. The ERDBD showed a high affinity for Cu and was completely occupied with 4 Cu bound; each Cu ion was evidently bound to only two ligand residues (net loss of only 2 Da per bound Cu). The Cu binding stoichiometry was confirmed by atomic absorption. These results (i) provide the first direct physical evidence for the ability of the estrogen receptor DNA-binding domain to bind Cu and (ii) document a twofold difference in the Zn- and Cu-binding capacity. Differences in the ERDBD domain structure with bound Zn and Cu are predicted. Given the relative intracellular contents of Zn and Cu, our findings demonstrate the need to investigate further the Cu occupancy of this and other zinc-finger domains both in vitro and in vivo.

Morphology variation, composition alteration and microstructure changes in ion-irradiated 1060 aluminum alloy

NASA Astrophysics Data System (ADS)

Wan, Hao; Si, Naichao; Wang, Quan; Zhao, Zhenjiang

2018-02-01

Morphology variation, composition alteration and microstructure changes in 1060 aluminum irradiated with 50 keV helium ions were characterized by field emission scanning electron microscopy (FESEM) equipped with x-ray elemental scanning, 3D measuring laser microscope and transmission electron microscope (TEM). The results show that, helium ions irradiation induced surface damage and Si-rich aggregates in the surfaces of irradiated samples. Increasing the dose of irradiation, more damages and Si-rich aggregates would be produced. Besides, defects such as dislocations, dislocation loops and dislocation walls were the primary defects in the ion implanted layer. The forming of surface damages were related with preferentially sputtering of Al component. While irradiation-enhanced diffusion and irradiation-induced segregation resulted in the aggregation of impurity atoms. And the aggregation ability of impurity atoms were discussed based on the atomic radius, displacement energy, lattice binding energy and surface binding energy.

Benzophenone based fluorophore for selective detection of Sn2+ ion: Experimental and theoretical study.

PubMed

Jadhav, Amol G; Shinde, Suvidha S; Lanke, Sandip K; Sekar, Nagaiyan

2017-03-05

Synthesis of novel benzophenone-based chemosensor is presented for the selective sensing of Sn 2+ ion. Screening of competitive metal ions was performed by competitive experiments. The specific cation recognition ability of chemosensor towards Sn 2+ was investigated by experimental (UV-visible, fluorescence spectroscopy, 1 H NMR, 13 C NMR, FTIR and HRMS) methods and further supported by Density Functional Theory study. The stoichiometric binding ratio and binding constant (K a ) for complex is found to be 1:1 and 1.50×10 4 , respectively. The detection limit of Sn 2+ towards chemosensor was found to be 0.3898ppb. Specific selectivity and superiority of chemosensor over another recently reported chemosensor is presented. Copyright © 2016 Elsevier B.V. All rights reserved.

Inactivation of phosphorylase b by potassium ferrate, a new reactive analogue of the phosphate group.

PubMed

Lee, Y M; Benisek, W F

1976-03-25

Rabbit muscle phosphorylase b reacts with the phosphate-like reagent potassium ferrate, K2FeO4, a potent oxidizing agent. The reaction results in inactivation of the enzyme and abolition of the ability of the enzyme to bind 5'-AMP. Activating and nonactivating nucleotides which bind at the 5'-AMP binding site such as 5'-AMP, 2'-AMP, 3'-AMP, and 5'-IMP substantially protect the enzyme from inactivation by ferrate. One to two residues of tyrosine and approximately 1 residue of cysteine are modified by ferrate under the conditions employed. Tyrosine is protected by 5-AMP, whereas cysteine is not. The tyrosine modification is suggested as the inactivating chemical reaction. The location of the inactivating reaction is suggested to be in or near the 5'-AMP binding site. The structural and chemical properties of ferrate ion are discussed and compared to those of phosphate. Ferrate ion may be a reagent useful for phosphate group binding site-directed modification of proteins.

Na+ Shows a Markedly Higher Potential than K+ in DNA Compaction in a Crowded Environment

PubMed Central

Zinchenko, Anatoly A.; Yoshikawa, Kenichi

2005-01-01

Whereas many physicochemical investigations have shown that among monovalent cations Na+ ion possesses minimal potential for DNA binding, biological assays have shown that Na+ ion (in contrast to K+ ion) plays a primary role in chromatin compaction and related processes. It is difficult to explain this inverse relationship between the compaction potentials of Na+ and K+ and their binding abilities. In this study we sought to resolve this contradiction and emphasize the phenomenological distinction between DNA compaction and DNA binding processes in the case of DNA compaction by monocations. Using polyethylene glycol solutions as a model of a crowded cell environment, we studied DNA compaction by alkali metal salts LiCl, NaCl, KCl, RbCl, and CsCl, and found that all of these monocations promote DNA compaction. Among these monovalent cations Na+ produces the greatest compaction and the ratio of K+ cand Na+ oncentrations for DNA compaction is ∼1.5–2. A comparative analysis of recent experimental results indicates that a higher binding activity of monocation generally corresponds to a low compaction potential of the corresponding monovalent ion. This inverse relation is explained as a result of partial dehydration of monocations in the compact state. PMID:15778438

Investigation of the binding of Salvianolic acid B to human serum albumin and the effect of metal ions on the binding

NASA Astrophysics Data System (ADS)

Chen, Tingting; Cao, Hui; Zhu, Shajun; Lu, Yapeng; Shang, Yanfang; Wang, Miao; Tang, Yanfeng; Zhu, Li

2011-10-01

The studies on the interaction between HSA and drugs have been an interesting research field in life science, chemistry and clinical medicine. There are also many metal ions present in blood plasma, thus the research about the effect of metal ions on the interaction between drugs and plasma proteins is crucial. In this study, the interaction of Salvianolic acid B (Sal B) with human serum albumin (HSA) was investigated by the steady-state, synchronous fluorescence and circular dichroism (CD) spectroscopies. The results showed that Sal B had a strong ability to quench the intrinsic fluorescence of HSA through a static quenching mechanism. Binding parameters calculated showed that Sal B was bound to HSA with the binding affinities of 10 5 L mol -1. The thermodynamic parameters studies revealed that the binding was characterized by positive enthalpy and positive entropy changes, and hydrophobic interactions were the predominant intermolecular forces to stabilize the complex. The specific binding distance r (2.93 nm) between donor (HSA) and acceptor (Sal B) was obtained according to Förster non-radiative resonance energy transfer theory. The synchronous fluorescence experiment revealed that Sal B cannot lead to the microenvironmental changes around the Tyr and Trp residues of HSA, and the binding site of Sal B on HSA is located in hydrophobic cavity of subdomain IIA. The CD spectroscopy indicated the secondary structure of HSA is not changed in the presence of Sal B. Furthermore, The effect of metal ions (e.g. Zn 2+, Cu 2+, Co 2+, Ni 2+, Fe 3+) on the binding constant of Sal B-HSA complex was also discussed.

Comparison of Iron-Binding Ability Between Thr70-NapA and Ser70-NapA of Helicobacter pylori.

PubMed

Shan, Weiran; Kung, Hsiang-Fu; Ge, Ruiguang

2016-06-01

The neutrophil-activating protein (NapA) of Helicobacter pylori (H. pylori), with DNA-binding and iron seizing properties, is a fundamental virulence factor involved in H. pylori-related diseases. Compared with Ser70-NapA strain, Thr70-NapA strain is more intimately correlated with iron-deficiency anemia. To investigate whether two types of proteins differ in iron-binding ability, mutated Thr70-NapA and Ser70-NapA strains were established. Isothermal titration calorimetry (ITC) method was conducted to measure the binding between the NapA protein and Fe(2+) . The structural changes of NapA protein were also tested during iron interaction by fast protein liquid chromatography (FPLC) and circular dichroism (CD) methods. DNA-binding assay was performed for evaluate the affinity of both mutated and wild types of NapA with DNA. Mutated Thr70-NapA had higher iron-binding ability than wild Ser70-NapA. The structural stability of Thr70-NapA was disrupted and became more active along with the rising concentration of Fe(2+) , whereas no similar association was observed between Ser70-NapA and Fe(2+) level. When the iron/protein molar ratio ranged from 10 to 20, both Ser70-NapA and Thr70-NapA displayed weaker DNA-binding ability. Thr70-NapA has much stronger ability to sequester ferrous ion compared with Ser70-NapA in H. pylori. In addition, the DNA-binding property of NapA is dependent upon the Fe(2+) concentration. © 2015 John Wiley & Sons Ltd.

Competition Between Co(NH3)63+ and Inner Sphere Mg2+ Ions in the HDV Ribozyme

PubMed Central

Gong, Bo; Chen, Jui-Hui; Bevilacqua, Philip C.; Golden, Barbara L.; Carey, Paul R.

2009-01-01

Divalent cations play critical structural and functional roles in many RNAs. While the hepatitis delta virus (HDV) ribozyme can undergo self-cleavage in the presence of molar concentrations of monovalent cations, divalent cations such as Mg2+ are required for efficient catalysis under physiological conditions. Moreover, the cleavage reaction can be inhibited with Co(NH3)63+, an analog of Mg(H2O)62+. Here, the binding of Mg2+ and Co(NH3)63+ to the HDV ribozyme are studied by Raman microscopic analysis of crystals. Raman difference spectra acquired at different metal ion conditions reveal changes in the ribozyme. When Mg2+ alone is introduced to the ribozyme, inner sphere coordination of Mg(H2O)x2+ (x≤5) to non-bridging PO2− oxygen, and changes in base stretches and phosphodiester group conformation are observed. In addition, binding of Mg2+ induces deprotonation of a cytosine assigned to the general acid C75, consistent with solution studies. When Co(NH3)63+ alone is introduced, deprotonation of C75 is again observed, as are distinctive changes in base vibrational ring modes and phosphodiester backbone conformation. In contrast to Mg2+ binding, Co(NH3)63+ binding does not perturb PO2− group vibrations, consistent with its ability to make only outer sphere contacts. Surprisingly, competitive binding studies reveal that Co(NH3)63+ ions displace some inner sphere-coordinated magnesium species, including ions coordinated to PO2− groups or the N7 of a guanine, likely G1 at the active site. These observations contrast with the tenet that Co(NH3)63+ ions displace only outer sphere magnesium ions. Overall, our data support two classes of inner sphere Mg2+-PO2− binding sites: sites that Co(NH3)63+ can displace, and others it cannot. PMID:19888753

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

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

Molecular models of alginic acid: Interactions with calcium ions and calcite surfaces

NASA Astrophysics Data System (ADS)

Perry, Thomas D.; Cygan, Randall T.; Mitchell, Ralph

2006-07-01

Cation binding by polysaccharides is observed in many environments and is important for predictive environmental modeling, and numerous industrial and food technology applications. The complexities of these cation-organic interactions are well suited for predictive molecular modeling and the analysis of conformation and configuration of polysaccharides and their influence on cation binding. In this study, alginic acid was chosen as a model polymer system and representative disaccharide and polysaccharide subunits were developed. Molecular dynamics simulation of the torsion angles of the ether linkage between various monomeric subunits identified local and global energy minima for selected disaccharides. The simulations indicate stable disaccharide configurations and a common global energy minimum for all disaccharide models at Φ = 274 ± 7°, Ψ = 227 ± 5°, where Φ and Ψ are the torsion angles about the ether linkage. The ability of disaccharide subunits to bind calcium ions and to associate with the (101¯4) surface of calcite was also investigated. Molecular models of disaccharide interactions with calcite provide binding energy differences for conformations that are related to the proximity and residence densities of the electron-donating moieties with calcium ions on the calcite surface, which are controlled, in part, by the torsion of the ether linkage between monosaccharide units. Dynamically optimized configurations for polymer alginate models with calcium ions were also derived.

Interactions of casein micelles with calcium phosphate particles.

PubMed

Tercinier, Lucile; Ye, Aiqian; Anema, Skelte G; Singh, Anne; Singh, Harjinder

2014-06-25

Insoluble calcium phosphate particles, such as hydroxyapatite (HA), are often used in calcium-fortified milks as they are considered to be chemically unreactive. However, this study showed that there was an interaction between the casein micelles in milk and HA particles. The caseins in milk were shown to bind to the HA particles, with the relative proportions of bound β-casein, αS-casein, and κ-casein different from the proportions of the individual caseins present in milk. Transmission electron microscopy showed no evidence of intact casein micelles on the surface of the HA particles, which suggested that the casein micelles dissociated either before or during binding. The HA particles behaved as ion chelators, with the ability to bind the ions contained in the milk serum phase. Consequently, the depletion of the serum minerals disrupted the milk mineral equilibrium, resulting in dissociation of the casein micelles in milk.

Influences of urea, pH and metal ions on the interaction between cepharanthine and lysozyme by steady state fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Yang, Yumin; Li, Daojin; Xu, Chen

2015-03-01

The study on the binding mode of drug with protein is important to understand the pharmacokinetics and toxicity of the drug as well as the relationship of structure and function of the protein. In the study, the interaction between cepharanthine and lysozyme (Lys) in aqueous solution was first investigated by fluorescence spectroscopic techniques at pH 7.4. The obtained quenching rate constant and binding constant indicated the static quenching mechanism and medium binding force. The effect of cepharanthine on the conformation of Lys was analyzed using synchronous fluorescence and three-dimensional (3D) fluorescence. In addition, the effect of urea on the interaction of cepharanthine with Lys was studied and the binding capacity of cepharanthine to the denatured Lys deceases dramatically, as compared with that of cepharanthine to native Lys. Moreover, influence of pH on the interaction of cepharanthine with Lys was investigated. As compared with that at pH 7.4, the binding abilities of the drug to Lys under other pH conditions (pH 9.0, 5.5, 3.5, and 1.9) deceased. Furthermore, the effect of metal ions on the binding constant of cepharanthine with Lys was investigated.

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.

The impact of highly hydrophobic material on the structure of transferrin and its ability to bind iron.

PubMed

Drug, E; Fadeev, L; Gozin, M

2011-05-30

Transferrin is a blood-plasma glycoprotein, which is responsible for ferric-ion delivery and which functions as the most important ferric pool in the body. The reversible complexation process of Fe(3+) ions is associated with conformational changes of the three-dimensional structure of the transferrin. This conformational dynamics is attributed to a partial unfolding of the N-lobe of the protein and could be described as a transition between the holo to the apo forms of the transferrin. The aim of the present work is to demonstrate the unprecedented ability of the transferrin to solubilize various polycyclic aromatic hydrocarbons in physiological solution and to explore the impact of these materials on the structure and functionality of the transferrin. The synthesis and characterization of novel materials, consisting of complexes between human transferrin and hydrophobic high-carbon-content compounds, is reported here for the first time. Furthermore, it is shown that the preparation of these complexes from holo-transferrin leads to an irreversible loss of the ferric ions from the protein. Analytical studies of these novel complexes may shed a light on the mechanism by which transferrin could lose its ability to bind and thus to transport and store iron. These findings clearly demonstrate a possible damaging impact of various hydrophobic pollutants, which can enter an organism by inhalation or ingestion, on the functionality of the transferrin. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

[Modification of retinal photoreceptor membranes and Ca ion binding].

PubMed

Korchagin, V P; Berman, A L; Shukoliukov, S A; Rychkova, M P; Etingof, R N

1978-10-01

Calcium binding by modified photoreceptor membranes of cattle retina has been studied. Ca2+-binding the membranes significantly changes after C-phospholipase treatment, displaying the initial growth (less than 65% of lipid phosphorus removed) with subsequent decrease (more than 65% of phosphorus removed). Liposomes of the photoreceptor membranes lipids were found to bind more calcium than do the native photoreceptor membranes. Proteolytic enzymes (papaine, pronase) splitting some rhodopsin fragments do not affect the ability of the membrane to bind Ca2+. The increase of light-induced Ca-binding is observed only after the outer segments preincubation under conditions providing for rhodopsin phosphorylation. This effect was observed also after the splitting of the rhodopsin fragment by papaine. It is concluded that calcium binding in the photoreceptor membranes is mainly due to the phosphate groups of phospholipids.

Dicationic Surfactants with Glycine Counter Ions for Oligonucleotide Transportation.

PubMed

Pietralik, Zuzanna; Skrzypczak, Andrzej; Kozak, Maciej

2016-08-04

Gemini surfactants are good candidates to bind, protect, and deliver nucleic acids. Herein, the concept of amino acids (namely glycine) as counter ions of gemini surfactants for gene therapy application was explored. This study was conducted on DNA and RNA oligomers and two quaternary bis-imidazolium salts, having 2,5-dioxahexane and 2,8-dioxanonane spacer groups. The toxicity level of surfactants was assessed by an MTT assay, and their ability to bind nucleic acids was tested through electrophoresis. The nucleic acid conformation was established based on circular dichroism and infrared spectroscopic analyses. The structures of the formed complexes were characterized by small-angle scattering of synchrotron radiation. Both studied surfactants appear to be suitable for gene therapy; however, although they vary by only three methylene groups in the spacer, they differ in binding ability and toxicity. The tested oligonucleotides maintained their native conformations upon surfactant addition and the studied lipoplexes formed a variety of structures. In systems based on a 2,5-dioxahexane spacer, a hexagonal phase was observed for DNA-surfactant complexes and a micellar phase was dominant with RNA. For the surfactant with a 2,8-dioxanonane spacer group, the primitive cubic phase prevailed. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effects of increasing number of rings on the ion sensing ability of CdSe quantum dots: a theoretical study

NASA Astrophysics Data System (ADS)

Malik, Pragati; Kakkar, Rita

2018-04-01

A computational study on the structural and electronic properties of a special class of artificial atoms, known as quantum dots, has been carried out. These are semiconductors with unique optical and electronic properties and have been widely used in various applications, such as bio-sensing, bio-imaging, and so on. We have considered quantum dots belonging to II-VI types of semiconductors, due to their wide band gap, possession of large exciton binding energies and unique optical and electronic properties. We have studied their applications as chemical ion sensors by beginning with the study of the ion sensing ability of (CdSe) n ( n = 3, 6, 9 which are in the size range of 0.24, 0.49, 0.74 nm, respectively) quantum dots for cations of the zinc triad, namely Zn2+, Cd2+, Hg2+, and various anions of biological and environmental importance, and studied the effect of increasing number of rings on their ion sensing ability. The various structural, electronic, and optical properties, their interaction energies, and charge transfer on interaction with metal ions and anions have been calculated and reported. Our studies indicate that the CdSe quantum dots can be employed as sensors for both divalent cations and anions, but they can sense cations better than anions.

Dramatically stabilizing multiprotein complex structure in the absence of bulk water using tuned Hofmeister salts.

PubMed

Han, Linjie; Hyung, Suk-Joon; Ruotolo, Brandon T

2013-01-01

The role that water plays in the salt-based stabilization of proteins is central to our understanding of protein biophysics. Ion hydration and the ability of ions to alter water surface tension are typically invoked, along with direct ion-protein binding, to describe Hofmeister stabilization phenomena observed for proteins experimentally, but the relative influence of these forces has been extraordinarily difficult to measure directly. Recently, we have used gas-phase measurements of proteins and large multiprotein complexes, using a combination of innovative ion mobility (IM) and mass spectrometry (MS) techniques, to assess the ability of bound cations and anions to stabilize protein ions in the absence of the solvation forces described above. Our previous work has studied a broad set of 12 anions bound to a range of proteins and protein complexes, and while primarily motivated by the analytical challenges surrounding the gas-phase measurement of solution-phase relevant protein structures, our work has also lead to a detailed physical mechanism of anion-protein complex stabilization in the absence of bulk solvent. Our more-recent work has screened a similarly-broad set of cations for their ability to stabilize gas-phase protein structure, and we have discovered surprising differences between the operative mechanisms for cations and anions in gas-phase protein stabilization. In both cases, cations and anions affect protein stabilization in the absence of solvent in a manner that is generally reversed relative to their ability to stabilize the same proteins in solution. In addition, our evidence suggests that the relative solution-phase binding affinity of the anions and cations studied here is preserved in our gas-phase measurements, allowing us to study the influence of such interactions in detail. In this report, we collect and summarize such gas-phase measurements to distill a generalized picture of salt-based protein stabilization in the absence of bulk water. Further, we communicate our most recent efforts to study the combined effects of stabilizing cations and anions on gas-phase proteins, and identify those salts that bear anion/cation pairs having the strongest stabilizing influence on protein structures

Facile characterization of aptamer kinetic and equilibrium binding properties using surface plasmon resonance

PubMed Central

Chang, Andrew L.; McKeague, Maureen; Smolke, Christina D.

2015-01-01

Nucleic acid aptamers find widespread use as targeting and sensing agents in nature and biotechnology. Their ability to bind an extensive range of molecular targets, including small molecules, proteins, and ions, with high affinity and specificity enables their use in diverse diagnostic, therapeutic, imaging, and gene-regulatory applications. Here, we describe methods for characterizing aptamer kinetic and equilibrium binding properties using a surface plasmon resonance-based platform. This aptamer characterization platform is broadly useful for studying aptamer–ligand interactions, comparing aptamer properties, screening functional aptamers during in vitro selection processes, and prototyping aptamers for integration into nucleic acid devices. PMID:25432760

Structure and dynamics of microbe-exuded polymers and their interactions with calcite surfaces.

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

Cygan, Randall Timothy; Mitchell, Ralph; Perry, Thomas D.

2005-12-01

Cation binding by polysaccharides is observed in many environments and is important for predictive environmental modeling, and numerous industrial and food technology applications. The complexities of these organo-cation interactions are well suited to predictive molecular modeling studies for investigating the roles of conformation and configuration of polysaccharides on cation binding. In this study, alginic acid was chosen as a model polymer and representative disaccharide and polysaccharide subunits were modeled. The ability of disaccharide subunits to bind calcium and to associate with the surface of calcite was investigated. The findings were extended to modeling polymer interactions with calcium ions.

Expressing a bacterial mercuric ion binding protein in plant for phytoremediation of heavy metals.

PubMed

Hsieh, Ju-Liang; Chen, Ching-Yi; Chiu, Meng-Hsuen; Chein, Mei-Fang; Chang, Jo-Shu; Endo, Ginro; Huang, Chieh-Chen

2009-01-30

A specific mercuric ion binding protein (MerP) originating from transposon TnMERI1 of Bacillus megaterium strain MB1 isolated from Minamata Bay displayed good adsorption capability for a variety of heavy metals. In this study, the Gram-positive MerP protein was expressed in transgenic Arabidopsis to create a model system for phytoremediation of heavy metals. Under control of an actin promoter, the transgenic Arabidpsis showed higher tolerance and accumulation capacity for mercury, cadium and lead when compared with the control plant. Results from confocal microscopy analysis also indicate that MerP was localized at the cell membrane and vesicles of plant cells. The developed transgenic plants possessing excellent metal-accumulative ability could have potential applications in decontamination of heavy metals.

Selection of Inhibitor-Resistant Viral Potassium Channels Identifies a Selectivity Filter Site that Affects Barium and Amantadine Block

PubMed Central

Fujiwara, Yuichiro; Arrigoni, Cristina; Domigan, Courtney; Ferrara, Giuseppina; Pantoja, Carlos; Thiel, Gerhard; Moroni, Anna; Minor, Daniel L.

2009-01-01

Background Understanding the interactions between ion channels and blockers remains an important goal that has implications for delineating the basic mechanisms of ion channel function and for the discovery and development of ion channel directed drugs. Methodology/Principal Findings We used genetic selection methods to probe the interaction of two ion channel blockers, barium and amantadine, with the miniature viral potassium channel Kcv. Selection for Kcv mutants that were resistant to either blocker identified a mutant bearing multiple changes that was resistant to both. Implementation of a PCR shuffling and backcrossing procedure uncovered that the blocker resistance could be attributed to a single change, T63S, at a position that is likely to form the binding site for the inner ion in the selectivity filter (site 4). A combination of electrophysiological and biochemical assays revealed a distinct difference in the ability of the mutant channel to interact with the blockers. Studies of the analogous mutation in the mammalian inward rectifier Kir2.1 show that the T→S mutation affects barium block as well as the stability of the conductive state. Comparison of the effects of similar barium resistant mutations in Kcv and Kir2.1 shows that neighboring amino acids in the Kcv selectivity filter affect blocker binding. Conclusions/Significance The data support the idea that permeant ions have an integral role in stabilizing potassium channel structure, suggest that both barium and amantadine act at a similar site, and demonstrate how genetic selections can be used to map blocker binding sites and reveal mechanistic features. PMID:19834614

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.

Gallium as a Therapeutic Agent: A Thermodynamic Evaluation of the Competition between Ga(3+) and Fe(3+) Ions in Metalloproteins.

PubMed

Nikolova, Valia; Angelova, Silvia; Markova, Nikoleta; Dudev, Todor

2016-03-10

Gallium has been employed (in the form of soluble salts) to fight various forms of cancer, infectious, and inflammatory diseases. The rationale behind this lies in the ability of Ga(3+) cation to mimic closely in appearance the native ferric ion, Fe(3+), thus interfering with the biological processes requiring ferric cofactors. However, Ga(3+) ion cannot participate in redox reactions and, when substituting for the "native" Fe(3+) ion in the enzyme active site, renders it inactive. Although a significant body of information on the Ga(3+)-Fe(3+) competition in biological systems has been accumulated, the intimate mechanism of the process is still not well understood and several questions remain: What are the basic physical principles governing the competition between the two trivalent cations in proteins? What type of metal centers are the most likely targets for gallium therapy? To what extent are the Fe(3+)-binding sites in the key enzyme ribonucleotide reductase vulnerable to Ga(3+) substitution? Here, we address these questions by studying the competition between Ga(3+) and Fe(3+) ions in model metal binding sites of various compositions and charge states. The results obtained are in line with available experimental data and shed light on the intimate mechanism of the Ga(3+)/Fe(3+) selectivity in various model metal binding sites and biological systems such as serum transferrin and ribonucleotide reductase.

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.

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

Uncoupling metallonuclease metal ion binding sites via nudge mutagenesis.

PubMed

Papadakos, Grigorios A; Nastri, Horacio; Riggs, Paul; Dupureur, Cynthia M

2007-05-01

The hydrolysis of phosphodiester bonds by nucleases is critical to nucleic acid processing. Many nucleases utilize metal ion cofactors, and for a number of these enzymes two active-site metal ions have been detected. Testing proposed mechanistic roles for individual bound metal ions has been hampered by the similarity between the sites and cooperative behavior. In the homodimeric PvuII restriction endonuclease, the metal ion dependence of DNA binding is sigmoidal and consistent with two classes of coupled metal ion binding sites. We reasoned that a conservative active-site mutation would perturb the ligand field sufficiently to observe the titration of individual metal ion binding sites without significantly disturbing enzyme function. Indeed, mutation of a Tyr residue 5.5 A from both metal ions in the enzyme-substrate crystal structure (Y94F) renders the metal ion dependence of DNA binding biphasic: two classes of metal ion binding sites become distinct in the presence of DNA. The perturbation in metal ion coordination is supported by 1H-15N heteronuclear single quantum coherence spectra of enzyme-Ca(II) and enzyme-Ca(II)-DNA complexes. Metal ion binding by free Y94F is basically unperturbed: through multiple experiments with different metal ions, the data are consistent with two alkaline earth metal ion binding sites per subunit of low millimolar affinity, behavior which is very similar to that of the wild type. The results presented here indicate a role for the hydroxyl group of Tyr94 in the coupling of metal ion binding sites in the presence of DNA. Its removal causes the affinities for the two metal ion binding sites to be resolved in the presence of substrate. Such tuning of metal ion affinities will be invaluable to efforts to ascertain the contributions of individual bound metal ions to metallonuclease function.

Structure-guided design of a high-affinity platelet integrin αIIbβ3 receptor antagonist that disrupts Mg²⁺ binding to the MIDAS.

PubMed

Zhu, Jieqing; Choi, Won-Seok; McCoy, Joshua G; Negri, Ana; Zhu, Jianghai; Naini, Sarasija; Li, Jihong; Shen, Min; Huang, Wenwei; Bougie, Daniel; Rasmussen, Mark; Aster, Richard; Thomas, Craig J; Filizola, Marta; Springer, Timothy A; Coller, Barry S

2012-03-14

An integrin found on platelets, α(IIb)β(3) mediates platelet aggregation, and α(IIb)β(3) antagonists are effective antithrombotic agents in the clinic. Ligands bind to integrins in part by coordinating a magnesium ion (Mg(2+)) located in the β subunit metal ion-dependent adhesion site (MIDAS). Drugs patterned on the integrin ligand sequence Arg-Gly-Asp have a basic moiety that binds the α(IIb) subunit and a carboxyl group that coordinates the MIDAS Mg(2+) in the β(3) subunits. They induce conformational changes in the β(3) subunit that may have negative consequences such as exposing previously hidden epitopes and inducing the active conformation of the receptor. We recently reported an inhibitor of α(IIb)β(3) (RUC-1) that binds exclusively to the α(IIb) subunit; here, we report the structure-based design and synthesis of RUC-2, a RUC-1 derivative with a ~100-fold higher affinity. RUC-2 does not induce major conformational changes in β(3) as judged by monoclonal antibody binding, light scattering, gel chromatography, electron microscopy, and a receptor priming assay. X-ray crystallography of the RUC-2-α(IIb)β(3) headpiece complex in 1 mM calcium ion (Ca(2+))/5 mM Mg(2+) at 2.6 Å revealed that RUC-2 binds to α(IIb) the way RUC-1 does, but in addition, it binds to the β(3) MIDAS residue glutamic acid 220, thus displacing Mg(2+) from the MIDAS. When the Mg(2+) concentration was increased to 20 mM, however, Mg(2+) was identified in the MIDAS and RUC-2 was absent. RUC-2's ability to inhibit ligand binding and platelet aggregation was diminished by increasing the Mg(2+) concentration. Thus, RUC-2 inhibits ligand binding by a mechanism different from that of all other α(IIb)β(3) antagonists and may offer advantages as a therapeutic agent.

Ion Selectivity Mechanism in a Bacterial Pentameric Ligand-Gated Ion Channel

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

Fritsch, Sebastian M; Ivanov, Ivaylo N; Wang, Hailong

2011-01-01

The proton-gated ion channel from Gloeobacter violaceus (GLIC) is a prokaryotic homolog of the eukaryotic nicotinic acetylcholine receptor (nAChR) that responds to the binding of neurotransmitter acetylcholine and mediates fast signal transmission. Recent emergence of a high resolution crystal structure of GLIC captured in a potentially open state allowed detailed, atomic-level insight into ion conduction and selectivity mechanisms in these channels. Herein, we have examined the barriers to ion conduction and origins of ion selectivity in the GLIC channel by the construction of potential of mean force (PMF) profiles for sodium and chloride ions inside the transmembrane region. Our calculationsmore » reveal that the GLIC channel is open for a sodium ion to transport, but presents a ~10 kcal/mol free energy barrier for a chloride ion, which arises primarily from the unfavorable interactions with a ring of negatively charged glutamate residues (E-2 ) at the intracellular end and a ring of hydrophobic residues (I9 ) in the middle of the transmembrane domain. Our collective findings further suggest that the charge selection mechanism can, to a large extent, be attributed to the narrow intracellular end and a ring of glutamate residues in this position their strong negative electrostatics and ability to bind cations. By contrast, E19 at the extracellular entrance only plays a minor role in ion selectivity of GLIC. In addition to electrostatics, both ion hydration and protein dynamics are found to be crucial for ion conduction as well, which explains why a chloride ion experiences a much greater barrier than a sodium ion in the hydrophobic region of the pore.« less

New tris(dopamine) derivative as an iron chelator. Synthesis, solution thermodynamic stability, and antioxidant research.

PubMed

Zhang, Qingchun; Jin, Bo; Shi, Zhaotao; Wang, Xiaofang; Lei, Shan; Tang, Xingyan; Liang, Hua; Liu, Qiangqiang; Gong, Mei; Peng, Rufang

2017-06-01

A new tris(dopamine) derivative, containing three dopamine chelate moieties which were attached to a trimesic acid molecular scaffold, has been prepared and fully characterized by NMR, FTIR and HRMS. The solution thermodynamic stability of the chelator with Fe(III), Mg(II), Zn(II) and Fe(II) ions was investigated. Results demonstrated that the chelator exhibited effective binding ability and improved selectivity to Fe(III) ion. The chelator possessed affinity similar to that of diethylenetriaminepentaacetic acid chelator for Fe(III) ion. The high affinity could be attributed to the favorable geometric arrangement between the chelator and Fe(III) ion coordination preference. The chelator also exhibited high antioxidant activity and nontoxicity to neuron-like rat pheochromocytoma cells. Hence, the chelator could be used as chelating agent for iron overload situations without depleting essential metal ions, such as Mg(II) and Zn(II) ions. Copyright © 2017. Published by Elsevier Inc.

The tautomerization between keto- to phenol-hydrazone induced by anions in the solution

NASA Astrophysics Data System (ADS)

Shang, Xuefang; Yuan, Jianmei; Wang, Yingling; Zhang, Jinlian; Xu, Xiufang

2012-02-01

Two simple anion receptors, 2-[(2-hydroxy-5-nitrophenyl)methylene]hydrazone (1) and 2-[(3,5-dibromo-2-hydroxyphenyl)methylene]hydrazone (2) with -OH binding sites, were synthesized and characterized. The anion binding ability of receptors 1 and 2 with halide anions (F-, Cl-, Br- and I-), AcO- and HPO4- was investigated using visual (naked-eye), UV-vis titration experiments in dry DMSO together with DFT theoretical calculation. The addition of F-, AcO- and HPO4- to the host solution resulted in a red shift of the charge-transfer absorbance band accompanied by a color change from yellow to orange in the naked-eye experiments. Receptor 1 containing a nitro group at the para position and receptor 2 containing two bromine groups at the ortho and para positions both showed strong binding ability for HPO4- ion in the form of phenol-hydrazone. Moreover, receptor 1, induced by anion species in the solution, converted to the form of phenol-hydrazone from keto-hydrazone.

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.

Ion Selectivity Mechanism in a Bacterial Pentameric Ligand-Gated Ion Channel

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

Fritsch, Sebastian; Ivanov, Ivaylo; Wang, Hailong

2010-01-01

The proton-gated ion channel from Gloeobacter violaceus (GLIC) is a prokaryotic homolog of the eukaryotic nicotinic acetylcholine receptor that responds to the binding of neurotransmitter acetylcholine and mediates fast signal transmission. Recent emergence of a high-resolution crystal structure of GLIC captured in a potentially open state allowed detailed, atomic-level insight into ion conduction and selectivity mechanisms in these channels. Herein, we have examined the barriers to ion conduction and origins of ion selectivity in the GLIC channel by the construction of potential-of-mean-force profiles for sodium and chloride ions inside the transmembrane region. Our calculations reveal that the GLIC channel ismore » open for a sodium ion to transport, but presents a 11 kcal/mol free energy barrier for a chloride ion. Our collective findings identify three distinct contributions to the observed preference for the permeant ions. First, there is a substantial contribution due to a ring of negatively charged glutamate residues (E-2 ) at the narrow intracellular end of the channel. The negative electrostatics of this region and the ability of the glutamate side chains to directly bind cations would strongly favor the passage of sodium ions while hindering translocation of chloride ions. Second, our results imply a significant hydrophobic contribution to selectivity linked to differences in the desolvation penalty for the sodium versus chloride ions in the central hydrophobic region of the pore. This hydrophobic contribution is evidenced by the large free energy barriers experienced by Cl in the middle of the pore for both GLIC and the E-2 A mutant. Finally, there is a distinct contribution arising from the overall negative electrostatics of the channel.« less

Multi-shell model of ion-induced nucleic acid condensation

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

Tolokh, Igor S.; Drozdetski, Aleksander V.; Pollack, Lois

2016-04-21

We present a semi-quantitative model of condensation of short nucleic acid (NA) duplexes in- duced by tri-valent cobalt hexammine (CoHex) ions. The model is based on partitioning of bound counterion distribution around single NA duplex into “external” and “internal” ion binding shells distinguished by the proximity to duplex helical axis. The duplex aggregation free energy is de- composed into attraction and repulsion components represented by simple analytic expressions. The source of the short-range attraction between NA duplexes in the aggregated phase is the in- teraction of CoHex ions in the overlapping regions of the “external” shells with the oppositely chargedmore » duplexes. The attraction depends on CoHex binding affinity to the “external” shell of nearly neutralized duplex and the number of ions in the shell overlapping volume. For a given NA duplex sequence and structure, these parameters are estimated from molecular dynamics simula- tion. The attraction is opposed by the residual repulsion of nearly neutralized duplexes as well as duplex configurational entropy loss upon aggregation. The estimates of the aggregation free energy are consistent with the experimental range of NA duplex condensation propensities, including the unusually poor condensation of RNA structures and subtle sequence effects upon DNA conden- sation. The model predicts that, in contrast to DNA, RNA duplexes may condense into tighter packed aggregates with a higher degree of duplex neutralization. The model also predicts that longer NA fragments will condense easier than shorter ones. The ability of this model to explain experimentally observed trends in NA condensation, lends support to proposed NA condensation picture based on the multivalent “ion binding shells”.« less

Representation of Ion–Protein Interactions Using the Drude Polarizable Force-Field

PubMed Central

2016-01-01

Small metal ions play critical roles in numerous biological processes. Of particular interest is how metalloenzymes are allosterically regulated by the binding of specific ions. Understanding how ion binding affects these biological processes requires atomic models that accurately treat the microscopic interactions with the protein ligands. Theoretical approaches at different levels of sophistication can contribute to a deeper understanding of these systems, although computational models must strike a balance between accuracy and efficiency in order to enable long molecular dynamics simulations. In this study, we present a systematic effort to optimize the parameters of a polarizable force field based on classical Drude oscillators to accurately represent the interactions between ions (K+, Na+, Ca2+, and Cl–) and coordinating amino-acid residues for a set of 30 biologically important proteins. By combining ab initio calculations and experimental thermodynamic data, we derive a polarizable force field that is consistent with a wide range of properties, including the geometries and interaction energies of gas-phase ion/protein-like model compound clusters, and the experimental solvation free-energies of the cations in liquids. The resulting models display significant improvements relative to the fixed-atomic-charge additive CHARMM C36 force field, particularly in their ability to reproduce the many-body electrostatic nonadditivity effects estimated from ab initio calculations. The analysis clarifies the fundamental limitations of the pairwise additivity assumption inherent in classical fixed-charge force fields, and shows its dramatic failures in the case of Ca2+ binding sites. These optimized polarizable models, amenable to computationally efficient large-scale MD simulations, set a firm foundation and offer a powerful avenue to study the roles of the ions in soluble and membrane transport proteins. PMID:25578354

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

Agonist trapped in ATP-binding sites of the P2X2 receptor.

PubMed

Jiang, Ruotian; Lemoine, Damien; Martz, Adeline; Taly, Antoine; Gonin, Sophie; Prado de Carvalho, Lia; Specht, Alexandre; Grutter, Thomas

2011-05-31

ATP-gated P2X receptors are trimeric ion channels, as recently confirmed by X-ray crystallography. However, the structure was solved without ATP and even though extracellular intersubunit cavities surrounded by conserved amino acid residues previously shown to be important for ATP function were proposed to house ATP, the localization of the ATP sites remains elusive. Here we localize the ATP-binding sites by creating, through a proximity-dependent "tethering" reaction, covalent bonds between a synthesized ATP-derived thiol-reactive P2X2 agonist (NCS-ATP) and single cysteine mutants engineered in the putative binding cavities of the P2X2 receptor. By combining whole-cell and single-channel recordings, we report that NCS-ATP covalently and specifically labels two previously unidentified positions N140 and L186 from two adjacent subunits separated by about 18 Å in a P2X2 closed state homology model, suggesting the existence of at least two binding modes. Tethering reaction at both positions primes subsequent agonist binding, yet with distinct functional consequences. Labeling of one position impedes subsequent ATP function, which results in inefficient gating, whereas tethering of the other position, although failing to produce gating by itself, enhances subsequent ATP function. Our results thus define a large and dynamic intersubunit ATP-binding pocket and suggest that receptors trapped in covalently agonist-bound states differ in their ability to gate the ion channel.

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.

Electronically conductive polymer binder for lithium-ion battery electrode

DOEpatents

Liu, Gao; Xun, Shidi; Battaglia, Vincent S.; Zheng, Honghe

2017-05-16

A family of carboxylic acid group containing fluorene/fluorenon copolymers is disclosed as binders of silicon particles in the fabrication of negative electrodes for use with lithium ion batteries. These binders enable the use of silicon as an electrode material as they significantly improve the cycle-ability of silicon by preventing electrode degradation over time. In particular, these polymers, which become conductive on first charge, bind to the silicon particles of the electrode, are flexible so as to better accommodate the expansion and contraction of the electrode during charge/discharge, and being conductive promote the flow battery current.

Electronically conductive polymer binder for lithium-ion battery electrode

DOEpatents

Liu, Gao; Xun, Shidi; Battaglia, Vincent S; Zheng, Honghe

2014-10-07

A family of carboxylic acid group containing fluorene/fluorenon copolymers is disclosed as binders of silicon particles in the fabrication of negative electrodes for use with lithium ion batteries. These binders enable the use of silicon as an electrode material as they significantly improve the cycle-ability of silicon by preventing electrode degradation over time. In particular, these polymers, which become conductive on first charge, bind to the silicon particles of the electrode, are flexible so as to better accommodate the expansion and contraction of the electrode during charge/discharge, and being conductive promote the flow battery current.

Microbial biofilms for the removal of Cu²⁺ from CMP wastewater.

PubMed

Mosier, Aaron P; Behnke, Jason; Jin, Eileen T; Cady, Nathaniel C

2015-09-01

The modern semiconductor industry relies heavily on a process known as chemical mechanical planarization, which uses physical and chemical processes to remove excess material from the surface of silicon wafers during microchip fabrication. This process results in large volumes of wastewater containing dissolved metals including copper (Cu(2+)), which must then be filtered and treated before release into municipal waste systems. We have investigated the potential use of bacterial and fungal biomass as an alternative to the currently used ion-exchange resins for the adsorption of dissolved Cu(2+) from high-throughput industrial waste streams. A library of candidate microorganisms, including Lactobacillus casei and Pichia pastoris, was screened for ability to bind Cu(2+) from solution and to form static biofilm communities within packed-bed adsorption columns. The binding efficiency of these biomass-based adsorption columns was assessed under various flow conditions and compared to that of industrially used ion-exchange resins. We demonstrated the potential to regenerate the biomass within the adsorption columns through the use of a hydrochloric acid wash, and subsequently reuse the columns for additional copper binding. While the binding efficiency and capacity of the developed L. casei/P. pastoris biomass filters was inferior to ion-exchange resin, the potential for repeated reuse of these filters, coupled with the advantages of a more sustainable "green" adsorption process, make this technique an attractive candidate for use in industrial-scale CMP wastewater treatment. Copyright © 2015 Elsevier Ltd. All rights reserved.

Total external reflection X-ray fluorescence analysis of protein-metal ion interactions in biological systems

NASA Astrophysics Data System (ADS)

Novikova, N. N.; Kovalchuk, M. V.; Yur'eva, E. A.; Konovalov, O. V.; Rogachev, A. V.; Stepina, N. D.; Sukhorukov, V. S.; Tsaregorodtsev, A. D.; Chukhrai, E. S.; Yakunin, S. N.

2012-09-01

This paper presents the results of an investigation into hemoglobin-based protein films that were formed on a liquid surface. X-ray standing wave measurements were performed at the ID 10 beamline of the European Synchrotron Radiation Facility (ESRF) and at the Langmuir station of the Kurchatov Synchrotron Radiation Source. It was found that the ability of the protein to bind metal ions is substantially increased due to the conformational rearrangements of protein macromolecules caused by various damaging effects. The elemental composition of protein preparations, which were isolated from children and adults with chronic metabolic diseases accompanied by endogenous intoxication, was analyzed. The results of the investigations offer evidence that an increase in the ligand-binding properties of the protein molecules, which was observed in model experiments using protein films, is a common trait and corresponds to in vivo processes accompanying metabolic disturbances in the body.

Free Energy Simulations of Ligand Binding to the Aspartate Transporter GltPh

PubMed Central

Heinzelmann, Germano; Baştuğ, Turgut; Kuyucak, Serdar

2011-01-01

Glutamate/Aspartate transporters cotransport three Na+ and one H+ ions with the substrate and countertransport one K+ ion. The binding sites for the substrate and two Na+ ions have been observed in the crystal structure of the archeal homolog GltPh, while the binding site for the third Na+ ion has been proposed from computational studies and confirmed by experiments. Here we perform detailed free energy simulations of GltPh, giving a comprehensive characterization of the substrate and ion binding sites, and calculating their binding free energies in various configurations. Our results show unequivocally that the substrate binds after the binding of two Na+ ions. They also shed light into Asp/Glu selectivity of GltPh, which is not observed in eukaryotic glutamate transporters. PMID:22098736

Characterization of Lactobacillus brevis L62 strain, highly tolerant to copper ions.

PubMed

Mrvčić, Jasna; Butorac, Ana; Solić, Ema; Stanzer, Damir; Bačun-Družina, Višnja; Cindrić, Mario; Stehlik-Tomas, Vesna

2013-01-01

Lactic acid bacteria (LAB) as starter culture in food industry must be suitable for large-scale industrial production and possess the ability to survive in unfavorable processes and storage conditions. Approaches taken to address these problems include the selection of stress-resistant strains. In food industry, LAB are often exposed to metal ions induced stress. The interactions between LAB and metal ions are very poorly investigated. Because of that, the influence of non-toxic, toxic and antioxidant metal ions (Zn, Cu, and Mn) on growth, acid production, metal ions binding capacity of wild and adapted species of Leuconostoc mesenteroides L3, Lactobacillus brevis L62 and Lactobacillus plantarum L73 were investigated. The proteomic approach was applied to clarify how the LAB cells, especially the adapted ones, protect themselves and tolerate high concentrations of toxic metal ions. Results have shown that Zn and Mn addition into MRS medium in the investigated concentrations did not have effect on the bacterial growth and acid production, while copper ions were highly toxic, especially in static conditions. Leuc. mesenteroides L3 was the most efficient in Zn binding processes among the chosen LAB species, while L. plantarum L73 accumulated the highest concentration of Mn. L. brevis L62 was the most copper resistant species. Adaptation had a positive effect on growth and acid production of all species in the presence of copper. However, the adapted species incorporated less metal ions than the wild species. The exception was adapted L. brevis L62 that accumulated high concentration of copper ions in static conditions. The obtained results showed that L. brevis L62 is highly tolerant to copper ions, which allows its use as starter culture in fermentative processes in media with high concentration of copper ions.

Allosteric nature of P2X receptor activation probed by photoaffinity labelling

PubMed Central

Bhargava, Y; Rettinger, J; Mourot, A

2012-01-01

BACKGROUND AND PURPOSE In P2X receptors, agonist binding at the interface between neighbouring subunits is efficiently transduced to ion channel gating. However, the relationship between binding and gating is difficult to study because agonists continuously bind and unbind. Here, we covalently incorporated agonists in the binding pocket of P2X receptors and examined how binding site occupancy affects the ability of the channel to gate. EXPERIMENTAL APPROACH We used a strategy for tethering agonists to their ATP-binding pocket, while simultaneously probing ion channel gating using electrophysiology. The agonist 2′,3′-O-(4-benzoylbenzoyl)-ATP (BzATP), a photoaffinity analogue of ATP, enabled us to trap rat homomeric P2X2 receptor and a P2X2/1 receptor chimera in different agonist-bound states. UV light was used to control the degree of covalent occupancy of the receptors. KEY RESULTS Irradiation of the P2X2/1 receptor chimera – BzATP complex resulted in a persistent current that lasted even after extensive washout, consistent with photochemical tethering of the agonist BzATP and trapping of the receptors in an open state. Partial labelling with BzATP primed subsequent agonist binding and modulated gating efficiency for both full and partial agonists. CONCLUSIONS AND IMPLICATIONS Our photolabelling strategy provides new molecular insights into the activation mechanism of the P2X receptor. We show here that priming with full agonist molecules leads to an increase in gating efficiency after subsequent agonist binding. PMID:22725669

Route, mechanism, and implications of proton import during Na+/K+ exchange by native Na+/K+-ATPase pumps

PubMed Central

Vedovato, Natascia

2014-01-01

A single Na+/K+-ATPase pumps three Na+ outwards and two K+ inwards by alternately exposing ion-binding sites to opposite sides of the membrane in a conformational sequence coupled to pump autophosphorylation from ATP and auto-dephosphorylation. The larger flow of Na+ than K+ generates outward current across the cell membrane. Less well understood is the ability of Na+/K+ pumps to generate an inward current of protons. Originally noted in pumps deprived of external K+ and Na+ ions, as inward current at negative membrane potentials that becomes amplified when external pH is lowered, this proton current is generally viewed as an artifact of those unnatural conditions. We demonstrate here that this inward current also flows at physiological K+ and Na+ concentrations. We show that protons exploit ready reversibility of conformational changes associated with extracellular Na+ release from phosphorylated Na+/K+ pumps. Reversal of a subset of these transitions allows an extracellular proton to bind an acidic side chain and to be subsequently released to the cytoplasm. This back-step of phosphorylated Na+/K+ pumps that enables proton import is not required for completion of the 3 Na+/2 K+ transport cycle. However, the back-step occurs readily during Na+/K+ transport when external K+ ion binding and occlusion are delayed, and it occurs more frequently when lowered extracellular pH raises the probability of protonation of the externally accessible carboxylate side chain. The proton route passes through the Na+-selective binding site III and is distinct from the principal pathway traversed by the majority of transported Na+ and K+ ions that passes through binding site II. The inferred occurrence of Na+/K+ exchange and H+ import during the same conformational cycle of a single molecule identifies the Na+/K+ pump as a hybrid transporter. Whether Na+/K+ pump–mediated proton inflow may have any physiological or pathophysiological significance remains to be clarified. PMID:24688018

Chelating Tendencies of Bioactive Aminophosphonates

PubMed Central

Lázár, István; Kafarski, Pawel

1994-01-01

The metal-binding abilities of a wide variety of bioactive aminophosphonates, from the simple aminoethanephosphonic acids to the rather large macrocyclic polyaza derivatives, are discussed with special emphasis on a comparison of the analogous carboxylic acid and phosphonic acid systems. Examples are given of the biological importance of metal ion – aminophosphonate interactions in living systems, and also of their actual and potential applicability in medicine. PMID:18476237

Predicting Nonspecific Ion Binding Using DelPhi

PubMed Central

Petukh, Marharyta; Zhenirovskyy, Maxim; Li, Chuan; Li, Lin; Wang, Lin; Alexov, Emil

2012-01-01

Ions are an important component of the cell and affect the corresponding biological macromolecules either via direct binding or as a screening ion cloud. Although some ion binding is highly specific and frequently associated with the function of the macromolecule, other ions bind to the protein surface nonspecifically, presumably because the electrostatic attraction is strong enough to immobilize them. Here, we test such a scenario and demonstrate that experimentally identified surface-bound ions are located at a potential that facilitates binding, which indicates that the major driving force is the electrostatics. Without taking into consideration geometrical factors and structural fluctuations, we show that ions tend to be bound onto the protein surface at positions with strong potential but with polarity opposite to that of the ion. This observation is used to develop a method that uses a DelPhi-calculated potential map in conjunction with an in-house-developed clustering algorithm to predict nonspecific ion-binding sites. Although this approach distinguishes only the polarity of the ions, and not their chemical nature, it can predict nonspecific binding of positively or negatively charged ions with acceptable accuracy. One can use the predictions in the Poisson-Boltzmann approach by placing explicit ions in the predicted positions, which in turn will reduce the magnitude of the local potential and extend the limits of the Poisson-Boltzmann equation. In addition, one can use this approach to place the desired number of ions before conducting molecular-dynamics simulations to neutralize the net charge of the protein, because it was shown to perform better than standard screened Coulomb canned routines, or to predict ion-binding sites in proteins. This latter is especially true for proteins that are involved in ion transport, because such ions are loosely bound and very difficult to detect experimentally. PMID:22735539

Conformational heterogeneity of the calmodulin binding interface

NASA Astrophysics Data System (ADS)

Shukla, Diwakar; Peck, Ariana; Pande, Vijay S.

2016-04-01

Calmodulin (CaM) is a ubiquitous Ca2+ sensor and a crucial signalling hub in many pathways aberrantly activated in disease. However, the mechanistic basis of its ability to bind diverse signalling molecules including G-protein-coupled receptors, ion channels and kinases remains poorly understood. Here we harness the high resolution of molecular dynamics simulations and the analytical power of Markov state models to dissect the molecular underpinnings of CaM binding diversity. Our computational model indicates that in the absence of Ca2+, sub-states in the folded ensemble of CaM's C-terminal domain present chemically and sterically distinct topologies that may facilitate conformational selection. Furthermore, we find that local unfolding is off-pathway for the exchange process relevant for peptide binding, in contrast to prior hypotheses that unfolding might account for binding diversity. Finally, our model predicts a novel binding interface that is well-populated in the Ca2+-bound regime and, thus, a candidate for pharmacological intervention.

Computational Design of Metal Ion Sequestering Agents

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

Hay, Benjamin P.; Rapko, Brian M.

Organic ligands that exhibit a high degree of metal ion recognition are essential precursors for developing separation processes and sensors for metal ions. Since the beginning of the nuclear era, much research has focused on discovering ligands that target specific radionuclides. Members of the Group 1A and 2A cations (e.g., Cs, Sr, Ra) and the f-block metals (actinides and lanthanides) are of primary concern to DOE. Although there has been some success in identifying ligand architectures that exhibit a degree of metal ion recognition, the ability to control binding affinity and selectivity remains a significant challenge. The traditional approach formore » discovering such ligands has involved lengthy programs of organic synthesis and testing that, in the absence of reliable methods for screening compounds before synthesis, have resulted in much wasted research effort.« less

A Unitary Anesthetic Binding Site at High Resolution

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

Vedula, L. Sangeetha; Brannigan, Grace; Economou, Nicoleta J.

2009-10-21

Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABA{sub A} receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show thatmore » apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.« less

A Unitary Anesthetic Binding Site at High Resolution

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

L Vedula; G Brannigan; N Economou

2011-12-31

Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABA{sub A} receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show thatmore » apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.« less

A Unitary Anesthetic-Binding Site at High Resolution

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

Vedula, L.; Brannigan, G; Economou, N

2009-01-01

Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABAA receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show that apoferritinmore » also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.« 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.

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

Agonist trapped in ATP-binding sites of the P2X2 receptor

PubMed Central

Jiang, Ruotian; Lemoine, Damien; Martz, Adeline; Taly, Antoine; Gonin, Sophie; Prado de Carvalho, Lia; Specht, Alexandre; Grutter, Thomas

2011-01-01

ATP-gated P2X receptors are trimeric ion channels, as recently confirmed by X-ray crystallography. However, the structure was solved without ATP and even though extracellular intersubunit cavities surrounded by conserved amino acid residues previously shown to be important for ATP function were proposed to house ATP, the localization of the ATP sites remains elusive. Here we localize the ATP-binding sites by creating, through a proximity-dependent “tethering” reaction, covalent bonds between a synthesized ATP-derived thiol-reactive P2X2 agonist (NCS-ATP) and single cysteine mutants engineered in the putative binding cavities of the P2X2 receptor. By combining whole-cell and single-channel recordings, we report that NCS-ATP covalently and specifically labels two previously unidentified positions N140 and L186 from two adjacent subunits separated by about 18 Å in a P2X2 closed state homology model, suggesting the existence of at least two binding modes. Tethering reaction at both positions primes subsequent agonist binding, yet with distinct functional consequences. Labeling of one position impedes subsequent ATP function, which results in inefficient gating, whereas tethering of the other position, although failing to produce gating by itself, enhances subsequent ATP function. Our results thus define a large and dynamic intersubunit ATP-binding pocket and suggest that receptors trapped in covalently agonist-bound states differ in their ability to gate the ion channel. PMID:21576497

Ion Binding Energies Determining Functional Transport of ClC Proteins

NASA Astrophysics Data System (ADS)

Yu, Tao; Guo, Xu; Zou, Xian-Wu; Sang, Jian-Ping

2014-06-01

The ClC-type proteins, a large family of chloride transport proteins ubiquitously expressed in biological organisms, have been extensively studied for decades. Biological function of ClC proteins can be reflected by analyzing the binding situation of Cl- ions. We investigate ion binding properties of ClC-ec1 protein with the atomic molecular dynamics simulation approach. The calculated electrostatic binding energy results indicate that Cl- at the central binding site Scen has more binding stability than the internal binding site Sint. Quantitative comparison between the latest experimental heat release data isothermal titration calorimetry (ITC) and our calculated results demonstrates that chloride ions prefer to bind at Scen than Sint in the wild-type ClC-ec1 structure and prefer to bind at Sext and Scen than Sint in mutant E148A/E148Q structures. Even though the chloride ions make less contribution to heat release when binding to Sint and are relatively unstable in the Cl- pathway, they are still part contributors for the Cl- functional transport. This work provides a guide rule to estimate the importance of Cl- at the binding sites and how chloride ions have influences on the function of ClC proteins.

Beyond the Hofmeister Series: Ion-Specific Effects on Proteins and Their Biological Functions.

PubMed

Okur, Halil I; Hladílková, Jana; Rembert, Kelvin B; Cho, Younhee; Heyda, Jan; Dzubiella, Joachim; Cremer, Paul S; Jungwirth, Pavel

2017-03-09

Ions differ in their ability to salt out proteins from solution as expressed in the lyotropic or Hofmeister series of cations and anions. Since its first formulation in 1888, this series has been invoked in a plethora of effects, going beyond the original salting out/salting in idea to include enzyme activities and the crystallization of proteins, as well as to processes not involving proteins like ion exchange, the surface tension of electrolytes, or bubble coalescence. Although it has been clear that the Hofmeister series is intimately connected to ion hydration in homogeneous and heterogeneous environments and to ion pairing, its molecular origin has not been fully understood. This situation could have been summarized as follows: Many chemists used the Hofmeister series as a mantra to put a label on ion-specific behavior in various environments, rather than to reach a molecular level understanding and, consequently, an ability to predict a particular effect of a given salt ion on proteins in solutions. In this Feature Article we show that the cationic and anionic Hofmeister series can now be rationalized primarily in terms of specific interactions of salt ions with the backbone and charged side chain groups at the protein surface in solution. At the same time, we demonstrate the limitations of separating Hofmeister effects into independent cationic and anionic contributions due to the electroneutrality condition, as well as specific ion pairing, leading to interactions of ions of opposite polarity. Finally, we outline the route beyond Hofmeister chemistry in the direction of understanding specific roles of ions in various biological functionalities, where generic Hofmeister-type interactions can be complemented or even overruled by particular steric arrangements in various ion binding sites.

Cation binding to 15-TBA quadruplex DNA is a multiple-pathway cation-dependent process.

PubMed

Reshetnikov, Roman V; Sponer, Jiri; Rassokhina, Olga I; Kopylov, Alexei M; Tsvetkov, Philipp O; Makarov, Alexander A; Golovin, Andrey V

2011-12-01

A combination of explicit solvent molecular dynamics simulation (30 simulations reaching 4 µs in total), hybrid quantum mechanics/molecular mechanics approach and isothermal titration calorimetry was used to investigate the atomistic picture of ion binding to 15-mer thrombin-binding quadruplex DNA (G-DNA) aptamer. Binding of ions to G-DNA is complex multiple pathway process, which is strongly affected by the type of the cation. The individual ion-binding events are substantially modulated by the connecting loops of the aptamer, which play several roles. They stabilize the molecule during time periods when the bound ions are not present, they modulate the route of the ion into the stem and they also stabilize the internal ions by closing the gates through which the ions enter the quadruplex. Using our extensive simulations, we for the first time observed full spontaneous exchange of internal cation between quadruplex molecule and bulk solvent at atomistic resolution. The simulation suggests that expulsion of the internally bound ion is correlated with initial binding of the incoming ion. The incoming ion then readily replaces the bound ion while minimizing any destabilization of the solute molecule during the exchange. © The Author(s) 2011. Published by Oxford University Press.

Initial Binding of Ions to the Interhelical Loops of Divalent Ion Transporter CorA: Replica Exchange Molecular Dynamics Simulation Study

PubMed Central

Zhang, Tong; Mu, Yuguang

2012-01-01

Crystal structures of Thermotoga maritima magnesium transporter CorA, reported in 2006, revealed its homo-pentameric constructions. However, the structure of the highly conserved extracellular interhelical loops remains unsolved, due to its high flexibility. We have explored the configurations of the loops through extensive replica exchange molecular dynamics simulations in explicit solvent model with the presence of either Co(III) Hexamine ions or Mg2+ ions. We found that there are multiple binding sites available on the interhelical loops in which the negatively charged residues, E316 and E320, are located notably close to the positively charged ions during the simulations. Our simulations resolved the distinct binding patterns of the two kinds of ions: Co(III) Hexamine ions were found to bind stronger with the loop than Mg2+ ions with binding free energy −7.3 kJ/mol lower, which is nicely consistent with the previous data. Our study provides an atomic basis description of the initial binding process of Mg2+ ions on the extracellular interhelical loops of CorA and the detailed inhibition mechanism of Co(III) Hexamine ions on CorA ions transportation. PMID:22952795

Cation binding to 15-TBA quadruplex DNA is a multiple-pathway cation-dependent process

PubMed Central

Reshetnikov, Roman V.; Sponer, Jiri; Rassokhina, Olga I.; Kopylov, Alexei M.; Tsvetkov, Philipp O.; Makarov, Alexander A.; Golovin, Andrey V.

2011-01-01

A combination of explicit solvent molecular dynamics simulation (30 simulations reaching 4 µs in total), hybrid quantum mechanics/molecular mechanics approach and isothermal titration calorimetry was used to investigate the atomistic picture of ion binding to 15-mer thrombin-binding quadruplex DNA (G-DNA) aptamer. Binding of ions to G-DNA is complex multiple pathway process, which is strongly affected by the type of the cation. The individual ion-binding events are substantially modulated by the connecting loops of the aptamer, which play several roles. They stabilize the molecule during time periods when the bound ions are not present, they modulate the route of the ion into the stem and they also stabilize the internal ions by closing the gates through which the ions enter the quadruplex. Using our extensive simulations, we for the first time observed full spontaneous exchange of internal cation between quadruplex molecule and bulk solvent at atomistic resolution. The simulation suggests that expulsion of the internally bound ion is correlated with initial binding of the incoming ion. The incoming ion then readily replaces the bound ion while minimizing any destabilization of the solute molecule during the exchange. PMID:21893589

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.

Fulvic acid-sulfide ion competition for mercury ion binding in the Florida everglades

USGS Publications Warehouse

Reddy, M.M.; Aiken, G.R.

2001-01-01

Negatively charged functional groups of fulvic acid compete with inorganic sulfide ion for mercury ion binding. This competition is evaluated here by using a discrete site-electrostatic model to calculate mercury solution speciation in the presence of fulvic acid. Model calculated species distributions are used to estimate a mercury-fulvic acid apparent binding constant to quantify fulvic acid and sulfide ion competition for dissolved inorganic mercury (Hg(II)) ion binding. Speciation calculations done with PHREEQC, modified to use the estimated mercury-fulvic acid apparent binding constant, suggest that mercury-fulvic acid and mercury-sulfide complex concentrations are equivalent for very low sulfide ion concentrations (about 10-11 M) in Everglades' surface water. Where measurable total sulfide concentration (about 10-7 M or greater) is present in Everglades' surface water, mercury-sulfide complexes should dominate dissolved inorganic mercury solution speciation. In the absence of sulfide ion (for example, in oxygenated Everglades' surface water), fulvic acid binding should dominate Everglades' dissolved inorganic mercury speciation.

Supported Lipid Bilayers with Phosphatidylethanolamine as the Major Component.

PubMed

Sendecki, Anne M; Poyton, Matthew F; Baxter, Alexis J; Yang, Tinglu; Cremer, Paul S

2017-11-21

Phosphatidylethanolamine (PE) is notoriously difficult to incorporate into model membrane systems, such as fluid supported lipid bilayers (SLBs), at high concentrations because of its intrinsic negative curvature. Using fluorescence-based techniques, we demonstrate that having fewer sites of unsaturation in the lipid tails leads to high-quality SLBs because these lipids help to minimize the curvature. Moreover, shorter saturated chains can help maintain the membranes in the fluid phase. Using these two guidelines, we find that up to 70 mol % PE can be incorporated into SLBs at room temperature and up to 90 mol % PE can be incorporated at 37 °C. Curiously, conditions under which three-dimensional tubules project outward from the planar surface as well as conditions under which domain formation occurs can be found. We have employed these model membrane systems to explore the ability of Ni 2+ to bind to PE. It was found that this transition metal ion binds 1000-fold tighter to PE than to phosphatidylcholine lipids. In the future, this platform could be exploited to monitor the binding of other transition metal ions or the binding of antimicrobial peptides. It could also be employed to explore the physical properties of PE-containing membranes, such as phase domain behavior and intermolecular hydrogen bonding.

Mechanism of Metal Ion Activation of the Diphtheria Toxin Repressor DtxR

NASA Astrophysics Data System (ADS)

D'Aquino, J. Alejandro; Ringe, Dagmar

2006-08-01

The diphtheria toxin repressor, DtxR, is a metal ion-activated transcriptional regulator that has been linked to the virulence of Corynebacterium diphtheriae. Structure determination has shown that there are two metal ion binding sites per repressor monomer, and site-directed mutagenesis has demonstrated that binding site 2 (primary) is essential for recognition of the target DNA repressor, leaving the role of binding site 1 (ancillary) unclear (1 - 3). Calorimetric techniques have demonstrated that while binding site 1 (ancillary) has high affinity for metal ion with a binding constant of 2 × 10-7, binding site 2 (primary) is a low affinity binding site with a binding constant of 6.3 × 10-4. These two binding sites act independently and their contribution can be easily dissected by traditional mutational analysis. Our results clearly demonstrate that binding site 1 (ancillary) is the first one to be occupied during metal ion activation, playing a critical role in stabilization of the repressor. In addition, structural data obtained for the mutants Ni-DtxR(H79A,C102D), reported here and the previously reported DtxR(H79A) (4) has allowed us to propose a mechanism of metal ion activation for DtxR.

Adenosine Monophosphate Binding Stabilizes the KTN Domain of the Shewanella denitrificans Kef Potassium Efflux System.

PubMed

Pliotas, Christos; Grayer, Samuel C; Ekkerman, Silvia; Chan, Anthony K N; Healy, Jess; Marius, Phedra; Bartlett, Wendy; Khan, Amjad; Cortopassi, Wilian A; Chandler, Shane A; Rasmussen, Tim; Benesch, Justin L P; Paton, Robert S; Claridge, Timothy D W; Miller, Samantha; Booth, Ian R; Naismith, James H; Conway, Stuart J

2017-08-15

Ligand binding is one of the most fundamental properties of proteins. Ligand functions fall into three basic types: substrates, regulatory molecules, and cofactors essential to protein stability, reactivity, or enzyme-substrate complex formation. The regulation of potassium ion movement in bacteria is predominantly under the control of regulatory ligands that gate the relevant channels and transporters, which possess subunits or domains that contain Rossmann folds (RFs). Here we demonstrate that adenosine monophosphate (AMP) is bound to both RFs of the dimeric bacterial Kef potassium efflux system (Kef), where it plays a structural role. We conclude that AMP binds with high affinity, ensuring that the site is fully occupied at all times in the cell. Loss of the ability to bind AMP, we demonstrate, causes protein, and likely dimer, instability and consequent loss of function. Kef system function is regulated via the reversible binding of comparatively low-affinity glutathione-based ligands at the interface between the dimer subunits. We propose this interfacial binding site is itself stabilized, at least in part, by AMP binding.

Adenosine Monophosphate Binding Stabilizes the KTN Domain of the Shewanella denitrificans Kef Potassium Efflux System

PubMed Central

2017-01-01

Ligand binding is one of the most fundamental properties of proteins. Ligand functions fall into three basic types: substrates, regulatory molecules, and cofactors essential to protein stability, reactivity, or enzyme–substrate complex formation. The regulation of potassium ion movement in bacteria is predominantly under the control of regulatory ligands that gate the relevant channels and transporters, which possess subunits or domains that contain Rossmann folds (RFs). Here we demonstrate that adenosine monophosphate (AMP) is bound to both RFs of the dimeric bacterial Kef potassium efflux system (Kef), where it plays a structural role. We conclude that AMP binds with high affinity, ensuring that the site is fully occupied at all times in the cell. Loss of the ability to bind AMP, we demonstrate, causes protein, and likely dimer, instability and consequent loss of function. Kef system function is regulated via the reversible binding of comparatively low-affinity glutathione-based ligands at the interface between the dimer subunits. We propose this interfacial binding site is itself stabilized, at least in part, by AMP binding. PMID:28656748

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.

Ion-binding properties of Calnuc, Ca2+ versus Mg2+--Calnuc adopts additional and unusual Ca2+-binding sites upon interaction with G-protein.

PubMed

Kanuru, Madhavi; Samuel, Jebakumar J; Balivada, Lavanya M; Aradhyam, Gopala K

2009-05-01

Calnuc is a novel, highly modular, EF-hand containing, Ca(2+)-binding, Golgi resident protein whose functions are not clear. Using amino acid sequences, we demonstrate that Calnuc is a highly conserved protein among various organisms, from Ciona intestinalis to humans. Maximum homology among all sequences is found in the region that binds to G-proteins. In humans, it is known to be expressed in a variety of tissues, and it interacts with several important protein partners. Among other proteins, Calnuc is known to interact with heterotrimeric G-proteins, specifically with the alpha-subunit. Herein, we report the structural implications of Ca(2+) and Mg(2+) binding, and illustrate that Calnuc functions as a downstream effector for G-protein alpha-subunit. Our results show that Ca(2+) binds with an affinity of 7 mum and causes structural changes. Although Mg(2+) binds to Calnuc with very weak affinity, the structural changes that it causes are further enhanced by Ca(2+) binding. Furthermore, isothermal titration calorimetry results show that Calnuc and the G-protein bind with an affinity of 13 nm. We also predict a probable function for Calnuc, that of maintaining Ca(2+) homeostasis in the cell. Using Stains-all and terbium as Ca(2+) mimic probes, we demonstrate that the Ca(2+)-binding ability of Calnuc is governed by the activity-based conformational state of the G-protein. We propose that Calnuc adopts structural sites similar to the ones seen in proteins such as annexins, c2 domains or chromogrannin A, and therefore binds more calcium ions upon binding to Gialpha. With the number of organelle-targeted G-protein-coupled receptors increasing, intracellular communication mediated by G-proteins could become a new paradigm. In this regard, we propose that Calnuc could be involved in the downstream signaling of G-proteins.

Analytical strategies based on quantum dots for heavy metal ions detection.

PubMed

Vázquez-González, Margarita; Carrillo-Carrion, Carolina

2014-01-01

Heavy metal contamination is one of the major concerns to human health because these substances are toxic and retained by the ecological system. Therefore, in recent years, there has been a pressing need for fast and reliable methods for the analysis of heavy metal ions in environmental and biological samples. Quantum dots (QDs) have facilitated the development of sensitive sensors over the past decade, due to their unique photophysical properties, versatile surface chemistry and ligand binding ability, and the possibility of the encapsulation in different materials or attachment to different functional materials, while retaining their native luminescence property. This paper comments on different sensing strategies with QD for the most toxic heavy metal ions (i.e., cadmium, Cd2+; mercury, Hg2+; and lead, Pb2+). Finally, the challenges and outlook for the QD-based sensors for heavy metals ions are discussed.

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.

Sequence-specific binding of counterions to B-DNA

PubMed Central

Denisov, Vladimir P.; Halle, Bertil

2000-01-01

Recent studies by x-ray crystallography, NMR, and molecular simulations have suggested that monovalent counterions can penetrate deeply into the minor groove of B form DNA. Such groove-bound ions potentially could play an important role in AT-tract bending and groove narrowing, thereby modulating DNA function in vivo. To address this issue, we report here 23Na magnetic relaxation dispersion measurements on oligonucleotides, including difference experiments with the groove-binding drug netropsin. The exquisite sensitivity of this method to ions in long-lived and intimate association with DNA allows us to detect sequence-specific sodium ion binding in the minor groove AT tract of three B-DNA dodecamers. The sodium ion occupancy is only a few percent, however, and therefore is not likely to contribute importantly to the ensemble of B-DNA structures. We also report results of ion competition experiments, indicating that potassium, rubidium, and cesium ions bind to the minor groove with similarly weak affinity as sodium ions, whereas ammonium ion binding is somewhat stronger. The present findings are discussed in the light of previous NMR and diffraction studies of sequence-specific counterion binding to DNA. PMID:10639130

Decarboxylation of bovine prothrombin fragment 1 and prothrombin.

PubMed

Tuhy, P M; Bloom, J W; Mann, K G

1979-12-25

Bovine prothrombin fragment 1 and prothrombin undergo decarboxylation of their gamma-carboxyglutamic acid residues when the lyophilized proteins are heated in vacuo at 110 degrees C for several hours. The fully decarboxylated fragment 1 product has lost its barium-binding ability as well as the calcium-binding function which causes fluorescence quenching in the presence of 2 mM Ca2+. There is no sign of secondary structure alteration in solution upon analysis by fluorescence emission and circular dichroic spectroscopy. A family of partially decarboxylated fragment 1 species generated by heating for shorter periods shows that the initial decrease in calcium-binding ability occurs almost twice as rapidly as the loss of gamma-carboxyglutamic acid. This is consistent with the idea that differential functions can be ascribed to the 10 gamma-carboxyglutamic acid residues in fragment 1, including both high- and low-affinity metal ion binding sites. Prothrombin itself also undergoes total decarboxylation without any apparent alteration in secondary structure. However, in this case the latent thrombin activity is progressively diminished during the heating process in terms of both clotting activity and hydrolysis of the amide substrate H-D-Phe-Pip-Arg-pNA. The present results indicate that in vitro decarboxylation of gamma-carboxyglutamic acid in dried proteins is useful for analyzing the detailed calcium-binding proteins of vitamin K dependent coagulation factors.

Facile synthesis of Fe3O4@PDA core-shell microspheres functionalized with various metal ions: A systematic comparison of commonly-used metal ions for IMAC enrichment.

PubMed

Jiang, Jiebing; Sun, Xueni; Li, Yan; Deng, Chunhui; Duan, Gengli

2018-02-01

Metal ions differed greatly in affinity towards phosphopeptides, and thus it is essential to systematically compare the phosphopeptides enrichment ability of different metal ions usually used in the IMAC techniques. In this work, for the first time, eight metal ions, including Nb 5+ , Ti 4+ , Zr 4+ , Ga 3+ , Y 3+ , In 3+ , Ce 4+ , Fe 3+ , were immobilized on the polydopamine (PDA)-coated Fe 3 O 4 (denoted as Fe 3 O 4 @PDA-M n+ ), and systematically compared by the real biosamples, in addition to standard phosphopeptides. Fe 3 O 4 microspheres were synthesized via the solvothermal reaction, followed by self-polymerization of dopamine on the surface. Then through taking advantage of the hydroxyl and amino group of PDA, the eight metal ions were easily adhered to the surface of Fe 3 O 4 @PDA. After characterization, the resultant Fe 3 O 4 @PDA-M n+ microspheres were applied to phosphopeptides enrichment based on the binding affinity between metal ions and phosphopeptides. According to the results, different metal ions presented diverse phosphopeptides enrichment efficiency in terms of selectivity, sensitivity and the enrichment ability from real complex samples, and Fe 3 O 4 @PDA-Nb 5+ and Fe 3 O 4 @PDA-Ti 4+ showed obvious advantages of the phosphopeptides enrichment effect after the comparison. This systematic comparison may provide certain reference for the use and development of IMAC materials in the future. Copyright © 2017 Elsevier B.V. All rights reserved.

Multi-shell model of ion-induced nucleic acid condensation

NASA Astrophysics Data System (ADS)

Tolokh, Igor S.; Drozdetski, Aleksander V.; Pollack, Lois; Baker, Nathan A.; Onufriev, Alexey V.

2016-04-01

We present a semi-quantitative model of condensation of short nucleic acid (NA) duplexes induced by trivalent cobalt(iii) hexammine (CoHex) ions. The model is based on partitioning of bound counterion distribution around single NA duplex into "external" and "internal" ion binding shells distinguished by the proximity to duplex helical axis. In the aggregated phase the shells overlap, which leads to significantly increased attraction of CoHex ions in these overlaps with the neighboring duplexes. The duplex aggregation free energy is decomposed into attractive and repulsive components in such a way that they can be represented by simple analytical expressions with parameters derived from molecular dynamic simulations and numerical solutions of Poisson equation. The attractive term depends on the fractions of bound ions in the overlapping shells and affinity of CoHex to the "external" shell of nearly neutralized duplex. The repulsive components of the free energy are duplex configurational entropy loss upon the aggregation and the electrostatic repulsion of the duplexes that remains after neutralization by bound CoHex ions. The estimates of the aggregation free energy are consistent with the experimental range of NA duplex condensation propensities, including the unusually poor condensation of RNA structures and subtle sequence effects upon DNA condensation. The model predicts that, in contrast to DNA, RNA duplexes may condense into tighter packed aggregates with a higher degree of duplex neutralization. An appreciable CoHex mediated RNA-RNA attraction requires closer inter-duplex separation to engage CoHex ions (bound mostly in the "internal" shell of RNA) into short-range attractive interactions. The model also predicts that longer NA fragments will condense more readily than shorter ones. The ability of this model to explain experimentally observed trends in NA condensation lends support to proposed NA condensation picture based on the multivalent "ion binding shells."

Bi-functional, substrate mimicking RNA inhibits MSK1-mediated cAMP-response element-binding protein phosphorylation and reveals magnesium ion-dependent conformational changes of the kinase.

PubMed

Hamm, Jorg; Alessi, Dario R; Biondi, Ricardo M

2002-11-29

The design of specific inhibitors for protein kinases is an important step toward elucidation of intracellular signal transduction pathways and to guide drug discovery programs. We devised a model approach to generate specific, competitive kinase inhibitors by isolating substrate mimics containing two independent binding sites with an anti-idiotype strategy from combinatorial RNA libraries. As a general test for the ability to generate highly specific kinase inhibitors, we selected the transcription factor cAMP-response element-binding protein (CREB) that is phosphorylated on the same serine residue by the protein kinase MSK1 as well as by RSK1. The sequences and structures of these kinases are very similar, about 60% of their amino acids are identical. Nevertheless, we can demonstrate that the selected RNA inhibitors inhibit specifically CREB phosphorylation by MSK1 but do not affect CREB phosphorylation by RSK1. The inhibitors interact preferentially with the inactive form of MSK1. Furthermore, we demonstrate that RNA ligands can be conformation-specific probes, and this feature allowed us to describe magnesium ion-dependent conformational changes of MSK1 upon activation.

Iron-Binding Protein Degradation by Cysteine Proteases of Naegleria fowleri.

PubMed

Martínez-Castillo, Moisés; Ramírez-Rico, Gerardo; Serrano-Luna, Jesús; Shibayama, Mineko

2015-01-01

Naegleria fowleri causes acute and fulminant primary amoebic meningoencephalitis. This microorganism invades its host by penetrating the olfactory mucosa and then traveling up the mesaxonal spaces and crossing the cribriform plate; finally, the trophozoites invade the olfactory bulbs. During its invasion, the protozoan obtains nutrients such as proteins, lipids, carbohydrates, and cationic ions (e.g., iron, calcium, and sodium) from the host. However, the mechanism by which these ions are obtained, particularly iron, is poorly understood. In the present study, we evaluated the ability of N. fowleri to degrade iron-binding proteins, including hololactoferrin, transferrin, ferritin, and hemoglobin. Zymography assays were performed for each substrate under physiological conditions (pH 7 at 37°C) employing conditioned medium (CM) and total crude extracts (TCEs) of N. fowleri. Different degradation patterns with CM were observed for hololactoferrin, transferrin, and hemoglobin; however, CM did not cause ferritin degradation. In contrast, the TCEs degraded only hololactoferrin and transferrin. Inhibition assays revealed that cysteine proteases were involved in this process. Based on these results, we suggest that CM and TCEs of N. fowleri degrade iron-binding proteins by employing cysteine proteases, which enables the parasite to obtain iron to survive while invading the central nervous system.

Iron-Binding Protein Degradation by Cysteine Proteases of Naegleria fowleri

PubMed Central

Ramírez-Rico, Gerardo; Serrano-Luna, Jesús; Shibayama, Mineko

2015-01-01

Naegleria fowleri causes acute and fulminant primary amoebic meningoencephalitis. This microorganism invades its host by penetrating the olfactory mucosa and then traveling up the mesaxonal spaces and crossing the cribriform plate; finally, the trophozoites invade the olfactory bulbs. During its invasion, the protozoan obtains nutrients such as proteins, lipids, carbohydrates, and cationic ions (e.g., iron, calcium, and sodium) from the host. However, the mechanism by which these ions are obtained, particularly iron, is poorly understood. In the present study, we evaluated the ability of N. fowleri to degrade iron-binding proteins, including hololactoferrin, transferrin, ferritin, and hemoglobin. Zymography assays were performed for each substrate under physiological conditions (pH 7 at 37°C) employing conditioned medium (CM) and total crude extracts (TCEs) of N. fowleri. Different degradation patterns with CM were observed for hololactoferrin, transferrin, and hemoglobin; however, CM did not cause ferritin degradation. In contrast, the TCEs degraded only hololactoferrin and transferrin. Inhibition assays revealed that cysteine proteases were involved in this process. Based on these results, we suggest that CM and TCEs of N. fowleri degrade iron-binding proteins by employing cysteine proteases, which enables the parasite to obtain iron to survive while invading the central nervous system. PMID:26090408

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.

Friedel's salt formation in sulfoaluminate cements: A combined XRD and {sup 27}Al MAS NMR study

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

Paul, G.; Boccaleri, E., E-mail: enrico.boccaleri@mfn.unipmn.it; Buzzi, L.

Four different binders based on calcium sulfoaluminate cements have been submitted to accelerated chlorination through ionic exchange on hydrated pastes, in order to investigate their ability to chemically bind chloride ions that might reduce chloride penetration. The composition of hydrated cements before and after the treatment was evaluated by means of an X-Ray Diffraction–{sup 27}Al Magic Angle Spinning Nuclear Magnetic Resonance Spectroscopy combined study, allowing to take into account even partially amorphous phases and to make quantitative assumption on the relative abundance of the different aluminium-containing phases. It was found that low SO{sub 3} Sulfoaluminate–Portland ternary systems are the mostmore » effective in binding chloride ions and the active role played by different members of the AFm family in chloride uptake was confirmed. Moreover, a peculiar behavior related to the formation of Friedel's salt in different pH conditions was also established for the different cements.« less

Albumin Redhill, a human albumin variant.

PubMed

Brand, S; Hutchinson, D W; Donaldson, D

1984-01-31

Albumin Redhill, a variant human albumin with the same C-terminal amino acid as albumin A but with arginine at the N-terminus has been isolated by chromatofocusing from the sera of an English family. Albumin Redhill appears to contain two sites of mutation in its protein chain and is probably a proalbumin. The ability of albumin Redhill to bind Ni(II) or Cu(II) ions is considerably less than that of albumin A.

Ion-binding properties of the ClC chloride selectivity filter

PubMed Central

Lobet, Séverine; Dutzler, Raimund

2006-01-01

The ClC channels are members of a large protein family of chloride (Cl−) channels and secondary active Cl− transporters. Despite their diverse functions, the transmembrane architecture within the family is conserved. Here we present a crystallographic study on the ion-binding properties of the ClC selectivity filter in the close homolog from Escherichia coli (EcClC). The ClC selectivity filter contains three ion-binding sites that bridge the extra- and intracellular solutions. The sites bind Cl− ions with mM affinity. Despite their close proximity within the filter, the three sites can be occupied simultaneously. The ion-binding properties are found conserved from the bacterial transporter EcClC to the human Cl− channel ClC-1, suggesting a close functional link between ion permeation in the channels and active transport in the transporters. In resemblance to K+ channels, ions permeate the ClC channel in a single file, with mutual repulsion between the ions fostering rapid conduction. PMID:16341087

FecB, a periplasmic ferric-citrate transporter from E. coli, can bind different forms of ferric-citrate as well as a wide variety of metal-free and metal-loaded tricarboxylic acids.

PubMed

Banerjee, Sambuddha; Paul, Subrata; Nguyen, Leonard T; Chu, Byron C H; Vogel, Hans J

2016-01-01

The Escherichia coli Fec system, consisting of an outer membrane receptor (FecA), a periplasmic substrate binding protein (FecB) and an inner membrane permease-ATPase type transporter (FecC/D), plays an important role in the uptake and transport of Fe(3+)-citrate. Although several FecB sequences from various organisms have been reported, there are no biophysical or structural data available for this protein to date. In this work, using isothermal titration calorimetry (ITC), we report for the first time the ability of FecB to bind different species of Fe(3+)-citrate as well as other citrate complexes with trivalent (Ga(3+), Al(3+), Sc(3+) and In(3+)) and a representative divalent metal ion (Mg(2+)) with low μM affinity. Interestingly, ITC experiments with various iron-free di- and tricarboxylic acids show that FecB can bind tricarboxylates with μM affinity but not biologically relevant dicarboxylates. The ability of FecB to bind with metal-free citrate is also observed in (1)H,(15)N HSQC-NMR titration experiments reported here at two different pH values. Further, differential scanning calorimetry (DSC) experiments indicate that the ligand-bound form of FecB has greater thermal stability than ligand-free FecB under all pH and ligand conditions tested, which is consistent with the idea of domain closure subsequent to ligand binding for this type of periplasmic binding proteins.

Calcium ion binding to a soil fulvic acid using a donnan potential model

USGS Publications Warehouse

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

1999-01-01

Calcium ion binding to a soil fulvic acid (Armadale Bh Horizon) was evaluated over a range of calcium ion concentrations, from pH 3.8 to 7.3, using potentiometric titrations and calcium ion electrode measurements. Fulvic acid concentration was constant (100 milligrams per liter) and calcium ion concentration varied up to 8 X 10-4 moles per liter. Experiments discussed here included: (1) titrations of fulvic acid-calcium ion containing solutions with sodium hydroxide; and (2) titrations of fully neutralized fulvic acid with calcium chloride solutions. Apparent binding constants (expressed as the logarithm of the value, log ??app) vary with solution pH, calcium ion concentration, degree of acid dissociation, and ionic strength (from log ??app = 2.5 to 3.9) and are similar to those reported by others. Fulvic acid charge, and the associated Donnan Potential, influences calcium ion-fulvic acid ion pair formation. A Donnan Potential corrrection term allowed calculation of intrinsic calcium ion-fulvic acid binding constants. Intrinsic binding constants vary from 1.2 to 2.5 (the average value is about log??= 1.6) and are similar to, but somewhat higher than, stability constants for calcium ion-carboxylic acid monodentate complexes. ?? by Oldenbourg Wissenschaftsverlag, Mu??nchen.

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.

Synthesis, characterization and extraction studies of some metal (II) complexes containing (hydrazoneoxime and bis-acylhydrazone) moieties

NASA Astrophysics Data System (ADS)

Al-Ne'aimi, Mohammed Mahmmod; Al-Khuder, Mohammed Moudar

2013-03-01

In this study, diacetylmonoximebenzoylhydrazone (L1H2) and 1,4-diacetylbenzene bis(benzoyl hydrazone) (L2H2) were synthesized by the condensation of benzohydrazide with diacetyl monoxime and 1,4-diacetylbenzene, respectively. Complexes of these ligands with Co(II), Ni(II), Cu(II), Zn(II) and Cd(II) inos were prepared with a metal:ligand ratio of 1:2 for L1H2 ligand, and 1:1 for L2H2 ligand. The ligands and their complexes were elucidated on the basis of elemental analyses CHN, AAS, FT-IR, 1H- and 13C NMR spectra, UV-vis spectra and magnetic susceptibility measurements. Results show the L1H2 ligand act as monoanionic O,N,N-tridentate and coordination takes place in the enol form through the oxime nitrogen, the imine nitrogen and the enolate oxygen atoms with a N4O2 donor environment, while the L2H2 ligand act as a dianionic O,N,N,O-tetradentate and coordination takes place in the enol form through the enolate oxygen and the azomethine nitrogen atoms with a N2O2 donor environment. These results are consistent with the formation of mononuclear metal (II) complexes [M(L1H)2], and binuclear polymeric metal (II) complexes [{M2(L2)}n]. The extraction ability of both ligands were examined in chloroform by the liquid-liquid extraction of selected transition metal [Co2+, Ni2+, Cu2+, Zn2+ and Pb2+] cations. The effects of pH and contact time on the percentage extraction of metal (II) ions were studied under the optimum extraction conditions. The (L1H2) ligand shows strong binding ability toward copper(II) and lead(II) ions, while the (L2H2) ligand shows strong binding ability toward nickel(II) and zinc(II) ions.

Synthesis, characterization and extraction studies of some metal (II) complexes containing (hydrazoneoxime and bis-acylhydrazone) moieties.

PubMed

Al-Ne'aimi, Mohammed Mahmmod; Al-Khuder, Mohammed Moudar

2013-03-15

In this study, diacetylmonoximebenzoylhydrazone (L(1)H(2)) and 1,4-diacetylbenzene bis(benzoyl hydrazone) (L(2)H(2)) were synthesized by the condensation of benzohydrazide with diacetyl monoxime and 1,4-diacetylbenzene, respectively. Complexes of these ligands with Co(II), Ni(II), Cu(II), Zn(II) and Cd(II) inos were prepared with a metal:ligand ratio of 1:2 for L(1)H(2) ligand, and 1:1 for L(2)H(2) ligand. The ligands and their complexes were elucidated on the basis of elemental analyses CHN, AAS, FT-IR, (1)H- and (13)C NMR spectra, UV-vis spectra and magnetic susceptibility measurements. Results show the L(1)H(2) ligand act as monoanionic O,N,N-tridentate and coordination takes place in the enol form through the oxime nitrogen, the imine nitrogen and the enolate oxygen atoms with a N(4)O(2) donor environment, while the L(2)H(2) ligand act as a dianionic O,N,N,O-tetradentate and coordination takes place in the enol form through the enolate oxygen and the azomethine nitrogen atoms with a N(2)O(2) donor environment. These results are consistent with the formation of mononuclear metal (II) complexes [M(L(1)H)(2)], and binuclear polymeric metal (II) complexes [{M(2)(L(2))}(n)]. The extraction ability of both ligands were examined in chloroform by the liquid-liquid extraction of selected transition metal [Co(2+), Ni(2+), Cu(2+), Zn(2+) and Pb(2+)] cations. The effects of pH and contact time on the percentage extraction of metal (II) ions were studied under the optimum extraction conditions. The (L(1)H(2)) ligand shows strong binding ability toward copper(II) and lead(II) ions, while the (L(2)H(2)) ligand shows strong binding ability toward nickel(II) and zinc(II) ions. Copyright © 2012 Elsevier B.V. All rights reserved.

Physical chemistry and membrane properties of two phosphatidylinositol bisphosphate isomers.

PubMed

Slochower, David R; Wang, Yu-Hsiu; Radhakrishnan, Ravi; Janmey, Paul A

2015-05-21

The most highly charged phospholipids, polyphosphoinositides, are often involved in signaling pathways that originate at cell-cell and cell-matrix contacts, and different isomers of polyphosphoinositides have distinct biological functions that cannot be explained by separate highly specific protein ligand binding sites [Lemmon, Nat. Rev. Mol. Cell Biol., 2008, 9, 99-111]. PtdIns(3,5)P2 is a low abundance phosphoinositide localized to cytoplasmic-facing membrane surfaces, with relatively few known ligands, yet PtdIns(3,5)P2 plays a key role in controlling membrane trafficking events and cellular stress responses that cannot be duplicated by other phosphoinositides [Dove et al., Nature, 1997, 390, 187-192; Michell, FEBS J., 2013, 280, 6281-6294]. Here we show that PtdIns(3,5)P2 is structurally distinct from PtdIns(4,5)P2 and other more common phospholipids, with unique physical chemistry. Using multiscale molecular dynamics techniques on the quantum level, single molecule, and in bilayer settings, we found that the negative charge of PtdIns(3,5)P2 is spread over a larger area, compared to PtdIns(4,5)P2, leading to a decreased ability to bind divalent ions. Additionally, our results match well with experimental data characterizing the cluster forming potential of these isomers in the presence of Ca(2+) [Wang et al., J. Am. Chem. Soc., 2012, 134, 3387-3395; van den Bogaart et al., Nature, 2011, 479, 552-555]. Our results demonstrate that the different cellular roles of PtdIns(4,5)P2 and PtdIns(3,5)P2in vivo are not simply determined by their localization by enzymes that produce or degrade them, but also by their molecular size, ability to chelate ions, and the partial dehydration of those ions, which might affect the ability of PtdIns(3,5)P2 and PtdIns(4,5)P2 to form phosphoinositide-rich clusters in vitro and in vivo.

Receptor binding of somatostatin-14 and somatostatin-28 in rat brain: differential modulation by nucleotides and ions.

PubMed

Srikant, C B; Dahan, A; Craig, C

1990-02-04

The tissue-selective binding of the two principal bioactive forms of somatostatin, somatostatin-14 (SS-14) and somatostatin-28 (SS-28), their ability to modulate cAMP-dependent and -independent regulation of post-receptor events to different degrees and the documentation of specific labelling of SS receptor subtypes with SS-28 but not SS-14 in discrete regions of rat brain suggest the existence of distinct SS-14 and SS-28 binding sites. Receptor binding of SS-14 ligands has been shown to be modulated by nucleotides and ions, but the effect of these agents on SS-28 binding has not been studied. In the present study we investigated the effects of adenine and guanine nucleotides as well as monovalent and divalent cations on rat brain SS receptors quantitated with radioiodinated analogs of SS-14 ([125I-Tyr11]SS14, referred to in this paper as SS-14) and SS-28 ([Leu8, D-Trp22, 125I-Tyr25] SS-28, referred to as LTT* SS-28) in order to determine if distinct receptor sites for SS-14 and SS-28 could be distinguished on the basis of their modulation by nucleotides and ions. GTP as well as ATP exerted a dose-dependent inhibition (over a concentration range of 10(-7)-10(-3) M) of the binding of the two radioligands. The nucleotide inhibition of binding resulted in a decrease the Bmax of the SS receptors, the binding affinity remaining unaltered. GTP (10(-4) M) decreased the Bmax of LTT* SS-28 binding sites to a greater extent than ATP (145 +/- 10 and 228 +/- 16 respectively, compared to control value of 320 +/- 20 pmol mg-1). Under identical conditions GTP was less effective than ATP in reducing the number of T* SS-14 binding sites (Bmax = 227 +/- 8 and 182 +/- 15, respectively, compared to 340 +/- 15 pmol mg-1 in the absence of nucleotides). Monovalent cations inhibited the binding of both radioligands, Li+ and Na+ inhibited the binding of T* SS-14 to a greater extent than K+. The effect of divalent cations on the other hand was varied. At low concentration (2 mM) Mg2+, Ba2+, Mn2+, Ca2+ and Co2+ augmented the binding of both T* SS-14 and LTT* SS-28, while higher than 4 mM Co2+ inhibited binding of both ligands. LTT* SS-28 binding was reduced in the presence of high concentrations of Ba2+ and Mn2+ also. Interestingly Ca2+ at higher than 10 mM preferentially inhibited LTT* SS-28 binding and increased the affinity of SS-14 but not SS-28 for LTT* SS-28 binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)

Evolution reversed: the ability to bind iron restored to the N-lobe of the murine inhibitor of carbonic anhydrase by strategic mutagenesis.

PubMed

Mason, Anne B; Judson, Gregory L; Bravo, Maria Cristina; Edelstein, Andrew; Byrne, Shaina L; James, Nicholas G; Roush, Eric D; Fierke, Carol A; Bobst, Cedric E; Kaltashov, Igor A; Daughtery, Margaret A

2008-09-16

The murine inhibitor of carbonic anhydrase (mICA) is a member of the superfamily related to the bilobal iron transport protein transferrin (TF), which binds a ferric ion within a cleft in each lobe. Although the gene encoding ICA in humans is classified as a pseudogene, an apparently functional ICA gene has been annotated in mice, rats, cows, pigs, and dogs. All ICAs lack one (or more) of the amino acid ligands in each lobe essential for high-affinity coordination of iron and the requisite synergistic anion, carbonate. The reason why ICA family members have lost the ability to bind iron is potentially related to acquiring a new function(s), one of which is inhibition of certain carbonic anhydrase (CA) isoforms. A recombinant mutant of the mICA (W124R/S188Y) was created with the goal of restoring the ligands required for both anion (Arg124) and iron (Tyr188) binding in the N-lobe. Absorption and fluorescence spectra definitively show that the mutant binds ferric iron in the N-lobe. Electrospray ionization mass spectrometry confirms the presence of both ferric iron and carbonate. At the putative endosomal pH of 5.6, iron is released by two slow processes indicative of high-affinity coordination. Induction of specific iron binding implies that (1) the structure of mICA resembles those of other TF family members and (2) the N-lobe can adopt a conformation in which the cleft closes when iron binds. Because the conformational change in the N-lobe indicated by metal binding does not impact the inhibitory activity of mICA, inhibition of CA was tentatively assigned to the C-lobe. Proof of this assignment is provided by limited trypsin proteolysis of porcine ICA.

Binding of Capsaicin to the TRPV1 Ion Channel.

PubMed

Darré, Leonardo; Domene, Carmen

2015-12-07

Transient receptor potential (TRP) ion channels constitute a notable family of cation channels involved in the ability of an organisms to detect noxious mechanical, thermal, and chemical stimuli that give rise to the perception of pain, taste, and changes in temperature. One of the most experimentally studied agonist of TRP channels is capsaicin, which is responsible for the burning sensation produced when chili pepper is in contact with organic tissues. Thus, understanding how this molecule interacts and regulates TRP channels is essential to high impact pharmacological applications, particularly those related to pain treatment. The recent publication of a three-dimensional structure of the vanilloid receptor 1 (TRPV1) in the absence and presence of capsaicin from single particle electron cryomicroscopy experiments provides the opportunity to explore these questions at the atomic level. In the present work, molecular docking and unbiased and biased molecular dynamics simulations were employed to generate a structural model of the capsaicin-channel complex. In addition, the standard free energy of binding was estimated using alchemical transformations coupled with conformational, translational, and orientational restraints on the ligand. Key binding modes consistent with previous experimental data are identified, and subtle but essential dynamical features of the binding site are characterized. These observations shed some light into how TRPV1 interacts with capsaicin, and may help to refine design parameters for new TRPV1 antagonists, and potentially guide further developments of TRP channel modulators.

Bovine Leukemia Virus SU Protein Interacts with Zinc, and Mutations within Two Interacting Regions Differently Affect Viral Fusion and Infectivity In Vivo

PubMed Central

Gatot, Jean-Stéphane; Callebaut, Isabelle; Van Lint, Carine; Demonté, Dominique; Kerkhofs, Pierre; Portetelle, Daniel; Burny, Arsène; Willems, Luc; Kettmann, Richard

2002-01-01

Bovine leukemia virus (BLV) and human T-cell lymphotropic virus type 1 (HTLV-1) belong to the genus of deltaretroviruses. Their entry into the host cell is supposed to be mediated by interactions of the extracellular (SU) envelope glycoproteins with cellular receptors. To gain insight into the mechanisms governing this process, we investigated the ability of SU proteins to interact with specific ligands. In particular, by affinity chromatography, we have shown that BLV SU protein specifically interacted with zinc ions. To identify the protein domains involved in binding, 16 peptides distributed along the sequence were tested. Two of them appeared to be able to interact with zinc. To unravel the role of these SU regions in the biology of the virus, mutations were introduced into the env gene of a BLV molecular clone in order to modify residues potentially interacting with zinc. The fusogenic capacity of envelope mutated within the first zinc-binding region (104 to 123) was completely abolished. Furthermore, the integrity of this domain was also required for in vivo infectivity. In contrast, mutations within the second zinc-binding region (218 to 237) did not hamper the fusogenic capacity; indeed, the syncytia were even larger. In sheep, mutations in region 218 to 237 did not alter infectivity or viral spread. Finally, we demonstrated that the envelope of the related HTLV-1 was also able to bind zinc. Interestingly, zinc ions were found to be associated with the receptor-binding domain (RBD) of Friend murine leukemia virus (Fr-MLV) SU glycoprotein, further supporting their relevance in SU structure. Based on the sequence similarities shared with the Fr-MLV RBD, whose three-dimensional structure has been experimentally determined, we located the BLV zinc-binding peptide 104-123 on the opposite side of the potential receptor-binding surface. This observation supports the hypothesis that zinc ions could mediate interactions of the SU RBD either with the C-terminal part of SU, thereby contributing to the SU structural integrity, or with a partner(s) different from the receptor. PMID:12134000

Linked supramolecular building blocks for enhanced cluster formation

DOE PAGES

McLellan, Ross; Palacios, Maria A.; Beavers, Christine M.; ...

2015-01-09

Methylene-bridged calix[4]arenes have emerged as extremely versatile ligand supports in the formation of new polymetallic clusters possessing fascinating magnetic properties. Metal ion binding rules established for this building block allow one to partially rationalise the complex assembly process. The ability to covalently link calix[4]arenes at the methylene bridge provides significantly improved control over the introduction of different metal centres to resulting cluster motifs. Clusters assembled from bis-calix[4]arenes and transition metal ions or 3d-4f combinations display characteristic features of the analogous calix[4]arene supported clusters, thereby demonstrating an enhanced and rational approach towards the targeted synthesis of complex and challenging structures.

Electronically conductive polymer binder for lithium-ion battery electrode

DOEpatents

Liu, Gao; Xun, Shidi; Battaglia, Vincent S.; Zheng, Honghe; Wu, Mingyan

2015-07-07

A family of carboxylic acid groups containing fluorene/fluorenon copolymers is disclosed as binders of silicon particles in the fabrication of negative electrodes for use with lithium ion batteries. Triethyleneoxide side chains provide improved adhesion to materials such as, graphite, silicon, silicon alloy, tin, tin alloy. These binders enable the use of silicon as an electrode material as they significantly improve the cycle-ability of silicon by preventing electrode degradation over time. In particular, these polymers, which become conductive on first charge, bind to the silicon particles of the electrode, are flexible so as to better accommodate the expansion and contraction of the electrode during charge/discharge, and being conductive promote the flow battery current.

Electronically conductive polymer binder for lithium-ion battery electrode

DOEpatents

Liu, Gao; Battaglia, Vincent S.; Park, Sang -Jae

2015-10-06

A family of carboxylic acid groups containing fluorene/fluorenon copolymers is disclosed as binders of silicon particles in the fabrication of negative electrodes for use with lithium ion batteries. Triethyleneoxide side chains provide improved adhesion to materials such as, graphite, silicon, silicon alloy, tin, tin alloy. These binders enable the use of silicon as an electrode material as they significantly improve the cycle-ability of silicon by preventing electrode degradation over time. In particular, these polymers, which become conductive on first charge, bind to the silicon particles of the electrode, are flexible so as to better accommodate the expansion and contraction of the electrode during charge/discharge, and being conductive promote the flow battery current.

Electronically conductive polymer binder for lithium-ion battery electrode

DOEpatents

Liu, Gao; Xun, Shidi; Battaglia, Vincent S.; Zheng, Honghe; Wu, Mingyan

2017-08-01

A family of carboxylic acid groups containing fluorene/fluorenon copolymers is disclosed as binders of silicon particles in the fabrication of negative electrodes for use with lithium ion batteries. Triethyleneoxide side chains provide improved adhesion to materials such as, graphite, silicon, silicon alloy, tin, tin alloy. These binders enable the use of silicon as an electrode material as they significantly improve the cycle-ability of silicon by preventing electrode degradation over time. In particular, these polymers, which become conductive on first charge, bind to the silicon particles of the electrode, are flexible so as to better accommodate the expansion and contraction of the electrode during charge/discharge, and being conductive promote the flow battery current.

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

Equivalence of two approaches for modeling ion permeation through a transmembrane channel with an internal binding site

NASA Astrophysics Data System (ADS)

Zhou, Huan-Xiang

2011-04-01

Ion permeation through transmembrane channels has traditionally been modeled using two different approaches. In one approach, the translocation of the permeant ion through the channel pore is modeled as continuous diffusion and the rate of ion transport is obtained from solving the steady-state diffusion equation. In the other approach, the translocation of the permeant ion through the pore is modeled as hopping along a discrete set of internal binding sites and the rate of ion transport is obtained from solving a set of steady-state rate equations. In a recent work [Zhou, J. Phys. Chem. Lett. 1, 1973 (2010)], the rate constants for binding to an internal site were further calculated by modeling binding as diffusion-influenced reactions. That work provided the foundation for bridging the two approaches. Here we show that, by representing a binding site as an energy well, the two approaches indeed give the same result for the rate of ion transport.

Modeling the human Nav1.5 sodium channel: structural and mechanistic insights of ion permeation and drug blockade

PubMed Central

Ahmed, Marawan; Jalily Hasani, Horia; Ganesan, Aravindhan; Houghton, Michael; Barakat, Khaled

2017-01-01

Abnormalities in the human Nav1.5 (hNav1.5) voltage-gated sodium ion channel (VGSC) are associated with a wide range of cardiac problems and diseases in humans. Current structural models of hNav1.5 are still far from complete and, consequently, their ability to study atomistic interactions of this channel is very limited. Here, we report a comprehensive atomistic model of the hNav1.5 ion channel, constructed using homology modeling technique and refined through long molecular dynamics simulations (680 ns) in the lipid membrane bilayer. Our model was comprehensively validated by using reported mutagenesis data, comparisons with previous models, and binding to a panel of known hNav1.5 blockers. The relatively long classical MD simulation was sufficient to observe a natural sodium permeation event across the channel’s selectivity filters to reach the channel’s central cavity, together with the identification of a unique role of the lysine residue. Electrostatic potential calculations revealed the existence of two potential binding sites for the sodium ion at the outer selectivity filters. To obtain further mechanistic insight into the permeation event from the central cavity to the intracellular region of the channel, we further employed “state-of-the-art” steered molecular dynamics (SMD) simulations. Our SMD simulations revealed two different pathways through which a sodium ion can be expelled from the channel. Further, the SMD simulations identified the key residues that are likely to control these processes. Finally, we discuss the potential binding modes of a panel of known hNav1.5 blockers to our structural model of hNav1.5. We believe that the data presented here will enhance our understanding of the structure–property relationships of the hNav1.5 ion channel and the underlying molecular mechanisms in sodium ion permeation and drug interactions. The results presented here could be useful for designing safer drugs that do not block the hNav1.5 channel. PMID:28831242

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

Two distinct modes of metal ion binding in the nuclease active site of a viral DNA-packaging terminase: insight into the two-metal-ion catalytic mechanism

PubMed Central

Zhao, Haiyan; Lin, Zihan; Lynn, Anna Y.; Varnado, Brittany; Beutler, John A.; Murelli, Ryan P.; Le Grice, Stuart F. J.; Tang, Liang

2015-01-01

Many dsDNA viruses encode DNA-packaging terminases, each containing a nuclease domain that resolves concatemeric DNA into genome-length units. Terminase nucleases resemble the RNase H-superfamily nucleotidyltransferases in folds, and share a two-metal-ion catalytic mechanism. Here we show that residue K428 of a bacteriophage terminase gp2 nuclease domain mediates binding of the metal cofactor Mg2+. A K428A mutation allows visualization, at high resolution, of a metal ion binding mode with a coupled-octahedral configuration at the active site, exhibiting an unusually short metal-metal distance of 2.42 Å. Such proximity of the two metal ions may play an essential role in catalysis by generating a highly positive electrostatic niche to enable formation of the negatively charged pentacovalent phosphate transition state, and provides the structural basis for distinguishing Mg2+ from Ca2+. Using a metal ion chelator β-thujaplicinol as a molecular probe, we observed a second mode of metal ion binding at the active site, mimicking the DNA binding state. Arrangement of the active site residues differs drastically from those in RNase H-like nucleases, suggesting a drifting of the active site configuration during evolution. The two distinct metal ion binding modes unveiled mechanistic details of the two-metal-ion catalysis at atomic resolution. PMID:26450964

Differential Cationization of Fatty Acids with Monovalent Cations Studied by ESI-MS/MS and Computational Approach.

PubMed

Sudarshana Reddy, B; Pavankumar, P; Sridhar, L; Saha, Soumen; Narahari Sastry, G; Prabhakar, S

2018-04-24

The intercellular and intracellular transport of charged species (Na + /K + ) entail interaction of the ions with neutral organic molecules and formation of adduct ions. The rate of transport of the ions across the cell membrane(s) may depend on the stability of the adduct ions, which in turn rely on structural aspects of the organic molecules that interact with the ions. Positive ion ESI mass spectra were recorded for the solutions containing fatty acids (FAs) and monovalent cations (X=Li + , Na + , K + , Rb + and Cs + ). Product ion spectra of the [FA+X] + ions were recorded at different collision energies. Theoretical studies were exploited under both gas phase and solvent phase to investigate the structural effects of the fatty acids during cationization. Stability of [FA+X] + adduct ions were further estimated by means of AIM topological analyses and interaction energy (IE) values. Positive ion ESI-MS analyses of the solution of FAs and X + ions showed preferential binding of the K + ions to FAs. The K + ion binding increased with the increase in number of double bonds of FAs, while decreased with increase in the number of carbons of FAs. Dissociation curves of [FA+X] + ions indicated the relative stability order of the [FA+X] + ions and it was in line with the observed trends in ESI-MS. The solvent phase computational studies divulged the mode of binding and the binding efficiencies of different FAs with monovalent cations. Among the studied monovalent cations, the cationization of FAs follow the order K + >Na + >Li + >Rb + >Cs + . The docosahexaenoic acid showed high efficiency in binding with K + ion. The K + ion binding efficiency of FAs depends on the number of double bonds in unsaturated FAs and the carbon chain length in saturated FAs. The cationization trends of FAs obtained from the ESI-MS, ESI-MS/MS analyses were in good agreement with solvent phase computational studies. This article is protected by copyright. All rights reserved.

Mechanism of Metal Ion Activation of the Diphtheria Toxin Repressor DtxR

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

D'Aquino,J.; Tetenbaum-Novatt, J.; White, A.

2005-01-01

The diphtheria toxin repressor (DtxR) is a metal ion-activated transcriptional regulator that has been linked to the virulence of Corynebacterium diphtheriae. Structure determination has shown that there are two metal ion binding sites per repressor monomer, and site-directed mutagenesis has demonstrated that binding site 2 (primary) is essential for recognition of the target DNA repressor, leaving the role of binding site 1 (ancillary) unclear. Calorimetric techniques have demonstrated that although binding site 1 (ancillary) has high affinity for metal ion with a binding constant of 2 x 10{sup -7}, binding site 2 (primary) is a low-affinity binding site with amore » binding constant of 6.3 x 10{sup -4}. These two binding sites act in an independent fashion, and their contribution can be easily dissected by traditional mutational analysis. Our results clearly demonstrate that binding site 1 (ancillary) is the first one to be occupied during metal ion activation, playing a critical role in stabilization of the repressor. In addition, structural data obtained for the mutants Ni-DtxR(H79A, C102D), reported here, and the previously reported DtxR(H79A) have allowed us to propose a mechanism of metal activation for DtxR.« less

Copper attachment to prion protein at a non-octarepeat site

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Bernholc, Jerry

2011-03-01

Prion protein (PrP) plays a causative role in a group of neurodegenerative diseases, which include ``mad cow disease'' or its human form variant Creutzfeld-Jacob disease. Normal function of PrP remains unknown, but it is now well established that PrP can efficiently bind copper ions and this ability has been linked to its function. The primary binding sites are located in the so-called octarepeat region located between residues 60-91. While these are by now well characterized, the sites located outside these region remain mostly undetermined. In this work, we investigate the properties of Cu binding site located at His 111 using recently developed hybrid Kohn-Sham/orbital-free density functional simulations. Experimental data indicate that copper is coordinated by either four nitrogens or three nitrogens and one oxygen. We investigate both possibilities, comparing their energetics and attachment geometries. Similarities and differences with other binding sites and implications for PrP function will also be discussed.

Identifying Cu( ii )–amyloid peptide binding intermediates in the early stages of aggregation by resonance Raman spectroscopy: a simulation study

DOE PAGES

Ren, Hao; Zhang, Yu; Guo, Sibei; ...

2017-10-31

The aggregation of amyloid beta (Aβ) peptides plays a crucial role in the pathology and etiology of Alzheimer's disease. Experimental evidence shows that copper ion is an aggregation-prone species with the ability to coordinately bind to Aβ and further induce the formation of neurotoxic Aβ oligomers. However, the detailed structures of Cu(II)–Aβ complexes have not been illustrated, and the kinetics and dynamics of the Cu(II) binding are not well understood. Two Cu(II)–Aβ complexes have been proposed to exist under physiological conditions, and another two might exist at higher pH values. By using ab initio simulations for the spontaneous resonance Ramanmore » and time domain stimulated resonance Raman spectroscopy signals, we obtained the characteristic Raman vibronic features of each complex. Finally, these signals contain rich structural information with high temporal resolution, enabling the characterization of transient states during the fast Cu–Aβ binding and interconversion processes.« less

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.

Spectroscopic and Theoretical Study of CuI Binding to His111 in the Human Prion Protein Fragment 106–115

PubMed Central

2016-01-01

The ability of the cellular prion protein (PrPC) to bind copper in vivo points to a physiological role for PrPC in copper transport. Six copper binding sites have been identified in the nonstructured N-terminal region of human PrPC. Among these sites, the His111 site is unique in that it contains a MKHM motif that would confer interesting CuI and CuII binding properties. We have evaluated CuI coordination to the PrP(106–115) fragment of the human PrP protein, using NMR and X-ray absorption spectroscopies and electronic structure calculations. We find that Met109 and Met112 play an important role in anchoring this metal ion. CuI coordination to His111 is pH-dependent: at pH >8, 2N1O1S species are formed with one Met ligand; in the range of pH 5–8, both methionine (Met) residues bind to CuI, forming a 1N1O2S species, where N is from His111 and O is from a backbone carbonyl or a water molecule; at pH <5, only the two Met residues remain coordinated. Thus, even upon drastic changes in the chemical environment, such as those occurring during endocytosis of PrPC (decreased pH and a reducing potential), the two Met residues in the MKHM motif enable PrPC to maintain the bound CuI ions, consistent with a copper transport function for this protein. We also find that the physiologically relevant CuI-1N1O2S species activates dioxygen via an inner-sphere mechanism, likely involving the formation of a copper(II) superoxide complex. In this process, the Met residues are partially oxidized to sulfoxide; this ability to scavenge superoxide may play a role in the proposed antioxidant properties of PrPC. This study provides further insight into the CuI coordination properties of His111 in human PrPC and the molecular mechanism of oxygen activation by this site. PMID:26930130

Copper attachment to a non-octarepeat site in prion protein

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Bernholc, Jerry

2010-03-01

Prion protein, PrP, plays a causative role in several neurodegenerative diseases, including mad cow disease in cattle and Creutzfeldt-Jakob disease in humans. The PrP is known to efficiently bind copper ions and this ability has been linked to its function. PrP contains up to six binding sites, four of which are located in the so-called octarepeat region and are now well known. The binding sites outside this region are still largely undetermined, despite evidence of their relevance to prion diseases. Using a hybrid DFT/DFT, which combines Kohn-Sham DFT with orbital-free DFT to achieve accurate and efficient description of solvent effects in ab initio calculations, we have investigated copper attachment to the sequence GGGTH, which represents the copper binding site located at His96. We have considered both NNNN and NNNO types of copper coordination, as suggested by experiments. Our calculations have determined the geometry of copper attachment site and its energetics. Comparison to the already known binding sites provides insight into the process of copper uptake in PrP.

Subnanomolar indazole-5-carboxamide inhibitors of monoamine oxidase B (MAO-B) continued: indications of iron binding, experimental evidence for optimised solubility and brain penetration.

PubMed

Tzvetkov, Nikolay T; Antonov, Liudmil

2017-12-01

Pharmacological and physicochemical studies of N-unsubstituted indazole-5-carboxamides (subclass I) and their structurally optimised N1-methylated analogues (subclass II), initially developed as drug and radioligand candidates for the treatment and diagnosis of Parkinson's disease (PD), are presented. The compounds are highly brain permeable, selective, reversible, and competitive monoamine oxidase B (MAO-B) inhibitors with improved water-solubility and subnanomolar potency (pIC 50  >8.8). Using a well-validated, combined X-ray/modelling technology platform, we performed a semi-quantitative analysis of the binding modes of all compounds and investigated the role of the indazole N1 position for their MAO-B inhibitory activity. Moreover, compounds NTZ-1006, 1032, and 1441 were investigated for their ability to bind Fe 2+ and Fe 3+ ions using UV-visible spectroscopy.

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

Ren, Hao; Zhang, Yu; Guo, Sibei

The aggregation of amyloid beta (Aβ) peptides plays a crucial role in the pathology and etiology of Alzheimer's disease. Experimental evidence shows that copper ion is an aggregation-prone species with the ability to coordinately bind to Aβ and further induce the formation of neurotoxic Aβ oligomers. However, the detailed structures of Cu(II)–Aβ complexes have not been illustrated, and the kinetics and dynamics of the Cu(II) binding are not well understood. Two Cu(II)–Aβ complexes have been proposed to exist under physiological conditions, and another two might exist at higher pH values. By using ab initio simulations for the spontaneous resonance Ramanmore » and time domain stimulated resonance Raman spectroscopy signals, we obtained the characteristic Raman vibronic features of each complex. Finally, these signals contain rich structural information with high temporal resolution, enabling the characterization of transient states during the fast Cu–Aβ binding and interconversion processes.« less

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

Location of Bromide Ions in Tetragonal Lysozyme Crystals

NASA Technical Reports Server (NTRS)

Lim, Kap; Nadarajah, Arunan; Forsythe, Elizabeth L.; Pusey, Marc L.

1998-01-01

Anions have been shown to play a dominant role in the crystallization of chicken egg white lysozyme from salt solutions. Previous studies employing X-ray crystallography had found one chloride ion binding site in the tetragonal crystal form of the protein and four nitrate ion binding sites in the monoclinic form. In this study the anion positions in the tetragonal form were determined from the difference Fourier map obtained from lysozyme crystal grown in bromide and chloride solutions. Five possible anion binding sites were found in this manner. Some of these sites were in pockets containing basic residues while others were near neutral, but polar, residues. The sole chloride ion binding site found in previous studies was confirmed, while four of these sites corresponded to four binding sites found for nitrate ions in monoclinic crystals. The study suggests that most of the anion binding sites in lysozyme remain unchanged, even when different anions and different crystal forms of lysozyme are employed.

Locations of Bromide Ions in Tetragonal Lysozyme Crystals

NASA Technical Reports Server (NTRS)

Lim, Kap; Nadarajah, Arunan; Forsythe, Elizabeth L.; Pusey, Marc L.

1998-01-01

Anions have been shown to play a dominant role in the crystallization of chicken egg-white lysozyme from salt solutions. Previous studies employing X-ray crystallography have found one chloride ion binding site in the tetragonal crystal form of the protein and four nitrate ion binding sites in the monoclinic form. In this study the anion positions in the tetragonal form were determined from the difference Fourier map obtained from lysozyme crystals grown in bromide and chloride solutions. Five possible anion-binding sites were found in this manner. Some of these sites were in pockets containing basic residues while others were near neutral, but polar, residues. The sole chloride ion binding site found in previous studies was confirmed, while four further sites were found which corresponded to the four binding sites found for nitrate ions in monoclinic crystals. The study suggests that most of the anion-binding sites in lysozyme remain unchanged even when different anions and different crystal forms of lysozyme are employed.

An accurate cost effective DFT approach to study the sensing behaviour of polypyrrole towards nitrate ions in gas and aqueous phases.

PubMed

Wasim, Fatima; Mahmood, Tariq; Ayub, Khurshid

2016-07-28

Density functional theory (DFT) calculations have been performed to study the response of polypyrrole towards nitrate ions in gas and aqueous phases. First, an accurate estimate of interaction energies is obtained by methods calibrated against the gold standard CCSD(T) method. Then, a number of low cost DFT methods are also evaluated for their ability to accurately estimate the binding energies of polymer-nitrate complexes. The low cost methods evaluated here include dispersion corrected potential (DCP), Grimme's D3 correction, counterpoise correction of the B3LYP method, and Minnesota functionals (M05-2X). The interaction energies calculated using the counterpoise (CP) correction and DCP methods at the B3LYP level are in better agreement with the interaction energies calculated using the calibrated methods. The interaction energies of an infinite polymer (polypyrrole) with nitrate ions are calculated by a variety of low cost methods in order to find the associated errors. The electronic and spectroscopic properties of polypyrrole oligomers nPy (where n = 1-9) and nPy-NO3(-) complexes are calculated, and then extrapolated for an infinite polymer through a second degree polynomial fit. Charge analysis, frontier molecular orbital (FMO) analysis and density of state studies also reveal the sensing ability of polypyrrole towards nitrate ions. Interaction energies, charge analysis and density of states analyses illustrate that the response of polypyrrole towards nitrate ions is considerably reduced in the aqueous medium (compared to the gas phase).

Structures of the carbohydrate recognition domain of Ca2+-independent cargo receptors Emp46p and Emp47p.

PubMed

Satoh, Tadashi; Sato, Ken; Kanoh, Akira; Yamashita, Katsuko; Yamada, Yusuke; Igarashi, Noriyuki; Kato, Ryuichi; Nakano, Akihiko; Wakatsuki, Soichi

2006-04-14

Emp46p and Emp47p are type I membrane proteins, which cycle between the endoplasmic reticulum (ER) and the Golgi apparatus by vesicles coated with coat protein complexes I and II (COPI and COPII). They are considered to function as cargo receptors for exporting N-linked glycoproteins from the ER. We have determined crystal structures of the carbohydrate recognition domains (CRDs) of Emp46p and Emp47p of Saccharomyces cerevisiae, in the absence and presence of metal ions. Both proteins fold as a beta-sandwich, and resemble that of the mammalian ortholog, p58/ERGIC-53. However, the nature of metal binding is distinct from that of Ca(2+)-dependent p58/ERGIC-53. Interestingly, the CRD of Emp46p does not bind Ca(2+) ion but instead binds K(+) ion at the edge of a concave beta-sheet whose position is distinct from the corresponding site of the Ca(2+) ion in p58/ERGIC-53. Binding of K(+) ion to Emp46p appears essential for transport of a subset of glycoproteins because the Y131F mutant of Emp46p, which cannot bind K(+) ion fails to rescue the transport in disruptants of EMP46 and EMP47 genes. In contrast the CRD of Emp47p binds no metal ions at all. Furthermore, the CRD of Emp46p binds to glycoproteins carrying high mannosetype glycans and the is promoted by binding not the addition of Ca(2+) or K(+) ion in These results suggest that Emp46p can be regarded as a Ca(2+)-independent intracellular lectin at the ER exit sites.

Molecular mechanism of ATP binding and ion channel activation in P2X receptors

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

Hattori, Motoyuki; Gouaux, Eric

P2X receptors are trimeric ATP-activated ion channels permeable to Na{sup +}, K{sup +} and Ca{sup 2+}. The seven P2X receptor subtypes are implicated in physiological processes that include modulation of synaptic transmission, contraction of smooth muscle, secretion of chemical transmitters and regulation of immune responses. Despite the importance of P2X receptors in cellular physiology, the three-dimensional composition of the ATP-binding site, the structural mechanism of ATP-dependent ion channel gating and the architecture of the open ion channel pore are unknown. Here we report the crystal structure of the zebrafish P2X4 receptor in complex with ATP and a new structure ofmore » the apo receptor. The agonist-bound structure reveals a previously unseen ATP-binding motif and an open ion channel pore. ATP binding induces cleft closure of the nucleotide-binding pocket, flexing of the lower body {beta}-sheet and a radial expansion of the extracellular vestibule. The structural widening of the extracellular vestibule is directly coupled to the opening of the ion channel pore by way of an iris-like expansion of the transmembrane helices. The structural delineation of the ATP-binding site and the ion channel pore, together with the conformational changes associated with ion channel gating, will stimulate development of new pharmacological agents.« less

Graphane versus graphene: a computational investigation of the interaction of nucleobases, aminoacids, heterocycles, small molecules (CO2, H2O, NH3, CH4, H2), metal ions and onium ions.

PubMed

Umadevi, Deivasigamani; Narahari Sastry, G

2015-11-11

Graphane has emerged as a two-dimensional hydrocarbon with interesting physical properties and potential applications. Understanding the interaction of graphane with various molecules and ions is crucial to appreciate its potential applications. We investigated the interaction of nucleobases, aminoacids, saturated and unsaturated heterocycles, small molecules, metal ions and onium ions with graphane by using density functional theory calculations. The preferred orientations of these molecules and ions on the graphane surface have been analysed. The binding energies of graphane with these molecules have been compared with the corresponding binding energies of graphene. Our results reveal that graphane forms stable complexes with all the molecules and ions yet showing lesser binding affinity when compared to graphene. As an exemption, the preferential strong binding of H2O with graphane than graphene reveals the fact that graphane is more hydrophilic than graphene. Charge transfer between graphane and the molecules and ions have been found to be an important factor in determining the binding strength of the complexes. The effect of the interaction of these molecules and ions on the HOMO-LUMO energy gap of graphane has also been investigated.

Influences of Mutations on the Electrostatic Binding Free Energies of Chloride Ions in Escherichia Coli ClC

PubMed Central

Yu, Tao; Wang, Xiao-Qing; Sang, Jian-Ping; Pan, Chun-Xu; Zou, Xian-Wu; Chen, Tsung-Yu; Zou, Xiaoqin

2012-01-01

Mutations in ClC channel proteins may cause serious functional changes and even diseases. The function of ClC proteins mainly manifests as Cl− transport, which is related to the binding free energies of chloride ions. Therefore, the influence of a mutation on ClC function can be studied by investigating the mutational effect on the binding free energies of chloride ions. The present study provides quantitative and systematic investigations on the influences of residue mutations on the electrostatic binding free energies in Escherichia coli ClC (EcClC) proteins, using all-atom molecular dynamics simulations. It was found that the change of the electrostatic binding free energy decreases linearly with the increase of the residue-chloride ion distance for a mutation. This work reveals how changes in the charge of a mutated residue and in the distance between the mutated residue and the binding site govern the variations in the electrostatic binding free energies, and therefore influence the transport of chloride ions and conduction in EcClC. This work would facilitate our understanding of the mutational effects on transport of chloride ions and functions of ClC proteins, and provide a guideline to estimate which residue mutations will have great influences on ClC functions. PMID:22612693

Metal ion reactive thin films using spray electrostatic LbL assembly.

PubMed

Krogman, Kevin C; Lyon, Katharine F; Hammond, Paula T

2008-11-20

By using the spray-layer-by-layer (Spray-LbL) technique, the number of metal counterions trapped within LbL coatings is significantly increased by kinetically freezing the film short of equilibrium, potentially limiting interchain penetration and forcing chains to remain extrinsically compensated to a much greater degree than observed in the traditional dipped LbL technique. The basis for the enhanced entrapment of metal ions such as Cu2+, Fe2+, and Ag+ is addressed, including the equilibrium driving force for extrinsic compensation by soft versus hard metal ions and the impact of Spray-LbL on the kinetics of polymer-ion complexation. These polymer-bound metal-ion coatings are also demonstrated to be effective treatments for air filtration, functionalizing existing filters with the ability to strongly bind toxic industrial compounds such as ammonia or cyanide gases, as well as chemical warfare agent simulants such as chloroethyl ethyl sulfide. On the basis of results reported here, future work could extend this method to include other toxic soft-base ligands such as carbon monoxide, benzene, or organophosphate nerve agents.

Tris-amidoximate uranyl complexes via η2 binding mode coordinated in aqueous solution shown by X-ray absorption spectroscopy and density functional theory methods.

PubMed

Zhang, Linjuan; Qie, Meiying; Su, Jing; Zhang, Shuo; Zhou, Jing; Li, Jiong; Wang, Yu; Yang, Shitong; Wang, Shuao; Li, Jingye; Wu, Guozhong; Wang, Jian Qiang

2018-03-01

The present study sheds some light on the long-standing debate concerning the coordination properties between uranyl ions and the amidoxime ligand, which is a key ingredient for achieving efficient extraction of uranium. Using X-ray absorption fine structure combined with theoretical simulation methods, the binding mode and bonding nature of a uranyl-amidoxime complex in aqueous solution were determined for the first time. The results show that in a highly concentrated amidoxime solution the preferred binding mode between UO 2 2+ and the amidoxime ligand is η 2 coordination with tris-amidoximate species. In such a uranyl-amidoximate complex with η 2 binding motif, strong covalent interaction and orbital hybridization between U 5f/6d and (N, O) 2p should be responsible for the excellent binding ability of the amidoximate ligand to uranyl. The study was performed directly in aqueous solution to avoid the possible binding mode differences caused by crystallization of a single-crystal sample. This work also is an example of the simultaneous study of local structure and electronic structure in solution systems using combined diagnostic tools.

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

Laskin, Julia; Johnson, Grant E.; Prabhakaran, Venkateshkumar

Immobilization of complex molecules and clusters on supports plays an important role in a variety of disciplines including materials science, catalysis and biochemistry. In particular, deposition of clusters on surfaces has attracted considerable attention due to their non-scalable, highly size-dependent properties. The ability to precisely control the composition and morphology of clusters and small nanoparticles on surfaces is crucial for the development of next generation materials with rationally tailored properties. Soft- and reactive landing of ions onto solid or liquid surfaces introduces unprecedented selectivity into surface modification by completely eliminating the effect of solvent and sample contamination on the qualitymore » of the film. The ability to select the mass-to-charge ratio of the precursor ion, its kinetic energy and charge state along with precise control of the size, shape and position of the ion beam on the deposition target makes soft-landing an attractive approach for surface modification. High-purity uniform thin films on surfaces generated using mass-selected ion deposition facilitate understanding of critical interfacial phenomena relevant to catalysis, energy generation and storage, and materials science. Our efforts have been directed toward understanding charge retention by soft-landed metal and metal-oxide cluster ions, which may affect both their structure and reactivity. Specifically, we have examined the effect of the surface on charge retention by both positively and negatively charged cluster ions. We found that the electronic properties of the surface play an important role in charge retention by cluster cations. Meanwhile, the electron binding energy is a key factor determining charge retention by cluster anions. These findings provide the scientific foundation for the rational design of interfaces for advanced catalysts and energy storage devices. Further optimization of electrode-electrolyte interfaces for applications in energy storage and electrocatalysis may be achieved by understanding and controlling the properties of soft-landed cluster ions.« less

Free Energy Wells and Barriers to Ion Transport Across Membranes

NASA Astrophysics Data System (ADS)

Rempe, Susan

2014-03-01

The flow of ions across cellular membranes is essential to many biological processes. Ion transport is also important in synthetic materials used as battery electrolytes. Transport often involves specific ions and fast conduction. To achieve those properties, ion conduction pathways must solvate specific ions by just the ``right amount.'' The right amount of solvation avoids ion traps due to deep free energy wells, and avoids ion block due to high free energy barriers. Ion channel proteins in cellular membranes demonstrate this subtle balance in solvation of specific ions. Using ab initio molecular simulations, we have interrogated the link between binding site structure and ion solvation free energies in biological ion binding sites. Our results emphasize the surprisingly important role of the environment that surrounds ion-binding sites for fast transport of specific ions. We acknowledge support from Sandia's LDRD program. Sandia National Labs is a multi-program laboratory operated by Sandia Corp., a wholly owned subsidiary of Lockheed Martin Corp., for the US DOE's NNSA under contract DE-AC04-94AL85000.

Tuning the ion selectivity of tetrameric cation channels by changing the number of ion binding sites

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

Derebe, Mehabaw G.; Sauer, David B.; Zeng, Weizhong

2015-11-30

Selective ion conduction across ion channel pores is central to cellular physiology. To understand the underlying principles of ion selectivity in tetrameric cation channels, we engineered a set of cation channel pores based on the nonselective NaK channel and determined their structures to high resolution. These structures showcase an ensemble of selectivity filters with a various number of contiguous ion binding sites ranging from 2 to 4, with each individual site maintaining a geometry and ligand environment virtually identical to that of equivalent sites in K{sup +} channel selectivity filters. Combined with single channel electrophysiology, we show that only themore » channel with four ion binding sites is K{sup +} selective, whereas those with two or three are nonselective and permeate Na{sup +} and K{sup +} equally well. These observations strongly suggest that the number of contiguous ion binding sites in a single file is the key determinant of the channel's selectivity properties and the presence of four sites in K{sup +} channels is essential for highly selective and efficient permeation of K{sup +} ions.« less

Mg(II) and Ni(II) induce aggregation of poly(rA)poly(rU) to either tetra-aggregate or triplex depending on the metal ion concentration.

PubMed

Biver, Tarita; Busto, Natalia; García, Begoña; Leal, José M; Menichetti, Luisa; Secco, Fernando; Venturini, Marcella

2015-10-01

The ability of magnesium(II) and nickel(II) to induce dramatic conformational changes in the synthetic RNA poly(rA)poly(rU) has been investigated. Kinetic experiments, spectrofluorometric titrations, melting experiments and DSC measurements contribute in shedding light on a complex behaviour where the action of metal ions (Na(+), Mg(2+), Ni(2+)), in synergism with other operators as the intercalating dye coralyne and temperature, all concur in stabilising a peculiar RNA form. Mg(2+) and Ni(2+) (M) bind rapidly and almost quantitatively to the duplex (AU) to give a RNA/metal ion complex (AUM). Then, by the union of two AUM units, an unstable tetra-aggregate (UAUA(M2)*) is formed which, in the presence of a relatively modest excess of metal, evolves to the UAUM triplex by releasing a single AM strand. On the other hand, under conditions of high metal content, the UAUA(M2)* intermediate rearranges to give a more stable tetra-aggregate (UAUA(M2)). As concerns the role of coralyne (D), it is found that D strongly interacts with UAUA(M2). Also, in the presence of coralyne, the ability of divalent ions to promote the transition of AUD into UAUD is enhanced, according to the efficiency sequence [Ni(2+)]≫[Mg(2+)]≫[Na(+)]. Copyright © 2015 Elsevier Inc. All rights reserved.

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.

Structure and evolutionary aspects of matrix metalloproteinases: a brief overview.

PubMed

Das, Sudip; Mandal, Malay; Chakraborti, Tapati; Mandal, Amritlal; Chakraborti, Sajal

2003-11-01

The matrix metalloproteinases (MMPs) are zinc dependent endopeptidases known for their ability to cleave one or several extracellular matrix (ECM) constituents, as well as non-matrix proteins. They comprise a large family of proteinases that share common structural and functional elements and are products of different genes. All members of this family contain a signal peptide, a propeptide and a catalytic domain. The catalytic domain contains two zinc ions and at least one calcium ion coordinated to various residues. All MMPs, with the exception matrilysin, have a hemopexin/vitronectin-like domain that is connected to the catalytic domain by a hinge or linker region. The hemopexin-like domain influences tissue inhibitor of metalloproteinases (TIMP) binding, the binding of certain substrates, membrane activation, and some proteolytic activities. It has been proposed that the origin of MMPs could be traced to before the emergence of vertebrates from invertebrates. It appears conceivable that the domain assemblies occurred at an early stage of the diversification of different MMPs and that they progressed through the evolutionary process independent of one another, and perhaps parallel to each other.

A dansyl-rhodamine chemosensor for Fe(III) based on off-on FRET.

PubMed

Piao, Jingyu; Lv, Jia; Zhou, Xin; Zhao, Tong; Wu, Xue

2014-07-15

A novel fluorescent chemosensor bearing a rhodamine and a dansyl moiety was developed for highly selective detection of Fe(3+) based on fluorescence resonance energy transfer (FRET) mechanism. Binding of Fe(3+) to the chemosensor induced spirolactam ring opening in the rhodamine moiety and subsequent off-on FRET from the dansyl energy donor to the rhodamine energy acceptor due to the spectral overlap between the emission of the dansyl moiety and the absorption of the ring opened rhodamine moiety. Job's plot analysis indicated a 1:1 binding stoichiometry between the chemosensor and Fe(3+). The association constant was estimated to be 2.72×10(3) M(-1) according to the Benesi-Hildebrand method. With the feature of easy synthesis, simple structural skeleton and excellent sensing ability, the newly synthesized chemosensor provided the potential for applying as a highly selective fluorescent probe in complex samples containing various competitive metal ions and developing other metal ion chemosensors to fulfill various needs of biological and environmental field. Copyright © 2014 Elsevier B.V. All rights reserved.

IBiSA_Tools: A Computational Toolkit for Ion-Binding State Analysis in Molecular Dynamics Trajectories of Ion Channels.

PubMed

Kasahara, Kota; Kinoshita, Kengo

2016-01-01

Ion conduction mechanisms of ion channels are a long-standing conundrum. Although the molecular dynamics (MD) method has been extensively used to simulate ion conduction dynamics at the atomic level, analysis and interpretation of MD results are not straightforward due to complexity of the dynamics. In our previous reports, we proposed an analytical method called ion-binding state analysis to scrutinize and summarize ion conduction mechanisms by taking advantage of a variety of analytical protocols, e.g., the complex network analysis, sequence alignment, and hierarchical clustering. This approach effectively revealed the ion conduction mechanisms and their dependence on the conditions, i.e., ion concentration and membrane voltage. Here, we present an easy-to-use computational toolkit for ion-binding state analysis, called IBiSA_tools. This toolkit consists of a C++ program and a series of Python and R scripts. From the trajectory file of MD simulations and a structure file, users can generate several images and statistics of ion conduction processes. A complex network named ion-binding state graph is generated in a standard graph format (graph modeling language; GML), which can be visualized by standard network analyzers such as Cytoscape. As a tutorial, a trajectory of a 50 ns MD simulation of the Kv1.2 channel is also distributed with the toolkit. Users can trace the entire process of ion-binding state analysis step by step. The novel method for analysis of ion conduction mechanisms of ion channels can be easily used by means of IBiSA_tools. This software is distributed under an open source license at the following URL: http://www.ritsumei.ac.jp/~ktkshr/ibisa_tools/.

Nucleoside-2',3'/3',5'-bis(thio)phosphate antioxidants are also capable of disassembly of amyloid beta42-Zn(ii)/Cu(ii) aggregates via Zn(ii)/Cu(ii)-chelation.

PubMed

Hevroni, Bosmat Levi; Major, Dan Thomas; Dixit, Mudit; Mhashal, Anil Ranu; Das, Susanta; Fischer, Bilha

2016-05-18

Currently, there is an urgent need for biocompatible metal-ion chelators capable of antioxidant activity and disassembly of amyloid beta (Aβ)-aggregates as potential therapeutics for Alzheimer's disease (AD). We recently demonstrated the promising antioxidant activity of adenine/guanine 2',3' or 3',5'-bis(thio)phosphate analogues, 2'-dA/G3'5'PO/S and A2'3'PO/S, and their affinity to Zn(ii)-ions. These findings encouraged us to evaluate them as agents for the dissolution of Aβ42-Zn(ii)/Cu(ii) aggregates. Specifically, we explored their ability to bind Cu(ii)/Zn(ii)-ions, the geometry and stoichiometry of these complexes, Cu(ii)/Zn(ii)-binding-sites and binding mode, and the ability of these analogues to dissolve Aβ42-Zn(ii)/Cu(ii) aggregates, as well as their effect on the secondary structure of those aggregates. Finally, we identified the most promising agents for dissolution of Aβ42-Zn(ii)/Cu(ii) aggregates. Specifically, we observed the formation of a 1 : 1 complex between 2'-dG3'5'PO and Cu(ii), involving O4 ligands. Zn(ii) was coordinated by both thiophosphate groups of 2'-dA3'5'PS and A2'3'PS involving O2S2 ligands in a 1 : 1 stoichiometry. A2'3'PS dissolves Aβ42-Zn(ii) and Aβ42-Cu(ii) aggregates as effectively as, and 2.5-fold more effectively than EDTA, respectively. Furthermore, 2'-dG3'5'PS and A2'3'PS reverted the Aβ42-M(ii) structure, back to that of the free Aβ42. Finally, cryo-TEM and TEM images confirmed the disassembly of Aβ42 and Aβ42-M(ii) aggregates by A2'3'PS. Hence, 2'-dG3'5'PS and A2'3'PS may serve as promising scaffolds for new AD therapeutics, acting as both effective antioxidants and agents for solubilization of Aβ42-Cu(ii)/Zn(ii) aggregates.

Multi-shell model of ion-induced nucleic acid condensation

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

Tolokh, Igor S.; Drozdetski, Aleksander V.; Pollack, Lois

We present a semi-quantitative model of condensation of short nucleic acid (NA) duplexes induced by trivalent cobalt(III) hexammine (CoHex) ions. The model is based on partitioning of bound counterion distribution around single NA duplex into “external” and “internal” ion binding shells distinguished by the proximity to duplex helical axis. In the aggregated phase the shells overlap, which leads to significantly increased attraction of CoHex ions in these overlaps with the neighboring duplexes. The duplex aggregation free energy is decomposed into attractive and repulsive components in such a way that they can be represented by simple analytical expressions with parameters derivedmore » from molecular dynamic simulations and numerical solutions of Poisson equation. The attractive term depends on the fractions of bound ions in the overlapping shells and affinity of CoHex to the “external” shell of nearly neutralized duplex. The repulsive components of the free energy are duplex configurational entropy loss upon the aggregation and the electrostatic repulsion of the duplexes that remains after neutralization by bound CoHex ions. The estimates of the aggregation free energy are consistent with the experimental range of NA duplex condensation propensities, including the unusually poor condensation of RNA structures and subtle sequence effects upon DNA condensation. The model predicts that, in contrast to DNA, RNA duplexes may condense into tighter packed aggregates with a higher degree of duplex neutralization. An appreciable CoHex mediated RNA-RNA attraction requires closer inter-duplex separation to engage CoHex ions (bound mostly in the “internal” shell of RNA) into short-range attractive interactions. The model also predicts that longer NA fragments will condense more readily than shorter ones. The ability of this model to explain experimentally observed trends in NA condensation lends support to proposed NA condensation picture based on the multivalent “ion binding shells.”.« less

Probing Interactions at the Nanoscale by Ion Current through Nanopores and Nanovoids

NASA Astrophysics Data System (ADS)

Gamble, Trevor Patrick

Polymer nanopores offer themselves as excellent test beds for study of phenomena that occur on the nano-scale, such as Debye layer formation, surface charge modulation, current saturation, and rectification. Studying ions interactions within the Debye layer, for example, is not possible on the micro-scale, where the pore diameter can be 100 times the size of the zone where interactions of interest occur. However, in our nanopores with an opening diameter less than 10 nm, a slight change of the Debye length can lead to drastic changes of the recorded ion current. Here we present our nanopores' use as a tool to study geometrical and electrochemical properties of porous manganese oxide. There is great value in studying nano-scale properties of this material because of its importance in lithium ion batteries and newly developed nano-architectures within supercapacitors. We electrodeposited manganese oxide wires into our cylindrical nanopores, filling them completely. In this use, nanopores became a template to probe properties of the embedded material such as surface charge, ion selectivity, and porosity. This information was then reported to the Energy Frontier Research Center (EFRC) collaboration, so that other groups can incorporate these recently discovered characteristics into future their nano-architecture design. Additionally, we constructed conical nanopores to study interactions between the surface charges found on the walls and alkali metal ions. In particular we looked at lithium, as it is the electrochemically active ion during charge cycling in EFRC energy storage devices. We attempted to reveal lithium ion's affinity to bind to surface charges. We found this binding led to lowering of the effective surface charge of the pore walls, while also decreasing lithium's ability to move through channels or voids that have charged walls. In connection to manganese oxide, a porous, charged material with voids, information on lithium's interaction with these charges is paramount.

Multi-shell model of ion-induced nucleic acid condensation

PubMed Central

Tolokh, Igor S.; Drozdetski, Aleksander V.; Pollack, Lois; Onufriev, Alexey V.

2016-01-01

We present a semi-quantitative model of condensation of short nucleic acid (NA) duplexes induced by trivalent cobalt(iii) hexammine (CoHex) ions. The model is based on partitioning of bound counterion distribution around single NA duplex into “external” and “internal” ion binding shells distinguished by the proximity to duplex helical axis. In the aggregated phase the shells overlap, which leads to significantly increased attraction of CoHex ions in these overlaps with the neighboring duplexes. The duplex aggregation free energy is decomposed into attractive and repulsive components in such a way that they can be represented by simple analytical expressions with parameters derived from molecular dynamic simulations and numerical solutions of Poisson equation. The attractive term depends on the fractions of bound ions in the overlapping shells and affinity of CoHex to the “external” shell of nearly neutralized duplex. The repulsive components of the free energy are duplex configurational entropy loss upon the aggregation and the electrostatic repulsion of the duplexes that remains after neutralization by bound CoHex ions. The estimates of the aggregation free energy are consistent with the experimental range of NA duplex condensation propensities, including the unusually poor condensation of RNA structures and subtle sequence effects upon DNA condensation. The model predicts that, in contrast to DNA, RNA duplexes may condense into tighter packed aggregates with a higher degree of duplex neutralization. An appreciable CoHex mediated RNA-RNA attraction requires closer inter-duplex separation to engage CoHex ions (bound mostly in the “internal” shell of RNA) into short-range attractive interactions. The model also predicts that longer NA fragments will condense more readily than shorter ones. The ability of this model to explain experimentally observed trends in NA condensation lends support to proposed NA condensation picture based on the multivalent “ion binding shells.” PMID:27389241

Effects of ionic compositions of the medium on monosodium glutamate binding to taste epithelial cells.

PubMed

Hayashi, Y; Tsunenari, T; Mori, T

1999-03-01

Monosodium glutamate and nucleotides are umami taste substances in animals and have a synergistic effect on each other. We studied the ligand-binding properties of the glutamate receptors in taste epithelial cells isolated from bovine tongue. Specific glutamate binding was observed in an enriched suspension of taste receptor cells in Hanks' balanced salt solution, while no specific glutamate binding was apparent in the absence of divalent ions or when the cells had been depolarized by a high content of potassium in Hanks' balanced salt solution. There was no significant difference between the release of glutamate under depolarized or divalent ion-free conditions and under normal conditions. However, glutamate was easily released from the depolarized cells in the absence of divalent ions. These data suggest that the binding of glutamate to receptors depends on divalent ions, which also have an effect on maintaining binding between glutamate and receptors.

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.

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.

Hemin/G-quadruplex structure and activity alteration induced by magnesium cations.

PubMed

Kosman, J; Juskowiak, B

2016-04-01

The influence of metal cations on G-quadruplex structure and peroxidase-mimicking DNAzyme activity was investigated. Experiments revealed a significant role of magnesium ion, which in the presence of potassium cation influenced DNAzyme activity. This ability has been associated with alteration of G-quadruplex topology and consequently affinity to bind hemin molecule. It has been demonstrated that G-quadruplex based on PS2.M sequence under these conditions formed parallel topology, which exhibited lower activity than that observed in standard potassium-containing solution. On the other hand DNAzyme/magnesium ion system based on telomeric sequence, which did not undergo significant structural changes, exhibited higher peroxidase activity upon magnesium ion addition. In both cases, the stabilization effect of magnesium cations on G-quadruplex structure was observed. The mechanism of DNAzyme activity alteration by magnesium ion can be explained by its influence on the pKa value of DNAzyme. Magnesium ion decreased pKa for PS2.M based system but increased it for telomeric DNAzyme. Magnesium cation effect on G-quadruplex structure as well as DNAzyme activity is particularly important since this ion is one of the most common metal cations in biological samples. Copyright © 2016 Elsevier B.V. All rights reserved.

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.

Yeast enolase: mechanism of activation by metal ions.

PubMed

Brewer, J M

1981-01-01

Yeast enolase as prepared by current procedures is inherently chemically homogeneous, though deamidation and partial denaturation can produce electrophoretically distinct forms. A true isozyme of the enzyme exists but does not survive the purification procedure. The chemical sequence for both has been established. The enzyme behaves in solution like a compact, nearly spherical molecule of moderate hydration. Strong intramolecular forces maintain the structure of the individual subunits. The enzyme as isolated is dimeric. If dissociated in the presence of magnesium ions and substrate, then the subunits are active, but if the dissociation occurs in the absence of metal ions, they are inactive until they have reassociated and undergone a first order "annealing" process. Magnesium (II) enhances association. The interaction between the subunits is hydrophobic in character. The enzyme can bind up to 2 mol of most metal ions in "conformational" sites which then allows up to 2 mol of substrate or some substrate analogue to bind. This is not sufficient for catalysis, but conformational metal ions do more than just allow substrate binding. A change in the environment of the metal ions occurs on substrate or substrate analogue binding. There is an absolute correlation between the occurrence of a structural change undergone by the 3-amino analogue of phosphoenolpyruvate and whether the metal ions produce any level of enzymatic activity. For catalysis, two more moles of metal ions, called "catalytic", must bind. There is evidence that the enzymatic reaction involves a carbanion mechanism. It is likely that two more moles of metal ion can bind which inhibit the reaction. The requirement for 2 mol of metal ion per subunit which contribute in different ways to catalysis is exhibited by a number of other enzymes.

The crystal structures of the psychrophilic subtilisin S41 and the mesophilic subtilisin Sph reveal the same calcium-loaded state.

PubMed

Almog, Orna; González, Ana; Godin, Noa; de Leeuw, Marina; Mekel, Marlene J; Klein, Daniela; Braun, Sergei; Shoham, Gil; Walter, Richard L

2009-02-01

We determine and compare the crystal structure of two proteases belonging to the subtilisin superfamily: S41, a cold-adapted serine protease produced by Antarctic bacilli, at 1.4 A resolution and Sph, a mesophilic serine protease produced by Bacillus sphaericus, at 0.8 A resolution. The purpose of this comparison was to find out whether multiple calcium ion binding is a molecular factor responsible for the adaptation of S41 to extreme low temperatures. We find that these two subtilisins have the same subtilisin fold with a root mean square between the two structures of 0.54 A. The final models for S41 and Sph include a calcium-loaded state of five ions bound to each of these two subtilisin molecules. None of these calcium-binding sites correlate with the high affinity known binding site (site A) found for other subtilisins. Structural analysis of the five calcium-binding sites found in these two crystal structures indicate that three of the binding sites have two side chains of an acidic residue coordinating the calcium ion, whereas the other two binding sites have either a main-chain carbonyl, or only one acidic residue side chain coordinating the calcium ion. Thus, we conclude that three of the sites are of high affinity toward calcium ions, whereas the other two are of low affinity. Because Sph is a mesophilic subtilisin and S41 is a psychrophilic subtilisin, but both crystal structures were found to bind five calcium ions, we suggest that multiple calcium ion binding is not responsible for the adaptation of S41 to low temperatures. Copyright 2008 Wiley-Liss, Inc.

A differential scanning calorimetric study of the effects of metal ions, substrate/product, substrate analogues and chaotropic anions on the thermal denaturation of yeast enolase 1.

PubMed

Brewer, J M; Wampler, J E

2001-03-14

The thermal denaturation of yeast enolase 1 was studied by differential scanning calorimetry (DSC) under conditions of subunit association/dissociation, enzymatic activity or substrate binding without turnover and substrate analogue binding. Subunit association stabilizes the enzyme, that is, the enzyme dissociates before denaturing. The conformational change produced by conformational metal ion binding increases thermal stability by reducing subunit dissociation. 'Substrate' or analogue binding additionally stabilizes the enzyme, irrespective of whether turnover is occurring, perhaps in part by the same mechanism. More strongly bound metal ions also stabilize the enzyme more, which we interpret as consistent with metal ion loss before denaturation, though possibly the denaturation pathway is different in the absence of metal ion. We suggest that some of the stabilization by 'substrate' and analogue binding is owing to the closure of moveable polypeptide loops about the active site, producing a more 'closed' and hence thermostable conformation.

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).

Long wave fluorophore sensor compounds and other fluorescent sensor compounds in polymers

DOEpatents

Walsh, Joseph C.; Heiss, Aaron M.; Noronha, Glenn; Vachon, David J.; Lane, Stephen M.; Satcher, Jr., Joe H.; Peyser, Thomas A.; Van Antwerp, William Peter; Mastrototaro, John Joseph

2004-07-20

Fluorescent biosensor molecules, fluorescent biosensors and systems, as well as methods of making and using these biosensor molecules and systems are described. Embodiments of these biosensor molecules exhibit fluorescence emission at wavelengths greater than about 650 nm. Typical biosensor molecules include a fluorophore that includes an iminium ion, a linker moiety that includes a group that is an anilinic type of relationship to the fluorophore and a boronate substrate recognition/binding moiety, which binds glucose. The fluorescence molecules modulated by the presence or absence of polyhydroxylated analytes such as glucose. This property of these molecules of the invention, as well as their ability to emit fluorescent light at greater than about 650 nm, renders these biosensor molecules particularly well-suited for detecting and measuring in-vivo glucose concentrations.

Expression, purification, and DNA-binding activity of the Herbaspirillum seropedicae RecX protein.

PubMed

Galvão, Carolina W; Pedrosa, Fábio O; Souza, Emanuel M; Yates, M Geoffrey; Chubatsu, Leda S; Steffens, Maria Berenice R

2004-06-01

The Herbaspirillum seropedicae RecX protein participates in the SOS response: a process in which the RecA protein plays a central role. The RecX protein of the H. seropedicae, fused to a His-tag sequence (RecX His-tagged), was over-expressed in Escherichia coli and purified by metal-affinity chromatography to yield a highly purified and active protein. DNA band-shift assays showed that the RecX His-tagged protein bound to both circular and linear double-stranded DNA and also to circular single-stranded DNA. The apparent affinity of RecX for DNA decreased in the presence of Mg(2+) ions. The ability of RecX to bind DNA may be relevant to its function in the SOS response.

The structural role of the zinc ion can be dispensable in prokaryotic zinc-finger domains

PubMed Central

Baglivo, Ilaria; Russo, Luigi; Esposito, Sabrina; Malgieri, Gaetano; Renda, Mario; Salluzzo, Antonio; Di Blasio, Benedetto; Isernia, Carla; Fattorusso, Roberto; Pedone, Paolo V.

2009-01-01

The recent characterization of the prokaryotic Cys2His2 zinc-finger domain, identified in Ros protein from Agrobacterium tumefaciens, has demonstrated that, although possessing a similar zinc coordination sphere, this domain is structurally very different from its eukaryotic counterpart. A search in the databases has identified ≈300 homologues with a high sequence identity to the Ros protein, including the amino acids that form the extensive hydrophobic core in Ros. Surprisingly, the Cys2His2 zinc coordination sphere is generally poorly conserved in the Ros homologues, raising the question of whether the zinc ion is always preserved in these proteins. Here, we present a functional and structural study of a point mutant of Ros protein, Ros56–142C82D, in which the second coordinating cysteine is replaced by an aspartate, 5 previously-uncharacterized representative Ros homologues from Mesorhizobium loti, and 2 mutants of the homologues. Our results indicate that the prokaryotic zinc-finger domain, which in Ros protein tetrahedrally coordinates Zn(II) through the typical Cys2His2 coordination, in Ros homologues can either exploit a CysAspHis2 coordination sphere, previously never described in DNA binding zinc finger domains to our knowledge, or lose the metal, while still preserving the DNA-binding activity. We demonstrate that this class of prokaryotic zinc-finger domains is structurally very adaptable, and surprisingly single mutations can transform a zinc-binding domain into a nonzinc-binding domain and vice versa, without affecting the DNA-binding ability. In light of our findings an evolutionary link between the prokaryotic and eukaryotic zinc-finger domains, based on bacteria-to-eukaryota horizontal gene transfer, is discussed. PMID:19369210

Cooperative binding modes of Cu(II) in prion protein

NASA Astrophysics Data System (ADS)

Hodak, Miroslav; Chisnell, Robin; Lu, Wenchang; Bernholc, Jerry

2007-03-01

The misfolding of the prion protein, PrP, is responsible for a group of neurodegenerative diseases including mad cow disease and Creutzfeldt-Jakob disease. It is known that the PrP can efficiently bind copper ions; four high-affinity binding sites located in the octarepeat region of PrP are now well known. Recent experiments suggest that at low copper concentrations new binding modes, in which one copper ion is shared between two or more binding sites, are possible. Using our hybrid Thomas-Fermi/DFT computational scheme, which is well suited for simulations of biomolecules in solution, we investigate the geometries and energetics of two, three and four binding sites cooperatively binding one copper ion. These geometries are then used as inputs for classical molecular dynamics simulations. We find that copper binding affects the secondary structure of the PrP and that it stabilizes the unstructured (unfolded) part of the protein.

Aqueous alkali halide solutions: can osmotic coefficients be explained on the basis of the ionic sizes alone?

PubMed

Kalyuzhnyi, Yu V; Vlachy, Vojko; Dill, Ken A

2010-06-21

We use the AMSA, associative mean spherical theory of associative fluids, to study ion-ion interactions in explicit water. We model water molecules as hard spheres with four off-center square-well sites and ions as charged hard spheres with sticky sites that bind to water molecules or other ions. We consider alkali halide salts. The choice of model parameters is based on two premises: (i) The strength of the interaction between a monovalent ion and a water molecule is inversely proportional to the ionic (crystal) diameter sigma(i). Smaller ions bind to water more strongly than larger ions do, taking into account the asymmetry of the cation-water and anion-water interactions. (ii) The number of contacts an ion can make is proportional to sigma2(i). In short, small ions bind waters strongly, but only a few of them. Large ions bind waters weakly, but many of them. When both a monovalent cation and anion are large, it yields a small osmotic coefficient of the salt, since the water molecules avoid the space in between large ions. On the other hand, salts formed from one small and one large ion remain hydrated and their osmotic coefficient is high. The osmotic coefficients, calculated using this model in combination with the integral equation theory developed for associative fluids, follow the experimental trends, including the unusual behavior of caesium salts.

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.

DNA binding, antioxidant, cytotoxicity (MTT, lactate dehydrogenase, NO), and cellular uptake studies of structurally different nickel(II) thiosemicarbazone complexes: synthesis, spectroscopy, electrochemistry, and X-ray crystallography.

PubMed

Prabhakaran, R; Kalaivani, P; Huang, R; Poornima, P; Vijaya Padma, V; Dallemer, F; Natarajan, K

2013-02-01

Three new nickel(II) thiosemicarbazone complexes have been synthesized and characterized by analytical, spectral, and single-crystal X-ray diffraction studies. In complex 1, the ligand 2-hydroxy-1-naphthaldehydethiosemicarbazone coordinated as a monobasic tridentate donor, whereas in complexes 2 and 3, the ligands salicylaldehyde-4(N)-ethylthiosemicarbazone and 2-hydroxy-1-naphthaldehyde-4(N)-ethylthiosemicarbazone coordinated as a dibasic tridentate donor. The DNA binding ability of the complexes in calf thymus DNA was explored by absorption and emission titration experiments. The antioxidant property of the new complexes was evaluated to test their free-radical scavenging ability. In vitro cytotoxicity assays were performed for the new complexes in A549 and HepG2 cell lines. The new compounds overcome cisplatin resistance in the A549 cell line and they were also active in the HepG2 cell line. The cellular uptake study showed the accumulation of the complexes in tumor cells depended on the nature of the ligand attached to the nickel ion.

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...

Thermodynamics of binding interactions between extracellular polymeric substances and heavy metals by isothermal titration microcalorimetry.

PubMed

Yan, Peng; Xia, Jia-Shuai; Chen, You-Peng; Liu, Zhi-Ping; Guo, Jin-Song; Shen, Yu; Zhang, Cheng-Cheng; Wang, Jing

2017-05-01

Extracellular polymeric substances (EPS) play a crucial role in heavy metal bio-adsorption using activated sludge, but the interaction mechanism between heavy metals and EPS remains unclear. Isothermal titration calorimetry was employed to illuminate the mechanism in this study. The results indicate that binding between heavy metals and EPS is spontaneous and driven mainly by enthalpy change. Extracellular proteins in EPS are major participants in the binding process. Environmental conditions have significant impact on the adsorption performance. Divalent and trivalent cations severely impeded the binding of heavy metal ions to EPS. Electrostatic interaction mainly attributed to competition between divalent cations and heavy metal ions; trivalent cations directly competed with heavy metal ions for EPS binding sites. Trivalent cations were more competitive than divalent cations for heavy metal ion binding because they formed complexing bonds. This study facilitates a better understanding about the interaction between heavy metals and EPS in wastewater treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

Theoretical study of metal noble-gas positive ions

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Partridge, Harry; Langhoff, Stephen R.

1989-01-01

Theoretical calculations have been performed to determine the spectroscopic constant for the ground and selected low-lying electronic states of the transition-metal noble-gas ions Var(+), FeAr(+), CoAr(+), CuHe(+), CuAr(+), and CuKr(+). Analogous calculations have been performed for the ground states of the alkali noble-gas ions LiAr(+), LiKr(+), NaAr(+), and KAr(+) and the alkaline-earth noble-gas ion MgAr(+) to contrast the difference in binding energies between the simple and transition-metal noble-gas ions. The binding energies increase with increasing polarizability of the noble-gas ions, as expected for a charge-induced dipole bonding mechanism. It is found that the spectroscopic constants of the X 1Sigma(+) states of the alkali noble-gas ions are well described at the self-consistent field level. In contrast, the binding energies of the transition-metal noble-gas ions are substantially increased by electron correlation.

Screening of biologically important Zn2 + by a chemosensor with fluorescent turn on-off mechanism

NASA Astrophysics Data System (ADS)

Khan, Tanveer A.; Sheoran, Monika; Nikhil Raj M., Venkata; Jain, Surbhi; Gupta, Diksha; Naik, Sunil G.

2018-01-01

Reported herein the synthesis, characterization and biologically important zinc ion binding propensity of a weakly fluorescent chemosensor, 4-methyl-2,6-bis((E)-(2-(4-phenylthiazol-2-yl)hydrazono)methyl)phenol (1). 1H NMR spectroscopic titration experiment reveals the binding knack of 1 to the essential Zn2 +. The photo-physical studies of 1 exhibit an enhancement in the fluorescence by several folds upon binding with the zinc ions attributed to PET-off process, with a binding constant value of 5.22 × 103 M- 1. 1 exhibits an excellent detection range for Zn2 + with lower detection limit value of 2.31 × 10- 8 M. The selectivity of 1 was studied with various mono and divalent metal cations and it was observed that most cations either quenches the fluorescence or remains unchanged except for Cd2 +, which shows a slight enhancement in fluorescence intensity of 1. The ratiometric displacement of Cd2 + ions by Zn2 + ions shows an excellent selectivity towards in-situ detection of Zn2 + ions. Photo-physical studies also support the reversible binding of 1 to Zn2 + ions having on and off mechanism in presence of EDTA. Such recognition of the biologically important zinc ions finds potential application in live cell imaging.

Specificity in cationic interaction with poly(N-isopropylacrylamide).

PubMed

Du, Hongbo; Wickramasinghe, Sumith Ranil; Qian, Xianghong

2013-05-02

Classical molecular dynamics (MD) simulations were conducted for PNIPAM in 1 M monovalent alkali chloride salt solutions as well as in 0.5 M divalent Mg(2+) and Ca(2+) chloride salt solutions. It was found that the strength for the direct alkali ion-amide O binding is strongly correlated with the size of the ionic radius. The smallest Li(+) ion binds strongest to amide O, and the largest Cs(+) ion has the weakest interaction with the amide bond. For the divalent Mg(2+) and Ca(2+) ions, their interactions with the amide bond are weak and appear to be mediated by the water molecules, particularly in the case of Mg(2+), resulting from their strong hydration. The direct binding between the cations and amide O requires partial desovlation of the ions that is energetically unfavorable for Mg(2+) and also to a great extent for Ca(2+). The higher cation charge makes the electrostatic interaction more favorable but the dehydration process less favorable. This competition between electrostatic interaction and the dehydration process largely dictates whether the direct binding between the cation and amide O is energetically preferred or not. For monovalent alkali ions, it is energetically preferred to bind directly with the amide O. Moreover, Li(+) ion is also found to associate strongly with the hydrophobic residues on PNIPAM.

Control of Ion Selectivity in LeuT: Two Na+ Binding Sites with two different mechanisms

PubMed Central

Noskov, Sergei Y.; Roux, Benoît

2016-01-01

The x-ray structure of LeuT, a bacterial homologue of Na+/Cl−-dependent neurotransmitter transporter, provides a great opportunity to better understand the molecular basis of monovalent cation selectivity in ion-coupled transporters. LeuT possesses two ion-binding sites, NA1 and NA2, which are highly selective for Na+. Extensive all-atom free energy molecular dynamics simulations of LeuT embedded in an explicit membrane are performed at different temperatures and various occupancy states of the binding sites to dissect the molecular mechanism of ion selectivity. The results show that the two binding sites display robust selectivity for Na+ over K+ or Li+, the competing ions of most similar radii. Of particular interest, the mechanism primarily responsible for selectivity for each of the two binding sites appears to be different. In site NA1, selectivity for Na+ over K+ arises predominantly from the strong electrostatic field arising from the negatively charged carboxylate group of the leucine substrate coordinating the ion directly. In site NA2, which comprises only neutral ligands, selectivity for Na+ is enforced by the local structural restraints arising from the hydrogen-bonding network and the covalent connectivity of the poly-peptide chain surrounding the ion according to a snug-fit mechanism. PMID:18280500

Structures of E. coli peptide deformylase bound to formate: insight into the preference for Fe2+ over Zn2+ as the active site metal.

PubMed

Jain, Rinku; Hao, Bing; Liu, Ren-Peng; Chan, Michael K

2005-04-06

E. coli peptide deformylase (PDF) catalyzes the deformylation of nascent polypeptides generated during protein synthesis. While PDF was originally thought to be a zinc enzyme, subsequent studies revealed that the active site metal is iron. In an attempt to understand this unusual metal preference, high-resolution structures of Fe-, Co-, and Zn-PDF were determined in complex with its deformylation product, formate. In all three structures, the formate ion binds the metal and forms hydrogen-bonding interactions with the backbone nitrogen of Leu91, the amide side chain of Gln50, and the carboxylate side chain of Glu133. One key difference, however, is how the formate binds the metal. In Fe-PDF and Co-PDF, formate binds in a bidentate fashion, while in Zn-PDF, it binds in a monodentate fashion. Importantly, these structural results provide the first clues into the origins of PDF's metal-dependent activity differences. On the basis of these structures, we propose that the basis for the higher activity of Fe-PDF stems from the better ability of iron to bind and activate the tetrahedral transition state required for cleavage of the N-terminal formyl group.

Cr(3+) Binding to DNA Backbone Phosphate and Bases: Slow Ligand Exchange Rates and Metal Hydrolysis.

PubMed

Zhou, Wenhu; Yu, Tianmeng; Vazin, Mahsa; Ding, Jinsong; Liu, Juewen

2016-08-15

The interaction between chromium ions and DNA is of great interest in inorganic chemistry, toxicology, and analytical chemistry. Most previous studies focused on in situ reduction of Cr(VI), producing Cr(3+) for DNA binding. Recently, Cr(3+) was reported to activate the Ce13d DNAzyme for RNA cleavage. Herein, the Ce13d is used to study two types of Cr(3+) and DNA interactions. First, Cr(3+) binds to the DNA phosphate backbone weakly through reversible electrostatic interactions, which is weakened by adding competing inorganic phosphate. However, Cr(3+) coordinates with DNA nucleobases forming stable cross-links that can survive denaturing gel electrophoresis condition. The binding of Cr(3+) to different nucleobases was further studied in terms of binding kinetics and affinity by exploiting carboxyfluorescein-labeled DNA homopolymers. Once binding takes place, the stable Cr(3+)/DNA complex cannot be dissociated by EDTA, attributable to the ultraslow ligand exchange rate of Cr(3+). The binding rate follows the order of G > C > T ≈ A. Finally, Cr(3+) gradually loses its DNA binding ability after being stored at neutral or high pH, attributable to hydrolysis. This hydrolysis can be reversed by lowering the pH. This work provides a deeper insight into the bioinorganic chemistry of Cr(3+) coordination with DNA, clarifies some inconsistency in the previous literature, and offers practically useful information for generating reproducible results.

Study of the mechanisms of cadmium biosorption by dealginated seaweed waste.

PubMed

Romero-Gonzalez, M E; Williams, C J; Gardiner, P H

2001-07-15

The ability of dealginated seaweed waste, a waste material derived from the commercial processing of seaweed for alginate production, to remove cadmium from solution was determined. Cadmium sorption was found to be rapid (91% removal within 5 min), achieving a residual concentration of 0.8 mg L-1 after 1-h contact time from an initial solution concentration of 10 mg L-1. The binding of cadmium by dealginate was found to be pH dependent, optimal sorption occurring at around pH 6-8. The mechanism of cadmium ion binding by dealginate was investigated by a number of techniques. Potentiometric titration of the dealginate revealed two distinct pKa values, the first having a value similar to carboxyl groups and the second comparable with that of saturated thiols and amines. Esterification of the dealginate resulted in the subsequent reduction in cadmium sorption (95% to 17%), indicating that carboxyl groups are largely responsible for sorption. Evidence from FT-IR spectra confirmed the presence of carboxyl groups in untreated dealginate, while the number of carboxyl groups was markedly reduced in the esterified sample. Furthermore, the FT-IR spectrum for dealginate was found to be similar to that previously reported for mannuronic acid-rich calcium alginate. Determination of a molar ratio in the displacement of calcium by cadmium on dealginate further supported the presence of an ion-exchange relationship. The ion-exchange constant was calculated to be 0.329 x 10(-6). The speciation of cadmium in solution both before and after sorption was determined by an ion-selective electrode (ISE) technique. The findings of this study suggest that the sorption of cadmium by dealginate is mainly due to an ion-exchange mechanism.

Modification of hydroxyapatite with ion-selective complexants: 1-hydroxyethane-1,1-diphosphonic acid

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

Daniels, Yasmine; Lyczko, Nathalie; Nzihou, Ange

Hydroxyapatite (HAP) was modified with 1-hydroxyethane-1,1-diphosphonic acid (HEDP), and its effect on divalent metal ion binding was determined. HAP was synthesized from calcium hydroxide and phosphoric acid. After calcination, it was modified with HEDP, and the influence of time and temperature on the modification was investigated. HEDP incorporation increased as its initial solution concentration increased from 0.01 to 0.50 M. Unmodified and modified HAP were characterized using Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy, energy dispersive X-ray spectroscopy, and specific surface area analysis. Ca/P ratios, acid capacities, and phosphorus elemental analyses gave the effect of modification on compositionmore » and surface characteristics. A high reaction temperature produced new phosphonate bands at 993, 1082, and 1144 cm –1 that indicated the presence of HEDP. HAP modification at a high temperature–long reaction time had the highest HEDP loading and gave the sharpest XRD peaks. The emergence of new HAP–HEDP strands was observed in SEM images for treated samples while EDS showed high phosphorus contents in these strands. Modified HAP had a high acid capacity from the additional P–OH groups in HEDP. The P(O)OH groups maintain their ability to bind metal ions within the HAP matrix: contacting the modified HAP with 10–4 N nitrate solutions of five transition metal ions gives an affinity sequence of Pb(II) > Cd(II) > Zn(II) > Ni(II) > Cu(II). Here, this result is comparable to that of commercially available di(2-ethylhexyl)phosphoric acid, a common solvent extractant, and the trend is consistent with the Misono softness parameter of metal ion polarizabilities.« less

Aminoglycoside antibiotics as a tool for the study of the biological role of calcium ions. Historical overview.

PubMed

Corrado, A P; de Morais, I P; Prado, W A

1989-01-01

Beginning with the pioneering work of Vital-Brazil and Corrado (1957), which suggested a possible interaction between aminoglycoside antibiotics (AGA) and calcium ions at the neuromuscular junction, the authors review the studies that demonstrated the existence of a competitive antagonism between AGA and calcium ions. In view of the low liposolubility of AGA and their inability to cross biological membranes, this antagonism seems to occur exclusively at calcium-binding sites at the level of the outer opening of calcium channels of the N-subtype, which are also the sites of interaction of omega-conotoxin. Being highly water soluble, AGA are easily removed from their binding sites with a consequent rapid reversal of their effects, a factor of primary importance to explain their wide use as tools in the pharmacological analysis of the study of the biological role of calcium ion on the membrane's outer surface. This use has advantages over the use of inorganic di- and trivalent cations such as Mg2+, Mn2+, Cd2+, Ni2+, La3+, etc., since the latter, though they are considered to be the most specific competitive antagonists of calcium ions, may induce biphasic effects due to their ability to cross the membranes and replace calcium and/or increase intracellular calcium concentration. The performance of AGA is also superior when compared with the so-called "specific" organic calcium antagonists--verapamil and nifedipine derivatives--since the latter, in addition to inducing possible biphasic effects, antagonize calcium in a non-competitive manner. Finally, the authors remark that AGA-Ca2+ antagonism relevance is not limited only to basic aspects and that it may have therapeutic implications since it provides alternatives for reducing the toxic adverse effects of this important group of antibiotics.

Modification of hydroxyapatite with ion-selective complexants: 1-hydroxyethane-1,1-diphosphonic acid

DOE PAGES

Daniels, Yasmine; Lyczko, Nathalie; Nzihou, Ange; ...

2014-12-29

Hydroxyapatite (HAP) was modified with 1-hydroxyethane-1,1-diphosphonic acid (HEDP), and its effect on divalent metal ion binding was determined. HAP was synthesized from calcium hydroxide and phosphoric acid. After calcination, it was modified with HEDP, and the influence of time and temperature on the modification was investigated. HEDP incorporation increased as its initial solution concentration increased from 0.01 to 0.50 M. Unmodified and modified HAP were characterized using Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy, energy dispersive X-ray spectroscopy, and specific surface area analysis. Ca/P ratios, acid capacities, and phosphorus elemental analyses gave the effect of modification on compositionmore » and surface characteristics. A high reaction temperature produced new phosphonate bands at 993, 1082, and 1144 cm –1 that indicated the presence of HEDP. HAP modification at a high temperature–long reaction time had the highest HEDP loading and gave the sharpest XRD peaks. The emergence of new HAP–HEDP strands was observed in SEM images for treated samples while EDS showed high phosphorus contents in these strands. Modified HAP had a high acid capacity from the additional P–OH groups in HEDP. The P(O)OH groups maintain their ability to bind metal ions within the HAP matrix: contacting the modified HAP with 10–4 N nitrate solutions of five transition metal ions gives an affinity sequence of Pb(II) > Cd(II) > Zn(II) > Ni(II) > Cu(II). Here, this result is comparable to that of commercially available di(2-ethylhexyl)phosphoric acid, a common solvent extractant, and the trend is consistent with the Misono softness parameter of metal ion polarizabilities.« less

Nonbonded interactions in membrane active cyclic biopolymers. IV - Cation dependence

NASA Technical Reports Server (NTRS)

Radhakrishnan, R.; Srinivasan, S.; Prasad, C. V.; Brinda, S. R.; Macelroy, R. D.; Sundaram, K.

1980-01-01

Interactions of valinomycin and form of its analogs in several conformations with the central ions Li(+), Na(+), K(+), Rb(+) and Cs(+) are investigated as part of a study of the specific preference of valinomycin for potassium and the mechanisms of carrier-mediated ion transport across membranes. Ion binding energies and conformational potential energies are calculated taking into account polarization energy formulas and repulsive energy between the central ion and the ligand atoms for conformations representing various stages in ion capture and release for each of the two ring chiralities of valinomycin and its analogs. Results allow the prediction of the chirality and conformation most likely to be observed for a given analog, and may be used to synthesize analogs with a desired rigidity or flexibility. The binding energies with the alkali metal cations are found to decrease with increasing ion size, and to be smaller than the corresponding ion hydration energies. It is pointed out that the observed potassium preference may be explainable in terms of differences between binding and hydration energies. Binding energies are also noted to depend on ligand conformation.

Interaction of ligands with the opiate receptors of brain membranes: Regulation by ions and nucleotides

PubMed Central

Blume, Arthur J.

1978-01-01

This study shows that nucleotides, as well as ions, regulate the opiate receptors of brain. GMP-P(NH)P and Na+ reduce the amount of steady-state specific [3H]dihydromorphine binding and increase the rate of dissociation of the ligand from the opiate receptor. In contrast, Mn2+ decreases the rate of ligand dissociation and antagonizes the ability of Na+ to increase dissociation. The effects of GMP-P(NH)P on steady-state binding and dissociation are not reversed by washing. Only GTP, GDP, ITP, and IMP-P(NH)P, in addition to GMP-P(NH)P, increase the rate of dihydromorphine dissociation. The site of nucleotide action appears to have high affinity: <1 μM GMP-P(NH)P produces half-maximal increases in ligand dissociation. GMP-P(NH)P- and Na+-directed increases in dissociation have also been found for the opiate agonists [3H]etorphine, [3H]Leu-enkephalin, and [3H]Met-enkephalin and the opiate antagonist [3H]naltrexone. Mn2+-directed decreases in dissociation have been found for the agonist [3H]-etorphine and the antagonist [3H]naltrexone. Although the plasma membrane receptors for a number of other neuro-transmitters and hormones are also regulated by guanine nucleotides, the opiate receptors appear unique because only they show nucleotide regulation of both agonist and antagonist binding. PMID:205867

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.

A membrane-embedded pathway delivers general anesthetics to two interacting binding sites in the Gloeobacter violaceus ion channel.

PubMed

Arcario, Mark J; Mayne, Christopher G; Tajkhorshid, Emad

2017-06-09

General anesthetics exert their effects on the central nervous system by acting on ion channels, most notably pentameric ligand-gated ion channels. Although numerous studies have focused on pentameric ligand-gated ion channels, the details of anesthetic binding and channel modulation are still debated. A better understanding of the anesthetic mechanism of action is necessary for the development of safer and more efficacious drugs. Herein, we present a computational study identifying two anesthetic binding sites in the transmembrane domain of the Gloeobacter violaceus ligand-gated ion channel (GLIC) channel, characterize the putative binding pathway, and observe structural changes associated with channel function. Molecular simulations of desflurane reveal a binding pathway to GLIC via a membrane-embedded tunnel using an intrasubunit protein lumen as the conduit, an observation that explains the Meyer-Overton hypothesis, or why the lipophilicity of an anesthetic and its potency are generally proportional. Moreover, employing high concentrations of ligand led to the identification of a second transmembrane site (TM2) that inhibits dissociation of anesthetic from the TM1 site and is consistent with the high concentrations of anesthetics required to achieve clinical effects. Finally, asymmetric binding patterns of anesthetic to the channel were found to promote an iris-like conformational change that constricts and dehydrates the ion pore, creating a 13.5 kcal/mol barrier to ion translocation. Together with previous studies, the simulations presented herein demonstrate a novel anesthetic binding site in GLIC that is accessed through a membrane-embedded tunnel and interacts with a previously known site, resulting in conformational changes that produce a non-conductive state of the channel. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Development of peptoid-based ligands for the removal of cadmium from biological media

DOE PAGES

Knight, Abigail S.; Zhou, Effie Y.; Francis, Matthew B.

2015-05-14

Cadmium poisoning poses a serious health concern due to cadmium's increasing industrial use, yet there is currently no recommended treatment. The selective coordination of cadmium in a biological environment—i.e. in the presence of serum ions, small molecules, and proteins—is a difficult task. To address this challenge, a combinatorial library of peptoid-based ligands has been evaluated to identify structures that selectively bind to cadmium in human serum with minimal chelation of essential metal ions. Eighteen unique ligands were identified in this screening procedure, and the binding affinity of each was measured using metal titrations monitored by UV-vis spectroscopy. To evaluate themore » significance of each chelating moiety, sequence rearrangements and substitutions were examined. Analysis of a metal–ligand complex by NMR spectroscopy highlighted the importance of particular residues. Depletion experiments were performed in serum mimetics and human serum with exogenously added cadmium. These depletion experiments were used to compare and demonstrate the ability of these peptoids to remove cadmium from blood-like mixtures. In one of these depletion experiments, the peptoid sequence was able to deplete the cadmium to a level comparable to the reported acute toxicity limit. Evaluation of the metal selectivity in buffered solution and in human serum was performed to verify minimal off-target binding. These studies highlight a screening platform for the identification of metal–ligands that are capable of binding in a complex environment. They additionally demonstrate the potential utility of biologically-compatible ligands for the treatment of heavy metal poisoning.« less

Development of peptoid-based ligands for the removal of cadmium from biological media

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

Knight, Abigail S.; Zhou, Effie Y.; Francis, Matthew B.

Cadmium poisoning poses a serious health concern due to cadmium's increasing industrial use, yet there is currently no recommended treatment. The selective coordination of cadmium in a biological environment—i.e. in the presence of serum ions, small molecules, and proteins—is a difficult task. To address this challenge, a combinatorial library of peptoid-based ligands has been evaluated to identify structures that selectively bind to cadmium in human serum with minimal chelation of essential metal ions. Eighteen unique ligands were identified in this screening procedure, and the binding affinity of each was measured using metal titrations monitored by UV-vis spectroscopy. To evaluate themore » significance of each chelating moiety, sequence rearrangements and substitutions were examined. Analysis of a metal–ligand complex by NMR spectroscopy highlighted the importance of particular residues. Depletion experiments were performed in serum mimetics and human serum with exogenously added cadmium. These depletion experiments were used to compare and demonstrate the ability of these peptoids to remove cadmium from blood-like mixtures. In one of these depletion experiments, the peptoid sequence was able to deplete the cadmium to a level comparable to the reported acute toxicity limit. Evaluation of the metal selectivity in buffered solution and in human serum was performed to verify minimal off-target binding. These studies highlight a screening platform for the identification of metal–ligands that are capable of binding in a complex environment. They additionally demonstrate the potential utility of biologically-compatible ligands for the treatment of heavy metal poisoning.« less

Ion sensing by charge transfer absorption variations of benzocrown-bipyridinium conjugates with an alkyl chain.

PubMed

Kuwabara, Tetsuo; Satake, Ryota; Guo, Haocheng

2015-01-01

Two benzocrown ether-bipyridinium conjugates, 1 and 2, each having a different length of alkyl chains with butyl and dodecyl groups, respectively, have been synthesized for the purpose of developing a new guest-responsive color-change chemosensor. Both 1 and 2 showed yellow colors with broad absorption bands around 400 nm in acetonitrile. These are associated with the intramolecular charge transfer (CT) absorption, in which the benzocrown ether and bipyridinium units act as the donor and acceptor, respectively. Upon addition of the guest; such as Na(+), they faded in color due to the blue shift in their intramolecular charge transfer absorption bands. These are associated with the formation of 1:1 host-guest inclusion complex. Analogues, 3 and 4, both being similar in structure to 1 and 2 with non-crown ether unit, also showed intramolecular CT absorptions around 400 nm, but did not change their absorption spectra upon addition of the guest because of the lack of guest-binding abilities. The guest-induced color change of 1 and 2 can be used for alkali and alkaline metal ion sensing. Both 1 and 2 could detect divalent cations such as Mg(2+) and Ca(2+) rather than univalent ones, Li(+), Na(+), K(+), Rb(+), and Cs(+). Although a marked difference between 1 and 2 was not observed in their guest sensing abilities, the remarkable recognition of 1 and 2 for Mg(2+) and Ca(2+) was found compared with that of 5, which has benzyl unit instead of alkyl chains of 1 and 2. The sensitivity values of 1 and 2 were roughly proportional to their binding constants, as shown by the binding constants with Li(+), Na(+), Mg(2+), and Ca(2+) with the values of 910, 260, 820, and 2300 M(-1) for 1 and 930, 290, 1270, and 2790 M(-1) for 2, while the binding constants of 5 were estimated to be 930, 440, 210, and 1200 M(-1) for Li(+), Na(+), Mg(2+), and Ca(2+), respectively. The limit concentration of detection of 2 for Ca(2+) was estimated to be 0.016 mM, which was the smallest value in this system.

Multitargeting by curcumin as revealed by molecular interaction studies

PubMed Central

Gupta, Subash C.; Prasad, Sahdeo; Kim, Ji Hye; Patchva, Sridevi; Webb, Lauren J.; Priyadarsini, Indira K.

2012-01-01

Curcumin (diferuloylmethane), the active ingredient in turmeric (Curcuma longa), is a highly pleiotropic molecule with anti-inflammatory, anti-oxidant, chemopreventive, chemosensitization, and radiosensitization activities. The pleiotropic activities attributed to curcumin come from its complex molecular structure and chemistry, as well as its ability to influence multiple signaling molecules. Curcumin has been shown to bind by multiple forces directly to numerous signaling molecules, such as inflammatory molecules, cell survival proteins, protein kinases, protein reductases, histone acetyltransferase, histone deacetylase, glyoxalase I, xanthine oxidase, proteasome, HIV1 integrase, HIV1 protease, sarco (endo) plasmic reticulum Ca2+ ATPase, DNA methyltransferases 1, FtsZ protofilaments, carrier proteins, and metal ions. Curcumin can also bind directly to DNA and RNA. Owing to its β-diketone moiety, curcumin undergoes keto–enol tautomerism that has been reported as a favorable state for direct binding. The functional groups on curcumin found suitable for interaction with other macromolecules include the α, β-unsaturated β-diketone moiety, carbonyl and enolic groups of the β-diketone moiety, methoxy and phenolic hydroxyl groups, and the phenyl rings. Various biophysical tools have been used to monitor direct interaction of curcumin with other proteins, including absorption, fluorescence, Fourier transform infrared (FTIR) and circular dichroism (CD) spectroscopy, surface plasmon resonance, competitive ligand binding, Forster type fluorescence resonance energy transfer (FRET), radiolabeling, site-directed mutagenesis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), immunoprecipitation, phage display biopanning, electron microscopy, 1-anilino-8-naphthalene-sulfonate (ANS) displacement, and co-localization. Molecular docking, the most commonly employed computational tool for calculating binding affinities and predicting binding sites, has also been used to further characterize curcumin’s binding sites. Furthermore, the ability of curcumin to bind directly to carrier proteins improves its solubility and bioavailability. In this review, we focus on how curcumin directly targets signaling molecules, as well as the different forces that bind the curcumin–protein complex and how this interaction affects the biological properties of proteins. We will also discuss various analogues of curcumin designed to bind selective targets with increased affinity. PMID:21979811

Review: Milk Proteins as Nanocarrier Systems for Hydrophobic Nutraceuticals.

PubMed

Kimpel, Florian; Schmitt, Joachim J

2015-11-01

Milk proteins and milk protein aggregates are among the most important nanovehicles in food technology. Milk proteins have various functional properties that facilitate their ability to carry hydrophobic nutraceutical substances. The main functional transport properties that were examined in the reviewed studies are binding of molecules or ions, surface activity, aggregation, gelation, and interaction with other polymers. Hydrophobic binding has been investigated using caseins and isolated β-casein as well as whey proteins. Surface activity of caseins has been used to create emulsion-based carrier systems. Furthermore, caseins are able to self-assemble into micelles, which can incorporate molecules. Gelation and interaction with other polymers can be used to encapsulate molecules into protein networks. The release of transported substances mainly depends on pH and swelling behavior of the proteins. The targeted use of nanocarrier systems requires specific knowledge about the binding mechanisms between the proteins and the carried substances in a certain food matrix. © 2015 Institute of Food Technologists®

Thiophene-based rhodamine as selectivef luorescence probe for Fe(III) and Al(III) in living cells.

PubMed

Wang, Kun-Peng; Chen, Ju-Peng; Zhang, Si-Jie; Lei, Yang; Zhong, Hua; Chen, Shaojin; Zhou, Xin-Hong; Hu, Zhi-Qiang

2017-09-01

The thiophene-modified rhodamine 6G (GYJ) has been synthesized as a novel chemosensor. The sensor has sufficiently high selectivity and sensitivity for the detection of Fe 3+ and Al 3+ ions (M 3+ ) by fluorescence and ultraviolet spectroscopy with a strong ability for anti-interference performance. The binding ratio of M 3+ -GYJ complex was determined to be 2:1 according to the Job's plot. The binding constants for Fe 3+ and Al 3+ were calculated to be 3.91 × 10 8 and 5.26 × 10 8  M -2 , respectively. All these unique features made it particularly favorable for cellular imaging applications. The obvious fluorescence microscopy experiments demonstrated that the probes could contribute to the detection of Fe 3+ and Al 3+ in related cells and biological organs with satisfying resolution. Graphical abstract GYJ has high selectivity and sensitivity for the detection of Fe(III) and Al(III) with the binding ratio of 2:1.

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.

Structure of Urtica dioica agglutinin isolectin I: dimer formation mediated by two zinc ions bound at the sugar-binding site.

PubMed

Harata, K; Schubert, W D; Muraki, M

2001-11-01

Ultica dioica agglutinin, a plant lectin from the stinging nettle, consists of a total of seven individual isolectins. One of these structures, isolectin I, was determined at 1.9 A resolution by the X-ray method. The crystals belong to the space group P2(1) and the asymmetric unit contains two molecules related by local twofold symmetry. The molecule consists of two hevein-like chitin-binding domains lacking distinct secondary structure, but four disulfide bonds in each domain maintain the tertiary structure. The backbone structure of the two independent molecules is essentially identical and this is similarly true of the sugar-binding sites. In the crystal, the C-terminal domains bind Zn(2+) ions at the sugar-binding site. Owing to their location near a pseudo-twofold axis, the two zinc ions link the two independent molecules in a tail-to-tail arrangement: thus, His47 of molecule 1 and His67 of molecule 2 coordinate the first zinc ion, while the second zinc ion links Asp75 of molecule 1 and His47 of molecule 2.

Synthesis and characterization of glucosyl-curcuminoids as Fe3+ suppliers in the treatment of iron deficiency.

PubMed

Ferrari, Erika; Arezzini, Beatrice; Ferrali, Marco; Lazzari, Sandra; Pignedoli, Francesca; Spagnolo, Ferdinando; Saladini, Monica

2009-10-01

The Fe(3+) chelating ability of some curcumin glucosyl derivatives (Glc-H; Glc-OH; Glc-OCH(3)) is tested by means of UV and NMR study. The pK(a) values of the ligands and the overall stability constants of Fe(3+) and Ga(3+) complexes are evaluated from UV spectra. The only metal binding site of the ligand is the beta-diketo moiety in the keto-enolic form; the glucosyl moiety does not interact with metal ion but it contributes to the stability of metal/ligand 1:2 complexes by means of hydrophilic interactions. These glucosyl derivatives are able to bind Fe(3+) in a wide pH rage, forming complex species thermodynamically more stable than those of other ligands commonly used in the treatment of iron deficiency. In addition they demonstrate to have a poor affinity for competitive biological metal ions such as Ca(2+). All ligands and their iron complexes have a good lypophilicity (log P > -0.7) suggesting an efficient gastrointestinal absorption in view of their possible use as iron supplements in oral therapy. The ligand molecules are also tested for their antioxidant properties in "ex vivo" biological system.

Using 15N-Ammonium to Characterise and Map Potassium Binding Sites in Proteins by NMR Spectroscopy

PubMed Central

Werbeck, Nicolas D; Kirkpatrick, John; Reinstein, Jochen; Hansen, D Flemming

2014-01-01

A variety of enzymes are activated by the binding of potassium ions. The potassium binding sites of these enzymes are very specific, but ammonium ions can often replace potassium ions in vitro because of their similar ionic radii. In these cases, ammonium can be used as a proxy for potassium to characterise potassium binding sites in enzymes: the 1H,15N spin-pair of enzyme-bound 15NH4+ can be probed by 15N-edited heteronuclear NMR experiments. Here, we demonstrate the use of NMR spectroscopy to characterise binding of ammonium ions to two different enzymes: human histone deacetylase 8 (HDAC8), which is activated allosterically by potassium, and the bacterial Hsp70 homologue DnaK, for which potassium is an integral part of the active site. Ammonium activates both enzymes in a similar way to potassium, thus supporting this non-invasive approach. Furthermore, we present an approach to map the observed binding site onto the structure of HDAC8. Our method for mapping the binding site is general and does not require chemical shift assignment of the enzyme resonances. PMID:24520048

Lanthanide binding and IgG affinity construct: Potential applications in solution NMR, MRI, and luminescence microscopy

PubMed Central

Barb, Adam W; Ho, Tienhuei Grace; Flanagan-Steet, Heather; Prestegard, James H

2012-01-01

Paramagnetic lanthanide ions when bound to proteins offer great potential for structural investigations that utilize solution nuclear magnetic resonance spectroscopy, magnetic resonance imaging, or optical microscopy. However, many proteins do not have native metal ion binding sites and engineering a chimeric protein to bind an ion while retaining affinity for a protein of interest represents a significant challenge. Here we report the characterization of an immunoglobulin G-binding protein redesigned to include a lanthanide binding motif in place of a loop between two helices (Z-L2LBT). It was shown to bind Tb3+ with 130 nM affinity. Ions such as Dy3+, Yb3+, and Ce3+ produce paramagnetic effects on NMR spectra and the utility of these effects is illustrated by their use in determining a structural model of the metal-complexed Z-L2LBT protein and a preliminary characterization of the dynamic distribution of IgG Fc glycan positions. Furthermore, this designed protein is demonstrated to be a novel IgG-binding reagent for magnetic resonance imaging (Z-L2LBT:Gd3+ complex) and luminescence microscopy (Z-L2LBT: Tb3+ complex). PMID:22851279

Ion Selectivity in the KcsA Potassium Channel from the Perspective of the Ion Binding Site

PubMed Central

Dixit, Purushottam D.; Merchant, Safir; Asthagiri, D.

2009-01-01

To understand the thermodynamic exclusion of Na+ relative to K+ from the S2 site of the selectivity filter, the distribution PX(ɛ) (X = K+ or Na+) of the binding energy (ɛ) of the ion with the channel is analyzed using the potential distribution theorem. By expressing the excess chemical potential of the ion as a sum of mean-field 〈ɛ〉 and fluctuation μexflux,X contributions, we find that selectivity arises from a higher value of μflux,Na+ex relative to μflux,K+ex. To understand the role of site-site interactions on μexflux,X, we decompose PX(ɛ) into n-dependent distributions, where n is the number of ion-coordinating ligands within a distance λ from the ion. For λ comparable to typical ion-oxygen bond distances, investigations building on this multistate model reveal an inverse correlation between favorable ion-site and site-site interactions: the ion-coordination states that most influence the thermodynamics of the ion are also those for which the binding site is energetically less strained and vice versa. This correlation motivates understanding entropic effects in ion binding to the site and leads to the finding that μexflux,X is directly proportional to the average site-site interaction energy, a quantity that is sensitive to the chemical type of the ligand coordinating the ion. Increasing the coordination number around Na+ only partially accounts for the observed magnitude of selectivity; acknowledging the chemical type of the ion-coordinating ligand is essential. PMID:19289040

Importance of length and sequence order on magnesium binding to surface-bound oligonucleotides studied by second harmonic generation and atomic force microscopy.

PubMed

Holland, Joseph G; Geiger, Franz M

2012-06-07

The binding of magnesium ions to surface-bound single-stranded oligonucleotides was studied under aqueous conditions using second harmonic generation (SHG) and atomic force microscopy (AFM). The effect of strand length on the number of Mg(II) ions bound and their free binding energy was examined for 5-, 10-, 15-, and 20-mers of adenine and guanine at pH 7, 298 K, and 10 mM NaCl. The binding free energies for adenine and guanine sequences were calculated to be -32.1(4) and -35.6(2) kJ/mol, respectively, and invariant with strand length. Furthermore, the ion density for adenine oligonucleotides did not change as strand length increased, with an average value of 2(1) ions/strand. In sharp contrast, guanine oligonucleotides displayed a linear relationship between strand length and ion density, suggesting that cooperativity is important. This data gives predictive capabilities for mixed strands of various lengths, which we exploit for 20-mers of adenines and guanines. In addition, the role sequence order plays in strands of hetero-oligonucleotides was examined for 5'-A(10)G(10)-3', 5'-(AG)(10)-3', and 5'-G(10)A(10)-3' (here the -3' end is chemically modified to bind to the surface). Although the free energy of binding is the same for these three strands (averaged to be -33.3(4) kJ/mol), the total ion density increases when several guanine residues are close to the 3' end (and thus close to the solid support substrate). To further understand these results, we analyzed the height profiles of the functionalized surfaces with tapping-mode atomic force microscopy (AFM). When comparing the average surface height profiles of the oligonucleotide surfaces pre- and post- Mg(II) binding, a positive correlation was found between ion density and the subsequent height decrease following Mg(II) binding, which we attribute to reductions in Coulomb repulsion and strand collapse once a critical number of Mg(II) ions are bound to the strand.

Inhibition of ferric ion to oxalate oxidase shed light on the substrate binding site.

PubMed

Pang, Yu; Lan, Wanjun; Huang, Xuelei; Zuo, Guanke; Liu, Hui; Zhang, Jingyan

2015-10-01

Oxalate oxidase (OxOx), a well known enzyme catalyzes the cleavage of oxalate to carbon dioxide with reduction of dioxygen to hydrogen peroxide, however its catalytic process is not well understood. To define the substrate binding site, interaction of Fe(3+) ions with OxOx was systemically investigated using biochemical method, circular dichrosim spectroscopy, microscale thermophoresis, and computer modeling. We demonstrated that Fe(3+) is a non-competitive inhibitor with a milder binding affinity to OxOx, and the secondary structure of the OxOx was slightly altered upon its binding. On the basis of the structural properties of the OxOx and its interaction with Fe(3+) ions, two residue clusters of OxOx were assigned as potential Fe(3+) binding sites, the mechanism of the inhibition of Fe(3+) was delineated. Importantly, the residues that interact with Fe(3+) ions are involved in the substrate orienting based on computer docking. Consequently, the interaction of OxOx with Fe(3+) highlights insight into substrate binding site in OxOx.

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.

Calorimetric and counterion binding studies of the interactions between micelles and ions. The observation of lyotropic series

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

Larsen, J.W.; Magid, L.J.

1974-09-04

Heats of transfer of a variety of salts from water to solutions of hexadecyltrimethylammonium bromide (CTAB), dodecyltrimethylammonium bromide (DTAB), and sodium dodecyl sulfate (NaLS) were measured. Lyotropic series for both cations and anions were observed for all soaps, the series for the 2 cationic soaps being almost identical. The dependence of the observed heats of transfer for anions from H/sub 2/O to CTAB and DTAB solutions and for cations from H2O to NaLS solutions on the hydrated radii of the ions involved supports the contention that favorable binding of counterions depends on how closely they can approach the charged micellarmore » surfaces. It is clear that a lyotropic series similar to that existing for proteins exists for ion binding to micelles. The controlling factor in this binding seems to be the distance of closest approach of the ion to the micelle, although polarizable organic ions may be the exceptions. Chain length has little effect on binding. It is felt that the work discussed has established the usefulness of a calorimetric investigation and the use of ion-specific electrodes for characterizing surfactant systems containing more than one species of counterions. (37 refs.)« less

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.

Fusaric acid induces a notochord malformation in zebrafish via copper chelation.

PubMed

Yin, Emily S; Rakhmankulova, Malika; Kucera, Kaury; de Sena Filho, Jose Guedes; Portero, Carolina E; Narváez-Trujillo, Alexandra; Holley, Scott A; Strobel, Scott A

2015-08-01

Over a thousand extracts were tested for phenotypic effects in developing zebrafish embryos to identify bioactive molecules produced by endophytic fungi. One extract isolated from Fusarium sp., a widely distributed fungal genus found in soil and often associated with plants, induced an undulated notochord in developing zebrafish embryos. The active compound was isolated and identified as fusaric acid. Previous literature has shown this phenotype to be associated with copper chelation from the active site of lysyl oxidase, but the ability of fusaric acid to bind copper ions has not been well described. Isothermal titration calorimetry revealed that fusaric acid is a modest copper chelator with a binding constant of 4.4 × 10(5) M(-1). These results shed light on the toxicity of fusaric acid and the potential teratogenic effects of consuming plants infected with Fusarium sp.

Simulation of a model nanopore sensor: Ion competition underlies device behavior.

PubMed

Mádai, Eszter; Valiskó, Mónika; Dallos, András; Boda, Dezső

2017-12-28

We study a model nanopore sensor with which a very low concentration of analyte molecules can be detected on the basis of the selective binding of the analyte molecules to the binding sites on the pore wall. The bound analyte ions partially replace the current-carrier cations in a thermodynamic competition. This competition depends both on the properties of the nanopore and the concentrations of the competing ions (through their chemical potentials). The output signal given by the device is the current reduction caused by the presence of the analyte ions. The concentration of the analyte ions can be determined through calibration curves. We model the binding site with the square-well potential and the electrolyte as charged hard spheres in an implicit background solvent. We study the system with a hybrid method in which we compute the ion flux with the Nernst-Planck (NP) equation coupled with the Local Equilibrium Monte Carlo (LEMC) simulation technique. The resulting NP+LEMC method is able to handle both strong ionic correlations inside the pore (including finite size of ions) and bulk concentrations as low as micromolar. We analyze the effect of bulk ion concentrations, pore parameters, binding site parameters, electrolyte properties, and voltage on the behavior of the device.

Simulation of a model nanopore sensor: Ion competition underlies device behavior

NASA Astrophysics Data System (ADS)

Mádai, Eszter; Valiskó, Mónika; Dallos, András; Boda, Dezső

2017-12-01

We study a model nanopore sensor with which a very low concentration of analyte molecules can be detected on the basis of the selective binding of the analyte molecules to the binding sites on the pore wall. The bound analyte ions partially replace the current-carrier cations in a thermodynamic competition. This competition depends both on the properties of the nanopore and the concentrations of the competing ions (through their chemical potentials). The output signal given by the device is the current reduction caused by the presence of the analyte ions. The concentration of the analyte ions can be determined through calibration curves. We model the binding site with the square-well potential and the electrolyte as charged hard spheres in an implicit background solvent. We study the system with a hybrid method in which we compute the ion flux with the Nernst-Planck (NP) equation coupled with the Local Equilibrium Monte Carlo (LEMC) simulation technique. The resulting NP+LEMC method is able to handle both strong ionic correlations inside the pore (including finite size of ions) and bulk concentrations as low as micromolar. We analyze the effect of bulk ion concentrations, pore parameters, binding site parameters, electrolyte properties, and voltage on the behavior of the device.

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.

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

FRET study of G-quadruplex forming fluorescent oligonucleotide probes at the lipid monolayer interface.

PubMed

Swiatkowska, Angelika; Kosman, Joanna; Juskowiak, Bernard

2016-01-05

Spectral properties and G-quadruplex folding ability of fluorescent oligonucleotide probes at the cationic dioctadecyldimethylammonium bromide (DODAB) monolayer interface are reported. Two oligonucleotides, a 19-mer bearing thrombin binding aptamer sequence and a 21-mer with human telomeric sequence, were end-labeled with fluorescent groups (FAM and TAMRA) to give FRET probes F19T and F21T, respectively. The probes exhibited abilities to fold into a quadruplex structure and to bind metal cations (Na(+) and K(+)). Fluorescence spectra of G-quadruplex FRET probes at the monolayer interface are reported for the first time. Investigations included film balance measurements (π-A isotherms) and fluorescence spectra recording using a fiber optic accessory interfaced with a spectrofluorimeter. The effect of the presence of DODAB monolayer, metal cations and the surface pressure of monolayer on spectral behavior of FRET probes were examined. Adsorption of probe at the cationic monolayer interface resulted in the FRET signal enhancement even in the absence of metal cations. Variation in the monolayer surface pressure exerted rather modest effect on the spectral properties of probes. The fluorescence energy transfer efficiency of monolayer adsorbed probes increased significantly in the presence of sodium or potassium ion in subphase, which indicated that the probes retained their cation binding properties when adsorbed at the monolayer interface. Copyright © 2015 Elsevier B.V. All rights reserved.

Multiple Ion Binding Equilibria, Reaction Kinetics, and Thermodynamics in Dynamic Models of Biochemical Pathways

PubMed Central

Vinnakota, Kalyan C.; Wu, Fan; Kushmerick, Martin J.; Beard, Daniel A.

2009-01-01

The operation of biochemical systems in vivo and in vitro is strongly influenced by complex interactions between biochemical reactants and ions such as H+, Mg2+, K+, and Ca2+. These are important second messengers in metabolic and signaling pathways that directly influence the kinetics and thermodynamics of biochemical systems. Herein we describe the biophysical theory and computational methods to account for multiple ion binding to biochemical reactants and demonstrate the crucial effects of ion binding on biochemical reaction kinetics and thermodynamics. In simulations of realistic systems, the concentrations of these ions change with time due to dynamic buffering and competitive binding. In turn, the effective thermodynamic properties vary as functions of cation concentrations and important environmental variables such as temperature and overall ionic strength. Physically realistic simulations of biochemical systems require incorporating all of these phenomena into a coherent mathematical description. Several applications to physiological systems are demonstrated based on this coherent simulation framework. PMID:19216922

Structure and electronic properties of ion pairs accompanying cyclic morpholinium cation and alkylphosphite anion based ionic liquids

NASA Astrophysics Data System (ADS)

Verma, Prakash L.; Singh, Priti; Gejji, Shridhar P.

2017-07-01

Molecular insights for the formation of ion pairs accompanying the cyclic ammonium cation based room temperature ionic liquids (RTILs) composed of alkyl substituted N-methylmorpholinium (RMMor) and alkylphosphite [(Rsbnd O)2PHdbnd O] (Rdbnd ethyl, butyl, hexyl, octyl) anion have been derived from the M06-2x level of theory. Electronic structures, binding energies, and spectral characteristics of the ion pairs underlying these RTILs have been characterized. The ion pair formation is largely governed by Csbnd H⋯O and other intermolecular interactions. Calculated binding energies increase with the increasing alkyl chain on either cation or alkylphosphite anion. The cation-anion binding reveals signature in the frequency down-(red) shift of the characteristic anionic Pdbnd O stretching whereas the Psbnd H stretching exhibits a shift in the opposite direction in vibrational spectra which has further been rationalized through molecular electron density topography. Correlations of measured electrochemical stability with the separation of frontier orbital energies and binding energies in the ion pairs have further been established.

Ammonium Ion Binding to DNA G-Quadruplexes: Do Electrospray Mass Spectra Faithfully Reflect the Solution-Phase Species?

NASA Astrophysics Data System (ADS)

Balthasart, Françoise; Plavec, Janez; Gabelica, Valérie

2013-01-01

G-quadruplex nucleic acids can bind ammonium ions in solution, and these complexes can be detected by electrospray mass spectrometry (ESI-MS). However, because ammonium ions are volatile, the extent to which ESI-MS quantitatively could provide an accurate reflection of such solution-phase equilibria is unclear. Here we studied five G-quadruplexes having known solution-phase structure and ammonium ion binding constants: the bimolecular G-quadruplexes (dG4T4G4)2, (dG4T3G4)2, and (dG3T4G4)2, and the intramolecular G-quadruplexes dG4(T4G4)3 and dG2T2G2TGTG2T2G2 (thrombin binding aptamer). We found that not all mass spectrometers are equally suited to reflect the solution phase species. Ion activation can occur in the electrospray source, or in a high-pressure traveling wave ion mobility cell. When the softest instrumental conditions are used, ammonium ions bound between G-quartets, but also additional ammonium ions bound at specific sites outside the external G-quartets, can be observed. However, even specifically bound ammonium ions are in some instances too labile to be fully retained in the gas phase structures, and although the ammonium ion distribution observed by ESI-MS shows biases at specific stoichiometries, the relative abundances in solution are not always faithfully reflected. Ion mobility spectrometry results show that all inter-quartet ammonium ions are necessary to preserve the G-quadruplex fold in the gas phase. Ion mobility experiments, therefore, help assign the number of inner ammonium ions in the solution phase structure.[Figure not available: see fulltext.

Sequential water molecule binding enthalpies for aqueous nanodrops containing a mono-, di- or trivalent ion and between 20 and 500 water molecules† †Electronic supplementary information (ESI) available: Detailed description of the experimental and computational modeling methods. Isolation, BIRD and UVPD sequence for [Ru(NH3)6]3+·(H2O)169–171, nanoESI spectra for 2+ and 3+ ions. Detailed description of the isotope distribution simulation program. Comparison between experimental and simulated 1+, 2+ and 3+ ion isotope distributions. Wavelength dependence of the deduced sequential binding enthalpies. Comparison of experimental UVPD binding enthalpies to the liquid drop model at different temperatures. Complete list of binding enthalpies and average number of water molecules lost upon UVPD. See DOI: 10.1039/c6sc04957e Click here for additional data file.

PubMed Central

Heiles, Sven; Cooper, Richard J.; DiTucci, Matthew J.

2017-01-01

Sequential water molecule binding enthalpies, ΔH n,n–1, are important for a detailed understanding of competitive interactions between ions, water and solute molecules, and how these interactions affect physical properties of ion-containing nanodrops that are important in aerosol chemistry. Water molecule binding enthalpies have been measured for small clusters of many different ions, but these values for ion-containing nanodrops containing more than 20 water molecules are scarce. Here, ΔH n,n–1 values are deduced from high-precision ultraviolet photodissociation (UVPD) measurements as a function of ion identity, charge state and cluster size between 20–500 water molecules and for ions with +1, +2 and +3 charges. The ΔH n,n–1 values are obtained from the number of water molecules lost upon photoexcitation at a known wavelength, and modeling of the release of energy into the translational, rotational and vibrational motions of the products. The ΔH n,n–1 values range from 36.82 to 50.21 kJ mol–1. For clusters containing more than ∼250 water molecules, the binding enthalpies are between the bulk heat of vaporization (44.8 kJ mol–1) and the sublimation enthalpy of bulk ice (51.0 kJ mol–1). These values depend on ion charge state for clusters with fewer than 150 water molecules, but there is a negligible dependence at larger size. There is a minimum in the ΔH n,n–1 values that depends on the cluster size and ion charge state, which can be attributed to the competing effects of ion solvation and surface energy. The experimental ΔH n,n–1 values can be fit to the Thomson liquid drop model (TLDM) using bulk ice parameters. By optimizing the surface tension and temperature change of the logarithmic partial pressure for the TLDM, the experimental sequential water molecule binding enthalpies can be fit with an accuracy of ±3.3 kJ mol–1 over the entire range of cluster sizes. PMID:28451364

Production and Characterization of Desmalonichrome Relative Binding Affinity for Uranyl Ions in Relation to Other Siderophores

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

Mo, Kai-For; Dai, Ziyu; Wunschel, David S.

2016-06-24

Siderophores are Fe binding secondary metabolites that have been investigated for their uranium binding properties. Much of the previous work has focused on characterizing hydroxamate types of siderophores, such as desferrioxamine B, for their uranyl binding affinity. Carboxylate forms of these metabolites hold potential to be more efficient chelators of uranyl, yet they have not been widely studied and are more difficult to obtain. Desmalonichrome is a carboxylate siderophore which is not commercially available and so was obtained from the ascomycete fungus Fusarium oxysporum cultivated under Fe depleted conditions. The relative affinity for uranyl binding of desmalonichrome was investigated usingmore » a competitive analysis of binding affinities between uranyl acetate and different concentrations of iron(III) chloride using electrospray ionization mass spectrometry (ESI-MS). In addition to desmalonichrome, three other siderophores, including two hydroxamates (desferrioxamine B and desferrichrome) and one carboxylate (desferrichrome A) were studied to understand their relative affinities for the uranyl ion at two pH values. The binding affinities of hydroxymate siderophores to uranyl ion were found to decrease to a greater degree at lower pH as the concentration of Fe (III) ion increases. On the other hand, lowering pH has little impact on the binding affinities between carboxylate siderophores and uranyl ion. Desmalonichrome was shown to have the greatest relative affinity for uranyl at any pH and Fe(III) concentration. These results suggest that acidic functional groups in the ligands are critical for strong chelation with uranium at lower pH.« less

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.

Computational Assessment of Potassium and Magnesium Ion Binding to a Buried Pocket in GTPase-Associating Center RNA

PubMed Central

2016-01-01

An experimentally well-studied model of RNA tertiary structures is a 58mer rRNA fragment, known as GTPase-associating center (GAC) RNA, in which a highly negative pocket walled by phosphate oxygen atoms is stabilized by a chelated cation. Although such deep pockets with more than one direct phosphate to ion chelation site normally include magnesium, as shown in one GAC crystal structure, another GAC crystal structure and solution experiments suggest potassium at this site. Both crystal structures also depict two magnesium ions directly bound to the phosphate groups comprising this controversial pocket. Here, we used classical molecular dynamics simulations as well as umbrella sampling to investigate the possibility of binding of potassium versus magnesium inside the pocket and to better characterize the chelation of one of the binding magnesium ions outside the pocket. The results support the preference of the pocket to accommodate potassium rather than magnesium and suggest that one of the closely binding magnesium ions can only bind at high magnesium concentrations, such as might be present during crystallization. This work illustrates the complementary utility of molecular modeling approaches with atomic-level detail in resolving discrepancies between conflicting experimental results. PMID:27983843

Computational Assessment of Potassium and Magnesium Ion Binding to a Buried Pocket in GTPase-Associating Center RNA.

PubMed

Hayatshahi, Hamed S; Roe, Daniel R; Galindo-Murillo, Rodrigo; Hall, Kathleen B; Cheatham, Thomas E

2017-01-26

An experimentally well-studied model of RNA tertiary structures is a 58mer rRNA fragment, known as GTPase-associating center (GAC) RNA, in which a highly negative pocket walled by phosphate oxygen atoms is stabilized by a chelated cation. Although such deep pockets with more than one direct phosphate to ion chelation site normally include magnesium, as shown in one GAC crystal structure, another GAC crystal structure and solution experiments suggest potassium at this site. Both crystal structures also depict two magnesium ions directly bound to the phosphate groups comprising this controversial pocket. Here, we used classical molecular dynamics simulations as well as umbrella sampling to investigate the possibility of binding of potassium versus magnesium inside the pocket and to better characterize the chelation of one of the binding magnesium ions outside the pocket. The results support the preference of the pocket to accommodate potassium rather than magnesium and suggest that one of the closely binding magnesium ions can only bind at high magnesium concentrations, such as might be present during crystallization. This work illustrates the complementary utility of molecular modeling approaches with atomic-level detail in resolving discrepancies between conflicting experimental results.

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.

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.

Final Technical Report: Targeting DOE-Relevant Ions with Supramolecular Strategies, DE-SC0010555

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

Bowman-James, Kristin

The effectiveness of three popular supramolecular strategies to selectively target negatively charged ions (anions) was evaluated. Ions of interest included oxo anions, particularly sulfate, that hamper nuclear waste remediation. Three objectives were pursued using a simple building block strategies and by strategically placing anion-binding sites at appropriate positions on organic host molecules. The goal of the first objective was to assess the influence of secondary, tertiary and quaternized amines on binding tetrahedral anions using mixed amide/amine macrocyclic and urea/amine hosts containing aromatic or heteroaromatic spacers. Objective 2 focused on the design of ion pair hosts, using mixed macrocyclic anion hostsmore » joined through polyether linkages. Objective 3 was to explore the synthesis of new metal-linked extended macrocyclic frameworks to leverage anion binding. Key findings were that smaller 24-membered macrocycles provided the most complementary binding for sulfate ion and mixed urea/amine chelates showed enhanced binding over amide corollaries in addition to being highly selective for SO 4 2- in the presence of small quantities of water. In addition to obtaining prototype metal-linked macrocyclic anion hosts, a new dipincer ligand was designed that can be used to link macrocyclic or other supramolecular hosts in extended frameworks. When the tetraamide-based pincers are bound to two metal ions, an interesting phenomenon occurs. Upon deprotonation of the amides, two new protons appear between adjacent carbonyl pairs on the ligand, which may modify the chemistry, and metal-metal interactions in the complexes. Gel formation occurred for some of these extended hosts, and the physical properties are currently under investigation. The new tetracarboxamide-based pincers can also provide basic frameworks for double macrocycles capable of binding ion pairs as well as for binding metal ions and exploring intermetallic interactions through the pyrazine π system. Additionally appendages capable of influencing solvation effects can be introduced, and a number of other potential applications can be realized in areas such as soft materials chemistry, catalysis, sensing, and proton switches, the latter for binding and release of targeted guests. These findings provide a better foundation for understanding the selective binding of anions by targeted placement of hydrogen binding sites, and the strengths and weaknesses of various functional groups, that will allow for more the design of more effective anion sequestering agents. Our design strategy also used simple, cost-effective building blocks for host synthesis to allow for scale-up should real-world applications be forthcoming.« less

Mechanism of potassium ion uptake by the Na+/K+-ATPase

PubMed Central

Castillo, Juan P.; Rui, Huan; Basilio, Daniel; Das, Avisek; Roux, Benoît; Latorre, Ramon; Bezanilla, Francisco; Holmgren, Miguel

2015-01-01

The Na+/K+-ATPase restores sodium (Na+) and potassium (K+) electrochemical gradients dissipated by action potentials and ion-coupled transport processes. As ions are transported, they become transiently trapped between intracellular and extracellular gates. Once the external gate opens, three Na+ ions are released, followed by the binding and occlusion of two K+ ions. While the mechanisms of Na+ release have been well characterized by the study of transient Na+ currents, smaller and faster transient currents mediated by external K+ have been more difficult to study. Here we show that external K+ ions travelling to their binding sites sense only a small fraction of the electric field as they rapidly and simultaneously become occluded. Consistent with these results, molecular dynamics simulations of a pump model show a wide water-filled access channel connecting the binding site to the external solution. These results suggest a mechanism of K+ gating different from that of Na+ occlusion. PMID:26205423

Comparison of the bonding between ML(+) and ML2(+) (M = metal, L = noble gas)

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Partridge, Harry; Langhoff, Stephen R.

1990-01-01

Ab initio calculations are reported of the spectroscopic constants for the low-lying states of the molecular ions ML2(+), where M = Li, Na, Mg, V, Fe, Co, Ni and Cu, and where L is usually Ar. Comparison with existing analogous calculations on the ML(+) ions shows how the bonding and binding energy change with the addition of a second noble gas atom. The second binding energy is predicted to be essentially the same as the first for the Li, Na, Mg, and V ions, but larger for the Fe, Co, Ni and Cu ions. The binding energies of the transition metal noble gas ions are not accurately predicted at the SCF level, because correlation is required to describe their M(0)Ln(+) character. All trends can be explained in terms of promotion and hybridization on the metal ion.

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

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

Carbon dots as fluorescent probes for "off-on" detection of Cu2+ and L-cysteine in aqueous solution.

PubMed

Zong, Jie; Yang, Xiaoling; Trinchi, Adrian; Hardin, Simon; Cole, Ivan; Zhu, Yihua; Li, Chunzhong; Muster, Tim; Wei, Gang

2014-01-15

Copper ion (Cu(2+)) and L-cysteine (L-Cys) detection is critically important since an abnormal level of Cu(2+) or L-Cys is an indicator for many diseases. In this paper, we demonstrate an "off-on" approach for highly sensitive and selective detection of Cu(2+) and L-Cys using carbon dots (CDs) as fluorescent probes. CDs were prepared by using mesoporous silica (MS) spheres as nanoreactors. The binding ability of CDs towards metal ions was examined by comparing the fluorescence intensities of CDs before and after the addition of the metal ions. The addition of Cu(2+) cations leads to their absorption on the surface of CDs and the significant fluorescence quench of CDs (turn-off). The resulting in CDs-Cu(2+) system was found to be sensitive to L-Cys. The addition of L-Cys not only serves to shelter the CDs effectively from being quenched, but also to reverse the quenching and restore the fluorescence (turn-on) due to its ability to remove Cu(2+) from the surface of CDs. This method is facile, rapid, low cost, and environment-friendly. A detection limit as low as 2.3×10(-8) M for Cu(2+) and 3.4×10(-10) M for L-Cys is obtained, which is promising for biological applications. © 2013 Elsevier B.V. All rights reserved.

The effects of para-chloromercuribenzoic acid and different oxidative and sulfhydryl agents on a novel, non-AT1, non-AT2 angiotensin binding site identified as neurolysin

PubMed Central

Santos, Kira L.; Vento, Megan A; Wright, John W.; Speth, Robert C.

2013-01-01

A novel, non-AT1, non-AT2 brain binding site for angiotensin peptides that is unmasked by p-chloromercuribenzoate (PCMB) has been identified as a membrane associated variant of neurolysin. The ability of different organic and inorganic oxidative and sulfhydryl reactive agents to unmask or inhibit 125I-Sar1Ile8 angiotensin II (SI-Ang II) binding to this site was presently examined. In tissue membranes from homogenates of rat brain and testis incubated in assay buffer containing losartan (10 μM) and PD123319 (10 μM) plus 100 μM PCMB, 5 of the 39 compounds tested inhibited 125I-SI Ang II binding in brain and testis. Mersalyl acid, mercuric chloride (HgCl2) and silver nitrate (AgNO3) most potently inhibited 125I-SI Ang II binding with IC50’s ~1–20 μM This HgCl2 inhibition was independent of any interaction of HgCl2 with angiotensin II (Ang II) based on the lack of effect of HgCl2 on the dipsogenic effects of intracerebroventricularly administered Ang II and 125I-SI Ang II binding to AT1 receptors in the liver. Among sulfhydryl reagents, cysteamine and reduced glutathione (GSH), but not oxidized glutathione (GSSG) up to 1 mM, inhibited PCMB-unmasked 125I-SI Ang II binding in brain and testis. Thimerosal and 4-hydroxymercuribenzoate moderately inhibited PCMB-unmasked 125I-SI Ang II binding in brain and testis at 100 μM; however, they also unmasked non-AT1, non-AT2 binding independent of PCMB. 4-hydroxybenzoic acid did not promote 125 I-SI Ang II binding to this binding site indicating that only specific organomercurial compounds can unmask the binding site. The common denominator for all of these interacting substances is the ability to bind to protein cysteine sulfur. Comparison of cysteines between neurolysin and the closely related enzyme thimet oligopeptidase revealed an unconserved cysteine (cys650, based on the full length variant) in the proposed ligand binding channel (Brown et al., 2001) [1] near the active site of neurolysin. It is proposed that the mercuric ion in PCMB and closely related organomercurial compounds binds to cys650, while the acidic anion forms an ionic bond with a nearby arginine or lysine along the channel to effect a conformational change in neurolysin that promotes Ang II binding. PMID:23511333

Reversible cobalt ion binding to imidazole-modified nanopipettes

PubMed Central

Sa, Niya; Fu, Yaqin; Baker, Lane A.

2010-01-01

In this report, we demonstrate that quartz nanopipettes modified with an imidazole-terminated silane respond to metal ions (Co2+) in solution. The response of nanopipettes is evaluated through examination of the ion current rectification response. By cycling nanopipettes between solutions of different pH, adsorbed Co2+ can be released from the nanopipette surface, to regenerate binding sites of the nanopipette. These results demonstrate that rectification-based sensing strategies for nanopore sensors can benefit from selection of recognition elements with intermediate binding affinities, such that reversible responses to be attained. PMID:21090777

Reversible cobalt ion binding to imidazole-modified nanopipettes.

PubMed

Sa, Niya; Fu, Yaqin; Baker, Lane A

2010-12-15

In this report, we demonstrate that quartz nanopipettes modified with an imidazole-terminated silane respond to metal ions (Co(2+)) in solution. The response of nanopipettes is evaluated through examination of the ion current rectification ratio. When nanopipettes are cycled between solutions of different pH, adsorbed Co(2+) can be released from the nanopipette surface, to regenerate binding sites of the nanopipette. These results demonstrate that rectification-based sensing strategies for nanopore sensors can benefit from selection of recognition elements with intermediate binding affinities, such that reversible responses can be attained.

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.

Flavonoids-Rich Orthosiphon stamineus Extract as New Candidate for Angiotensin I-Converting Enzyme Inhibition: A Molecular Docking Study.

PubMed

Shafaei, Armaghan; Sultan Khan, Md Shamsuddin; F A Aisha, Abdalrahim; Abdul Majid, Amin Malik Shah; Hamdan, Mohammad Razak; Mordi, Mohd Nizam; Ismail, Zhari

2016-11-09

This study aims to evaluate the in vitro angiotensin-converting enzyme (ACE) inhibition activity of different extracts of Orthosiphon stamineus (OS) leaves and their main flavonoids, namely rosmarinic acid (RA), sinensetin (SIN), eupatorin (EUP) and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone (TMF). Furthermore, to identify possible mechanisms of action based on structure-activity relationships and molecular docking. The in vitro ACE inhibition activity relied on determining hippuric acid (HA) formation from ACE-specific substrate (hippuryl-histidyl-leucine (HHL)) by the action of ACE enzyme. A High Performance Liquid Chromatography method combined with UV detection was developed and validated for measurement the concentration of produced HA. The chelation ability of OS extract and its reference compounds was evaluated by tetramethylmurexide reagent. Furthermore, molecular docking study was performed by LeadIT-FlexX : BioSolveIT's LeadIT program. OS ethanolic extract (OS-E) exhibited highest inhibition and lowest IC 50 value (45.77 ± 1.17 µg/mL) against ACE compared to the other extracts. Among the tested reference compounds, EUP with IC 50 15.35 ± 4.49 µg/mL had highest inhibition against ACE and binding ability with Zn (II) (56.03% ± 1.26%) compared to RA, TMF and SIN. Molecular docking studies also confirmed that flavonoids inhibit ACE via interaction with the zinc ion and this interaction is stabilized by other interactions with amino acids in the active site. In this study, we have demonstrated that changes in flavonoids active core affect their capacity to inhibit ACE. Moreover, we showed that ACE inhibition activity of flavonoids compounds is directly related to their ability to bind with zinc ion in the active site of ACE enzyme. It was also revealed that OS extract contained high amount of flavonoids other than RA, TMF, SIN and EUP. As such, application of OS extract is useful as inhibitors of ACE.

Identification of two pentatricopeptide repeat genes required for RNA editing and zinc binding by C-terminal cytidine deaminase-like domains.

PubMed

Hayes, Michael L; Giang, Karolyn; Berhane, Beniam; Mulligan, R Michael

2013-12-20

Many transcripts expressed from plant organelle genomes are modified by C-to-U RNA editing. Nuclear encoded pentatricopeptide repeat (PPR) proteins are required as RNA binding specificity determinants in the RNA editing mechanism. Bioinformatic analysis has shown that most of the Arabidopsis PPR proteins necessary for RNA editing events include a C-terminal portion that shares structural characteristics with a superfamily of deaminases. The DYW deaminase domain includes a highly conserved zinc binding motif that shares characteristics with cytidine deaminases. The Arabidopsis PPR genes, ELI1 and DOT4, both have DYW deaminase domains and are required for single RNA editing events in chloroplasts. The ELI1 DYW deaminase domain was expressed as a recombinant protein in Escherichia coli and was shown to bind two zinc atoms per polypeptide. Thus, the DYW deaminase domain binds a zinc metal ion, as expected for a cytidine deaminase, and is potentially the catalytic component of an editing complex. Genetic complementation experiments demonstrate that large portions of the DYW deaminase domain of ELI1 may be eliminated, but the truncated genes retain the ability to restore editing site conversion in a mutant plant. These results suggest that the catalytic activity can be supplied in trans by uncharacterized protein(s) of the editosome.

[Structure-functional organization of eukaryotic high-affinity copper importer CTR1 determines its ability to transport copper, silver and cisplatin].

PubMed

Skvortsov, A N; Zatulovskiĭ, E A; Puchkova, L V

2012-01-01

It was shown recently, that high affinity Cu(I) importer eukaryotic protein CTR1 can also transport in vitro abiogenic Ag(I) ions and anticancer drug cisplatin. At present there is no rational explanation how CTR1 can transfer platinum group, which is different by coordination properties from highly similar Cu(I) and Ag(I). To understand this phenomenon we analyzed 25 sequences of chordate CTR1 proteins, and found out conserved patterns of organization of N-terminal extracellular part of CTR1 which correspond to initial metal binding. Extracellular copper-binding motifs were qualified by their coordination properties. It was shown that relative position of Met- and His-rich copper-binding motifs in CTR1 predisposes the extracellular CTR1 part to binding of copper, silver and cisplatin. Relation between tissue-specific expression of CTR1 gene, steady-state copper concentration, and silver and platinum accumulation in organs of mice in vivo was analyzed. Significant positive but incomplete correlation exists between these variables. Basing on structural and functional peculiarities of N-terminal part of CTR1 a hypothesis of coupled transport of copper and cisplatin has been suggested, which avoids the disagreement between CTR1-mediated cisplatin transport in vitro, and irreversible binding of platinum to Met-rich peptides.

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

Theoretical study of the BeLi, BeNa, MgLi, MgNa, and AlBe molecules and their negative ions

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Langhoff, Stephen R.; Partridge, Harry

1992-01-01

The alkaline earth-alkali diatomics are found to have weak bonds, because the diffuse alkali valence s orbitals cannot form a bond of sufficient strength to pay the promotion energy of the alkaline-earth atoms. This leads to van der Waals bonding in the neutrals as well as the negative ions. In fact, the negative ions have larger binding energies than the neutrals as a result of the much larger polarizability of the negative ion. The binding energy of AlBe is significantly larger than the Be-alkali molecules, due to a covalent contribution to the bonding. The binding energy in AlBe(-) is considerably larger than AlBe; the binding energy of the X 3Sigma(-) state of AlBe(-) is computed to be 1.36 eV, as compared with 0.57 eV for the X 2Pi state of AlBe.

Cyanobacterial megamolecule sacran efficiently forms LC gels with very heavy metal ions.

PubMed

Okajima, Maiko K; Miyazato, Shinji; Kaneko, Tatsuo

2009-08-04

We extracted the megamolecular polysaccharide sacran, which contains carboxylate and sulfate groups, from the jellylike extracellular matrix (ECM) of the cyanobacterium Aphanothece sacrum, which has mineral adsorption bioactivity. We investigated the gelation properties of sacran binding with various heavy metal ions. The sacran chain adsorbed heavier metal ions such as indium, rare earth metals, and lead ions more efficiently to form gel beads. In addition, trivalent metal ions adsorbed onto the sacran chains more efficiently than did divalent ions. The investigation of the metal ion binding ratio on sacran chains demonstrated that sacran adsorbed gadolinium trivalent ions more efficiently than indium trivalent ions. Gel bead formation may be closely correlated to the liquid-crystalline organization of sacran.

A critical review of three methods used for the measurement of mercury (Hg2+)-dissolved organic matter stability constants

USGS Publications Warehouse

Gasper, J.D.; Aiken, G.R.; Ryan, J.N.

2007-01-01

Three experimental techniques - ion exchange, liquid-liquid extraction with competitive ligand exchange, and solid-phase extraction with competitive ligand exchange (CLE-SPE) - were evaluated as methods for determining conditional stability constants (K) for the binding of mercury (Hg2+) to dissolved organic matter (DOM). To determine the utility of a given method to measure stability constants at environmentally relevant experimental conditions, experimental results should meet three criteria: (1) the data must be experimentally valid, in that they were acquired under conditions that meet all the requirements of the experimental method, (2) the Hg:DOM ratio should be determined and it should fall within levels that are consistent with environmental conditions, and (3) the stability constants must fall within the detection window of the method. The ion exchange method was found to be limited by its detection window, which constrains the method to stability constants with log K values less than about 14. The liquid-liquid extraction method was found to be complicated by the ability of Hg-DOM complexes to partition into the organic phase. The CLE-SPE method was found to be the most suitable of these methods for the measurement of Hg-DOM stability constants. Stability constants for DOM isolates measured using the CLE-SPE method at environmentally relevant Hg:DOM ratios were log K = 25-30 (M-1). These values are consistent with the strong Hg2+ binding expected for reduced S-containing binding sites. ?? 2007 Elsevier Ltd. All rights reserved.

Multiply Reduced Oligofluorenes: Their Nature and Pairing with THF-Solvated Sodium Ions

DOE PAGES

Wu, Qin; Zaikowski, Lori; Kaur, Parmeet; ...

2016-07-01

Conjugated oligofluorenes are chemically reduced up to five charges in tetrahydrofuran solvent and confirmed with clear spectroscopic evidence. Stimulated by these experimental results, we have conducted a comprehensive computational study of the electronic structure and the solvation structure of representative oligofluorene anions with a focus on the pairing between sodium ions and these multianions. In addition, using density functional theory (DFT) methods and a solvation model of both explicit solvent molecules and implicit polarizable continuum, we first elucidate the structure of tightly solvated free sodium ions, and then explore the pairing of sodium ions either in contact with reduced oligofluorenesmore » or as solvent-separated ion pairs. Computed time-dependent-DFT absorption spectra are compared with experiments to assign the dominant ion pairing structure for each multianion. Computed ion pair binding energies further support our assignment. Lastly, the availability of different length and reducing level of oligofluorenes enables us to investigate the effects of total charge and charge density on the binding with sodium ions, and our results suggest both factors play important roles in ion pairing for small molecules. However, as the oligofluorene size grows, its charge density determines the binding strength with the sodium ion.« less

Essential slow degrees of freedom in protein-surface simulations: A metadynamics investigation.

PubMed

Prakash, Arushi; Sprenger, K G; Pfaendtner, Jim

2018-03-29

Many proteins exhibit strong binding affinities to surfaces, with binding energies much greater than thermal fluctuations. When modelling these protein-surface systems with classical molecular dynamics (MD) simulations, the large forces that exist at the protein/surface interface generally confine the system to a single free energy minimum. Exploring the full conformational space of the protein, especially finding other stable structures, becomes prohibitively expensive. Coupling MD simulations with metadynamics (enhanced sampling) has fast become a common method for sampling the adsorption of such proteins. In this paper, we compare three different flavors of metadynamics, specifically well-tempered, parallel-bias, and parallel-tempering in the well-tempered ensemble, to exhaustively sample the conformational surface-binding landscape of model peptide GGKGG. We investigate the effect of mobile ions and ion charge, as well as the choice of collective variable (CV), on the binding free energy of the peptide. We make the case for explicitly biasing ions to sample the true binding free energy of biomolecules when the ion concentration is high and the binding free energies of the solute and ions are similar. We also make the case for choosing CVs that apply bias to all atoms of the solute to speed up calculations and obtain the maximum possible amount of information about the system. Copyright © 2017 Elsevier Inc. All rights reserved.

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.

Dynamics and lithium binding energies of polyelectrolytes based on functionalized poly(para-phenylene terephthalamide).

PubMed

Grozema, F C; Best, A S; van Eijck, L; Stride, J; Kearley, G J; de Leeuw, S W; Picken, S J

2005-04-28

Polyelectrolyte materials are an interesting class of electrolytes for use in fuel cell and battery applications. Poly(para-phenylene terephthalamide) (PPTA, Kevlar) is a liquid crystalline polymer that, when sulfonated, is a polyelectrolyte that exhibits moderate ion conductivity at elevated temperatures. In this work, quasi-elastic neutron scattering (QENS) experiments were performed to gain insight into the effect of the presence of lithium counterions on the chain dynamics in the material. It was found that the addition of lithium ions decreases the dynamics of the chains. Additionally, the binding of lithium ions to the sulfonic acids groups was investigated by density functional theory (DFT) calculations. It was found that the local surroundings of the sulfonic acid group have very little effect on the lithium-ion binding energy. Binding energies for a variety of different systems were all calculated to be around 150 kcal/mol. The DFT calculations also show the existence of a structure in which a single lithium ion interacts with two sulfonic acid moieties on different chains. The formation of such "electrostatic cross-links" is believed to be the source of the increased tendency to aggregate and the reduced dynamics in the presence of lithium ions.

Phosphorylation-Dependent Regulation of Ryanodine Receptors

PubMed Central

Marx, Steven O.; Reiken, Steven; Hisamatsu, Yuji; Gaburjakova, Marta; Gaburjakova, Jana; Yang, Yi-Ming; Rosemblit, Nora; Marks, Andrew R.

2001-01-01

Ryanodine receptors (RyRs), intracellular calcium release channels required for cardiac and skeletal muscle contraction, are macromolecular complexes that include kinases and phosphatases. Phosphorylation/dephosphorylation plays a key role in regulating the function of many ion channels, including RyRs. However, the mechanism by which kinases and phosphatases are targeted to ion channels is not well understood. We have identified a novel mechanism involved in the formation of ion channel macromolecular complexes: kinase and phosphatase targeting proteins binding to ion channels via leucine/isoleucine zipper (LZ) motifs. Activation of kinases and phosphatases bound to RyR2 via LZs regulates phosphorylation of the channel, and disruption of kinase binding via LZ motifs prevents phosphorylation of RyR2. Elucidation of this new role for LZs in ion channel macromolecular complexes now permits: (a) rapid mapping of kinase and phosphatase targeting protein binding sites on ion channels; (b) predicting which kinases and phosphatases are likely to regulate a given ion channel; (c) rapid identification of novel kinase and phosphatase targeting proteins; and (d) tools for dissecting the role of kinases and phosphatases as modulators of ion channel function. PMID:11352932

Counter-ion binding and mobility in the presence of hydrophobic polyions – combining molecular dynamics simulations and NMR

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

Druchok, Maksym; Malikova, Natalie; Rollet, Anne-Laure

Counter-ion binding and mobility in aqueous solutions of partially hydrophobic ionene oligoions is studied here by a combination of all-atomic molecular dynamics (MD) simulations and NMR ({sup 19}F and {sup 81}Br nuclei) measurements. We present results for 12, 12–ionenes in the presence of different halide ions (F{sup −}, Cl{sup −}, Br{sup −} and I{sup −}), as well as their mixtures; the latter allowing us to probe counter-ion selectivity of these oligoions. We consolidate both structural and dynamic information, in particular simulated radial distribution functions and average residence times of counter-ions in the vicinity of ionenes and NMR data in themore » form of counter-ion chemical shift and self-diffusion coefficients. On one hand, previously reported enthalpy of dilution and mixing measurements show a reverse counter-ion sequence for 12, 12–ionenes with respect to their less hydrophobic 3, 3– and 6, 6– analogues. On the other hand, the current MD and NMR data, reflecting the counter-ion binding tendencies to the ionene chain, give evidence for the same ordering as that observed by MD for 3, 3–ionenes. This is not seen as a contradiction and can be rationalized on the basis of increasing chain hydrophobicity, which has different consequences for enthalpy and ion-binding. The latter is reflecting free energy changes and as such includes both enthalpic and entropic contributions.« less

Conformation of the Phosphate D-alanine Zwitterion in Bacterial Teichoic Acid from Nuclear Magnetic Resonance Spectroscopy

PubMed Central

Garimella, Ravindranath; Halye, Jeffrey L.; Harrison, William; Klebba, Phillip E.; Rice, Charles V.

2009-01-01

The conformation of D-alanine (D-Ala) groups of bacterial teichoic acid is a central, yet untested, paradigm of microbiology. The D-Ala binds via the C-terminus, thereby allowing the amine to exist as a free cationic NH3+ group with the ability to form a contact-ion-pair with the nearby anionic phosphate group. This conformation hinders metal chelation by the phosphate because the zwitterion pair is charge neutral. To the contrary, the repulsion of cationic antimicrobial peptides (CAMPs) is attributed to the presence of the D-Ala cation; thus the ion-pair does not form in this model. Solid-state nuclear magnetic resonance (NMR) spectroscopy has been used to measure the distance between amine and phosphate groups within cell wall fragments of Bacillus subtilis. The bacteria were grown on media containing 15N D-Ala and β-chloroalanine racemase inhibitor. The rotational-echo double-resonance (REDOR) pulse sequence was used to measure the internuclear dipolar coupling and the results demonstrate: 1) the metal-free amine-to-phosphate distance is 4.4 Å and 2) the amine-to-phosphate distance increases to 5.4 Å in the presence of Mg2+ ions. As a result, the zwitterion exists in a nitrogen-oxygen ion-pair configuration providing teichoic acid with a positive charge to repel CAMPs. Additionally, the amine of D-Ala does not prevent magnesium chelation in contradiction to the prevailing view of teichoic acids in metal binding. Thus, the NMR-based description of teichoic acid structure resolves the contradictory models, advances the basic understanding of cell wall biochemistry, and provides possible insight into the creation of new antibiotic therapies. PMID:19746945

Fluorescence fingerprints and Cu2+-complexing ability of individual molecular size fractions in soil- and waste-borne DOM.

PubMed

Knoth de Zarruk, K; Scholer, G; Dudal, Y

2007-09-01

Land spreading of organic materials introduces large amounts of dissolved organic matter (DOM) into the soil. DOM has the ability to form stable complexes with heavy metals and can facilitate their transport towards the groundwater. The effects on soil processes are difficult to assess, because different DOM components might react differently towards metal ions. The objective of this study was to investigate the fluorescence signature and the Cu2+-binding capacity of individual molecular size fractions of DOM from various sources. DOM extracted from leaf compost, chicken manure, sugar cane vinasse and a fulvic hypercalcaric cambisol was fractionated by the means of dialysis into four molecular size classes: MW<500, 50012000-14000 Da. Vinasse and leaf compost contained around 80% and 70%, respectively, of the total organic carbon in the fractions with low molecular weight (MW<3500 Da); in chicken manure and soil these fractions accounted for 40% and 50% only. Fluorescence was highest in the fraction MW>12000 Da for leaf compost, chicken manure and soil. The opposite result was obtained for vinasse, where the fractions with low molecular weight showed highest fluorescence intensities, distinguishing it from all other samples. Vinasse showed the greatest ability to bind Cu2+ with a resulting complex concentration of 6.31mgl(-1) while in contact with an excess of Cu2+. Leaf compost, soil and chicken manure followed with 2.69, 1.12, and 0.85mgl(-1), respectively. Within vinasse, the fraction MW<500 Da was able to form the most DOM-Cu complexes, indicating the importance of low molecular weight fractions in metal binding.

Ligand binding to telomeric G-quadruplex DNA investigated by funnel-metadynamics simulations

PubMed Central

Moraca, Federica; Amato, Jussara; Ortuso, Francesco; Artese, Anna; Novellino, Ettore; Alcaro, Stefano; Parrinello, Michele; Limongelli, Vittorio

2017-01-01

G-quadruplexes (G4s) are higher-order DNA structures typically present at promoter regions of genes and telomeres. Here, the G4 formation decreases the replicative DNA at each cell cycle, finally leading to apoptosis. The ability to control this mitotic clock, particularly in cancer cells, is fascinating and passes through a rational understanding of the ligand/G4 interaction. We demonstrate that an accurate description of the ligand/G4 binding mechanism is possible using an innovative free-energy method called funnel-metadynamics (FM), which we have recently developed to investigate ligand/protein interaction. Using FM simulations, we have elucidated the binding mechanism of the anticancer alkaloid berberine to the human telomeric G4 (d[AG3(T2AG3)3]), computing also the binding free-energy landscape. Two ligand binding modes have been identified as the lowest energy states. Furthermore, we have found prebinding sites, which are preparatory to reach the final binding mode. In our simulations, the ions and the water molecules have been explicitly represented and the energetic contribution of the solvent during ligand binding evaluated. Our theoretical results provide an accurate estimate of the absolute ligand/DNA binding free energy (ΔGb0 = −10.3 ± 0.5 kcal/mol) that we validated through steady-state fluorescence binding assays. The good agreement between the theoretical and experimental value demonstrates that FM is a most powerful method to investigate ligand/DNA interaction and can be a useful tool for the rational design also of G4 ligands. PMID:28232513

Extraction of mercury from groundwater using immobilized algae.

PubMed

Barkley, N P

1991-10-01

Bio-Recovery Systems, Inc. conducted a project under the Emerging Technology portion of the United States Environmental Protection Agency's (EPAs) Superfund Innovative Technology Evaluation (SITE) Program to evaluate the ability of immobilized algae to adsorb mercury from contaminated groundwater in laboratory studies and pilot-scale field tests. Algal biomass was incorporated in a permeable polymeric matrix. The product, AlgaSORB, packed into adsorption columns, exhibited excellent flow characteristics, and functioned as a "biological" ion exchange resin. A sequence of eleven laboratory tests demonstrated the ability of this product to adsorb mercury from groundwater that contained high levels of total dissolved solids and hard water components. However, use of a single AlgaSORB preparation yielded nonrepeatable results with samples collected at different times of the year. The strategy of sequentially extracting the groundwater through two columns containing different preparations of AlgaSORB was developed and proved successful in laboratory and pilot-scale field tests. Field test results indicate that AlgaSORB could be economically competitive with ion exchange resins for removal of mercury, with the advantage that hardness and other dissolved solids do not appear to compete with heavy metals for binding capacity.

Effect of ionic strength on the interfacial viscoelasticity and stability of silk fibroin at the oil/water interface.

PubMed

Tang, Xiaoxiao; Qiao, Xiuying; Miller, Reinhard; Sun, Kang

2016-12-01

The amphiphilic character and surface activity endows silk fibroin with the ability to reside at fluid interfaces and effectively stabilize emulsions. However, the influence of relevant factors and their actual effect on the interfacial viscoelasticity and stability of silk fibroin at the oil/water interface has received less attention. In the present study, the effect of ionic strength on the interfacial viscoelasticity, emulsification effectiveness and stability of silk fibroin at the oil/water interface was investigated in detail. A higher ion concentration facilitates greater adsorption, stronger molecular interaction and faster structure reorganization of silk fibroin at the oil/water interface, thus causing quicker interfacial saturation adsorption, greater interfacial strength and lower interfacial structural fracture on large deformation. However, the presence of concentrated ions screens the charges in silk fibroin molecules and the zeta potential decreases as a result of electrostatic screening and ion-binding effects, which may result in emulsion droplet coalescence and a decrease in emulsion stability. The positively-charged ions significantly affect the interfacial elasticity and stability of silk fibroin layers at the oil/water interface as a result of the strong electrostatic interactions between counter-ions and the negatively-charged groups of silk fibroin. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Induced binding of proteins by ammonium sulfate in affinity and ion-exchange column chromatography.

PubMed

Arakawa, Tsutomu; Tsumoto, Kouhei; Ejima, Daisuke; Kita, Yoshiko; Yonezawa, Yasushi; Tokunaga, Masao

2007-04-10

In general, proteins bind to affinity or ion-exchange columns at low salt concentrations, and the bound proteins are eluted by raising the salt concentration, changing the solvent pH, or adding competing ligands. Blue-Sepharose is often used to remove bovine serum albumin (BSA) from samples, but when we applied BSA to Blue-Sepharose in 20 mM phosphate, pH 7.0, 50%-60% of the protein flowed through the column; however, complete binding of BSA was achieved by the addition of 2 M ammonium sulfate (AS) to the column equilibration buffer and the sample. The bound protein was eluted by decreasing the AS concentration or by adding 1 M NaCl or arginine. AS at high concentrations resulted in binding of BSA even to an ion-exchange column, Q-Sepharose, at pH 7.0. Thus, although moderate salt concentrations elute proteins from Blue-Sepharose or ion-exchange columns, proteins can be bound to these columns under extreme salting-out conditions. Similar enhanced binding of proteins by AS was observed with an ATP-affinity column.

A Cr(VI) selective probe based on a quinoline-amide calix[4]arene

NASA Astrophysics Data System (ADS)

Ferreira, Juliane F.; Bagatin, Izilda A.

2018-01-01

A new quinoline-amide calix[4]arene 3-receptor for detection of hazardous anions and cations have been synthesized. The 3-receptor was examined for its sensing properties towards several different anions (Cr2O72 -, SCN-, F-, Cl-, NO3-) and metal ions (Hg2+, Cd2+, Ag+) by UV-vis and fluorescence spectroscopies. It was detected that the 3-receptor has only sensing ability for Cr2O72 - and Hg2+ ions, resulting in the association constants higher for Cr2O72 - than to the Hg2+ ions. High selectivity towards Cr2O72 - were also observed by fluorescence measurement among other ions (F-, Cl-, SCN-, Hg2+, Cd2+, Ag+) with a low limit of detection (7.36 × 10-6 mol dm-3). Proton NMR anion-binding investigations revealed a strong interaction of Cr2O72 - anion with NH and CH groups of the receptor, showing that the combination with hydrogen-bonds donor groups strengthened the anion receptor association. Furthermore, remarkable association constants for dichromate anion obtained by all techniques strongly suggest the 3-receptor as a selective Cr(VI) sensor.

A Cr(VI) selective probe based on a quinoline-amide calix[4]arene.

PubMed

Ferreira, Juliane F; Bagatin, Izilda A

2018-01-15

A new quinoline-amide calix[4]arene 3-receptor for detection of hazardous anions and cations have been synthesized. The 3-receptor was examined for its sensing properties towards several different anions (Cr 2 O 7 2- , SCN - , F - , Cl - , NO 3 - ) and metal ions (Hg 2+ , Cd 2+ , Ag + ) by UV-vis and fluorescence spectroscopies. It was detected that the 3-receptor has only sensing ability for Cr 2 O 7 2- and Hg 2+ ions, resulting in the association constants higher for Cr 2 O 7 2- than to the Hg 2+ ions. High selectivity towards Cr 2 O 7 2- were also observed by fluorescence measurement among other ions (F - , Cl - , SCN - , Hg 2+ , Cd 2+ , Ag + ) with a low limit of detection (7.36×10 -6 moldm -3 ). Proton NMR anion-binding investigations revealed a strong interaction of Cr 2 O 7 2- anion with NH and CH groups of the receptor, showing that the combination with hydrogen-bonds donor groups strengthened the anion receptor association. Furthermore, remarkable association constants for dichromate anion obtained by all techniques strongly suggest the 3-receptor as a selective Cr(VI) sensor. Copyright © 2017 Elsevier B.V. All rights reserved.

A zinc fluorescent sensor used to detect mercury (II) and hydrosulfide.

PubMed

Jung, Jae Min; Lee, Jae Jun; Nam, Eunju; Lim, Mi Hee; Kim, Cheal; Harrison, Roger G

2017-05-05

A zinc sensor based on quinoline and morpholine has been synthesized. The sensor selectively fluoresces in the presence of Zn 2+ , while not for other metal ions. Absorbance changes in the 350nm region are observed when Zn 2+ binds, which binds in a 1:1 ratio. The sensor fluoresces due to Zn 2+ above pH values of 6.0 and in the biological important region. The Zn 2+ -sensor complex has the unique ability to detect both Hg 2+ and HS - . The fluorescence of the Zn 2+ -sensor complex is quenched when it is exposed to aqueous solutions of Hg 2+ with sub-micromolar detection levels for Hg 2+ . The fluorescence of the Zn 2+ -sensor complex is also quenched by aqueous solutions of hydrosulfide. The sensor was used to detect Zn 2+ and Hg 2+ in living cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Spectroscopic and molecular modeling studies of the interaction between cytidine and human serum albumin and its analytical application.

PubMed

Cui, Fengling; Wang, Junli; Yao, Xiaojun; Wang, Li; Zhang, Qiangzhai; Qu, Guirong

In this study, the interaction between cytidine and human serum albumin (HSA) was investigated for the first time by fluorescence spectroscopy in combination with UV absorption spectrum and molecular modeling under simulative physiological conditions. Experimental results indicated that cytidine had a strong ability to quench the intrinsic fluorescence of human serum albumin. The binding constants (K) at different temperatures, thermodynamic parameter enthalpy changes (DeltaH) and entropy changes (DeltaS) of HSA-cytidine had been calculated according to the relevant fluorescence data, which indicated that the hydrophobic and electrostatic interactions played a major role, which was in agreement with the results of molecular modeling study. In addition, the effects of other ions on the binding constants were also studied. Furthermore, synchronous fluorescence technology was successfully applied to the determination of human serum albumin added into the cytidine solution.

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

Wu, Qin; Zaikowski, Lori; Kaur, Parmeet

Conjugated oligofluorenes are chemically reduced up to five charges in tetrahydrofuran solvent and confirmed with clear spectroscopic evidence. Stimulated by these experimental results, we have conducted a comprehensive computational study of the electronic structure and the solvation structure of representative oligofluorene anions with a focus on the pairing between sodium ions and these multianions. In addition, using density functional theory (DFT) methods and a solvation model of both explicit solvent molecules and implicit polarizable continuum, we first elucidate the structure of tightly solvated free sodium ions, and then explore the pairing of sodium ions either in contact with reduced oligofluorenesmore » or as solvent-separated ion pairs. Computed time-dependent-DFT absorption spectra are compared with experiments to assign the dominant ion pairing structure for each multianion. Computed ion pair binding energies further support our assignment. Lastly, the availability of different length and reducing level of oligofluorenes enables us to investigate the effects of total charge and charge density on the binding with sodium ions, and our results suggest both factors play important roles in ion pairing for small molecules. However, as the oligofluorene size grows, its charge density determines the binding strength with the sodium ion.« less

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.

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

Zinc-binding Domain of the Bacteriophage T7 DNA Primase Modulates Binding to the DNA Template*

PubMed Central

Lee, Seung-Joo; Zhu, Bin; Akabayov, Barak; Richardson, Charles C.

2012-01-01

The zinc-binding domain (ZBD) of prokaryotic DNA primases has been postulated to be crucial for recognition of specific sequences in the single-stranded DNA template. To determine the molecular basis for this role in recognition, we carried out homolog-scanning mutagenesis of the zinc-binding domain of DNA primase of bacteriophage T7 using a bacterial homolog from Geobacillus stearothermophilus. The ability of T7 DNA primase to catalyze template-directed oligoribonucleotide synthesis is eliminated by substitution of any five-amino acid residue-long segment within the ZBD. The most significant defect occurs upon substitution of a region (Pro-16 to Cys-20) spanning two cysteines that coordinate the zinc ion. The role of this region in primase function was further investigated by generating a protein library composed of multiple amino acid substitutions for Pro-16, Asp-18, and Asn-19 followed by genetic screening for functional proteins. Examination of proteins selected from the screening reveals no change in sequence-specific recognition. However, the more positively charged residues in the region facilitate DNA binding, leading to more efficient oligoribonucleotide synthesis on short templates. The results suggest that the zinc-binding mode alone is not responsible for sequence recognition, but rather its interaction with the RNA polymerase domain is critical for DNA binding and for sequence recognition. Consequently, any alteration in the ZBD that disturbs its conformation leads to loss of DNA-dependent oligoribonucleotide synthesis. PMID:23024359

Curcumin derivatives as metal-chelating agents with potential multifunctional activity for pharmaceutical applications.

PubMed

Ferrari, Erika; Benassi, Rois; Sacchi, Stefania; Pignedoli, Francesca; Asti, Mattia; Saladini, Monica

2014-10-01

Curcuminoids represent new perspectives for the development of novel therapeutics for Alzheimer's disease (AD), one probable mechanism of action is related to their metal complexing ability. In this work we examined the metal complexing ability of substituted curcuminoids to propose new chelating molecules with biological properties comparable with curcumin but with improved stability as new potential AD therapeutic agents. The K2T derivatives originate from the insertion of a -CH2COOC(CH3)3 group on the central atom of the diketonic moiety of curcumin. They retain the diketo-ketoenol tautomerism which is solvent dependent. In aqueous solution the prevalent form is the diketo one but the addition of metal ion (Ga(3+), Cu(2+)) causes the dissociation of the enolic proton creating chelate complexes and shifting the tautomeric equilibrium towards the keto-enol form. The formation of metal complexes is followed by both NMR and UV-vis spectroscopy. The density functional theory (DFT) calculations on K2T21 complexes with Ga(3+) and Cu(2+) are performed and compared with those on curcumin complexes. [Ga(K2T21)2(H2O)2](+) was found more stable than curcumin one. Good agreement is detected between calculated and experimental (1)H and (13)C NMR data. The calculated OH bond dissociation energy (BDE) and the OH proton dissociation enthalpy (PDE), allowed to predict the radical scavenging ability of the metal ion complexed with K2T21, while the calculated electronic affinity (EA) and ionization potential (IP) represent yardsticks of antioxidant properties. Eventually theoretical calculations suggest that the proton-transfer-associated superoxide-scavenging activity is enhanced after binding metal ions, and that Ga(3+) complexes display possible superoxide dismutase (SOD)-like activity. Copyright © 2014 Elsevier Inc. All rights reserved.

Iron and copper chelation by flavonoids: an electrospray mass spectrometry study.

PubMed

Fernandez, M Tereza; Mira, M Lurdes; Florêncio, M Helena; Jennings, Keith R

2002-11-11

Flavonoids are well known as effective free radical scavengers exhibiting therefore an antioxidant behaviour. Another antioxidant mechanism however may result from the ability they have to chelate metal ions, rendering them inactive to participate in free radical generating reactions. Electrospray mass spectrometry has been used to study metal ion interactions with a set of flavonoids from different classes. Complexes with a range of stoichiometries, of metal: flavonoid, 1:1, 1:2, 2:2, 2:3 have been observed. The stoichiometry 1:2 is in general the preferred one. It is established for flavones and for the flavanone naringenin that the binding metal sites are preferentially at the 5-hydroxyl and 4-oxo groups. Redox reactions are also observed through the change of the oxidation state of the metal, jointly with the oxidation of the flavonoid by loss of hydrogen. Structures of the oxidized species of some flavonoids are proposed.

Block copolymer with simultaneous electric and ionic conduction for use in lithium ion batteries

DOEpatents

Javier, Anna Esmeralda K; Balsara, Nitash Pervez; Patel, Shrayesh Naran; Hallinan, Jr., Daniel T

2013-10-08

Redox reactions that occur at the electrodes of batteries require transport of both ions and electrons to the active centers. Reported is the synthesis of a block copolymer that exhibits simultaneous electronic and ionic conduction. A combination of Grignard metathesis polymerization and click reaction was used successively to synthesize the block copolymer containing regioregular poly(3-hexylthiophene) (P3HT) and poly(ethylene oxide) (PEO) segments. The P3HT-PEO/LiTFSI mixture was then used to make a lithium battery cathode with LiFePO.sub.4 as the only other component. All-solid lithium batteries of the cathode described above, a solid electrolyte and a lithium foil as the anode showed capacities within experimental error of the theoretical capacity of the battery. The ability of P3HT-PEO to serve all of the transport and binding functions required in a lithium battery electrode is thus demonstrated.

Chemically functionalized gold nanoparticles: Synthesis, characterization, and applications

NASA Astrophysics Data System (ADS)

Daniel, Weston Lewis

This thesis focuses on the development and application of gold nanoparticle based detection systems and biomimetic structures. Each class of modified nanoparticle has properties that are defined by its chemical moieties that interface with solution and the gold nanoparticle core. In Chapter 2, a comparison of the biomolecular composition and binding properties of various preparations of antibody oligonucleotide gold nanoparticle conjugates is presented. These constructs differed significantly in terms of their structure and binding properties. Chapter 3 reports the use of electroless gold deposition as a light scattering signal enhancer in a multiplexed, microarray-based scanometric immunoassay using the gold nanoparticle probes evaluated in Chapter 2. The use of gold development results in greater signal enhancement than the typical silver development, and multiple rounds of metal development were found to increase the resulting signal compared to one development. Chapter 4 describes an amplified scanometric detection method for human telomerase activity. Gold nanoparticles functionalized with specific oligonucleotide sequences can efficiently capture telomerase enzymes and subsequently be elongated. Both the elongated and unmodified oligonucleotide sequences are simultaneously measured. At low telomerase concentrations, elongated strands cannot be detected, but the unmodified sequences, which come from the same probe particles, can be detected because their concentration is higher, providing a novel form of amplification. Chapter 5 reports the development of a novel colorimetric nitrite and nitrate ion assay based upon gold nanoparticle probes functionalized with Griess reaction reagents. This assay takes advantage of the distance-dependent plasmonic properties of the gold nanoparticles and the ability of nitrite ion to facilitate the cross coupling of novel nanoparticle probes. The assay works on the concept of a kinetic end point and can be triggered at the EPA limit for this ion in drinking water. Finally, Chapter 6 describes the synthesis of high density lipoprotein biomimetic nanoparticles capable of binding cholesterol. These structures use a gold nanoparticle core to template the assembly of a mixed phospholipid layer and the adsorption of apolipoprotein A-I. These synthesized structures have the general size and surface composition of natural HDL and bind free cholesterol with a Kd of 4 nM.

The Role and Specificity of the Catalytic and Regulatory Cation-binding Sites of the Na+-pumping NADH:Quinone Oxidoreductase from Vibrio cholerae*

PubMed Central

Juárez, Oscar; Shea, Michael E.; Makhatadze, George I.; Barquera, Blanca

2011-01-01

The Na+-translocating NADH:quinone oxidoreductase is the entry site for electrons into the respiratory chain and the main sodium pump in Vibrio cholerae and many other pathogenic bacteria. In this work, we have employed steady-state and transient kinetics, together with equilibrium binding measurements to define the number of cation-binding sites and characterize their roles in the enzyme. Our results show that sodium and lithium ions stimulate enzyme activity, and that Na+-NQR enables pumping of Li+, as well as Na+ across the membrane. We also confirm that the enzyme is not able to translocate other monovalent cations, such as potassium or rubidium. Although potassium is not used as a substrate, Na+-NQR contains a regulatory site for this ion, which acts as a nonessential activator, increasing the activity and affinity for sodium. Rubidium can bind to the same site as potassium, but instead of being activated, enzyme turnover is inhibited. Activity measurements in the presence of both sodium and lithium indicate that the enzyme contains at least two functional sodium-binding sites. We also show that the binding sites are not exclusively responsible for ion selectivity, and other steps downstream in the mechanism also play a role. Finally, equilibrium-binding measurements with 22Na+ show that, in both its oxidized and reduced states, Na+-NQR binds three sodium ions, and that the affinity for sodium is the same for both of these states. PMID:21652714

Influence of Na+ and Mg2+ ions on RNA structures studied with molecular dynamics simulations.

PubMed

Fischer, Nina M; Polêto, Marcelo D; Steuer, Jakob; van der Spoel, David

2018-06-01

The structure of ribonucleic acid (RNA) polymers is strongly dependent on the presence of, in particular Mg2+ cations to stabilize structural features. Only in high-resolution X-ray crystallography structures can ions be identified reliably. Here, we perform molecular dynamics simulations of 24 RNA structures with varying ion concentrations. Twelve of the structures were helical and the others complex folded. The aim of the study is to predict ion positions but also to evaluate the impact of different types of ions (Na+ or Mg2+) and the ionic strength on structural stability and variations of RNA. As a general conclusion Mg2+ is found to conserve the experimental structure better than Na+ and, where experimental ion positions are available, they can be reproduced with reasonable accuracy. If a large surplus of ions is present the added electrostatic screening makes prediction of binding-sites less reproducible. Distinct differences in ion-binding between helical and complex folded structures are found. The strength of binding (ΔG‡ for breaking RNA atom-ion interactions) is found to differ between roughly 10 and 26 kJ/mol for the different RNA atoms. Differences in stability between helical and complex folded structures and of the influence of metal ions on either are discussed.

Insight into the coordination and the binding sites of Cu(2+) by the histidyl-6-tag using experimental and computational tools.

PubMed

Watly, Joanna; Simonovsky, Eyal; Wieczorek, Robert; Barbosa, Nuno; Miller, Yifat; Kozlowski, Henryk

2014-07-07

His-tags are specific sequences containing six to nine subsequent histydyl residues, and they are used for purification of recombinant proteins by use of IMAC chromatography. Such polyhistydyl tags, often used in molecular biology, can be also found in nature. Proteins containing histidine-rich domains play a critical role in many life functions in both prokaryote and eukaryote organisms. Binding mode and the thermodynamic properties of the system depend on the specific metal ion and the histidine sequence. Despite the wide application of the His-tag for purification of proteins, little is known about the properties of metal-binding to such tag domains. This inspired us to undertake detailed studies on the coordination of Cu(2+) ion to hexa-His-tag. Experiments were performed using the potentiometric, UV-visible, CD, and EPR techniques. In addition, molecular dynamics (MD) simulations and density functional theory (DFT) calculations were applied. The experimental studies have shown that the Cu(2+) ion binds most likely to two imidazoles and one, two, or three amide nitrogens, depending on the pH. The structures and stabilities of the complexes for the Cu(2+)-Ac-(His)6-NH2 system using experimental and computational tools were established. Polymorphic binding states are suggested, with a possibility of the formation of α-helix structure induced by metal ion coordination. Metal ion is bound to various pairs of imidazole moieties derived from the tag with different efficiencies. The coordination sphere around the metal ion is completed by molecules of water. Finally, the Cu(2+) binding by Ac-(His)6-NH2 is much more efficient compared to other multihistidine protein domains.

Effects of the central potassium ions on the G-quadruplex and stabilizer binding.

PubMed

Wang, Zhiguo; Liu, Jun-Ping

2017-03-01

Human telomeres undertake the structure of intra-molecular parallel G-quadruplex in the presence of K + in eukaryotic cell. Stabilization of the telomere G-quadruplex represents a potential strategy to prevent telomere lengthening by telomerase in cancer therapy. Current work demonstrates that the binding of central K + with the parallel G-quadruplex is a coordinated water directed step-wise process. The K + above the top G-tetrad is prone to leak into environment and the 5'-adenine quickly flips over the top G-tetrad, leading to the bottom gate of G-tetrads as the only viable pathway of K + binding. Present molecular dynamics studies on the two most potent stabilizers RHPS4 and BRACO-19 reveal that the central K + has little influence on the binding conformations of the bound stabilizers. But without the central K + , either RHPS4 or BRACO-19 cannot stabilize the structure of G-quadruplex. The binding strength of stabilizers evaluated by the MM-PBSA method follows the order of BRACO-19> RHPS4, which agrees with the experimental results. The difference in binding affinities between RHPS4 and BRACO-19 is probably related to the ability to form intramolecular hydrogen bonds and favorable van del Waals interactions with G-quadruplex. In the models that have one central K + located at the upper/lower binding site, the corresponding top/bottom stacked stabilizers show more favorable binding affinities, indicating the apparent promoting effect of central K + on the stabilizer binding. Our findings provide further insights into the regulatory effect of K + on the G-quadruplex targeted binding, which is meaningful to the development of G-quadruplex stabilizers. Copyright © 2017 Elsevier Inc. All rights reserved.

Unambiguous identification and discovery of bacterial siderophores by direct injection 21 Tesla Fourier transform ion cyclotron resonance mass spectrometry.

PubMed

Walker, Lawrence R; Tfaily, Malak M; Shaw, Jared B; Hess, Nancy J; Paša-Tolić, Ljiljana; Koppenaal, David W

2017-01-25

Under iron-limiting conditions, bacteria produce low molecular mass Fe(iii) binding molecules known as siderophores to sequester the Fe(iii), along with other elements, increasing their bioavailability. Siderophores are thought to influence iron cycling and biogeochemistry in both marine and terrestrial ecosystems and hence the need for rapid, confident characterization of these compounds has increased. In this study, the type of siderophores produced by two marine bacterial species, Synechococcus sp. PCC 7002 and Vibrio cyclitrophicus 1F53, were characterized by use of a newly developed 21 T Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (FTICR MS) with direct injection electrospray ionization. This technique allowed for the rapid detection of synechobactins from Synechococcus sp. PCC 7002 as well as amphibactins from Vibrio cyclitrophicus 1F53 based on high mass accuracy and resolution allowing for observation of specific Fe isotopes and isotopic fine structure enabling highly confident identification of these siderophores. When combined with molecular network analysis two new amphibactins were discovered and verified by tandem MS. These results show that high-field FTICR MS is a powerful technique that will greatly improve the ability to rapidly identify and discover metal binding species in the environment.

Unambiguous identification and discovery of bacterial siderophores by direct injection 21 Tesla Fourier transform ion cyclotron resonance mass spectrometry

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

Walker, Lawrence R.; Tfaily, Malak M.; Shaw, Jared B.

Under iron-limiting conditions, bacteria produce low molecular mass Fe(III) binding molecules known as siderophores to sequester the Fe(III), along with other elements, increasing their bioavailibility. Siderophores are thought to influence iron cycling and biogeochemistry in both marine and terrestrial ecosystems and hence the need for rapid, confident characterization of these compounds has increased. In this study, the type of siderophores produced by two marine bacterial species, Synechococcus sp. PCC 7002 and Vibrio cyclitrophicus 1F53, were characterized using a newly developed 21T Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (FTICR MS) with direct injection electrospray ionization. This technique allowed for themore » rapid detection of synechobactins from Synechococcus sp. PCC 7002 as well as amphibactins from Vibrio cyclitrophicus 1F53 based on high mass accuracy and resolution allowing for observation of specific Fe isotopic peaks and fine isotopic structure enables highly confident identification of these sideropohores. When combined with molecular network analysis two new amphibactins were discovered and verified by tandem MS. These results show that high-field FTICR MS is a powerful technique that will greatly improve the ability to rapidly identify and discover metal binding species in the environment.« less

Low Temperature Extends the Lifespan of Bursaphelenchus xylophilus through the cGMP Pathway

PubMed Central

Wang, Bowen; Ma, Ling; Wang, Feng; Wang, Buyong; Hao, Xin; Xu, Jiayao; Ma, Yan

2017-01-01

The causal agent of pine wilt disease, pine wood nematode (PWN) (Bursaphelenchus xylophilus), revealed extended lifespan at low temperature. To discover the molecular mechanism of this phenomenon, we attempted to study the molecular characterization, transcript abundance, and functions of three genes of the cyclic guanosine monophosphate (cGMP) pathway from B. xylophilus. Three cGMP pathway genes were identified from B. xylophilus. Bioinformatic software was utilized to analyze the characteristics of the three putative proteins. Function of the three genes in cold tolerance was studied with RNA interference (RNAi). The results showed that the deduced protein of Bx-DAF-11 has an adenylate and guanylate cyclase catalytic domain, indicating an ability to bind to extracellular ligands and synthesizing cGMP. Both Bx-TAX-2 and Bx-TAX-4 have cyclic nucleotide-binding domains and ion transport protein domains, illustrating that they are cGMP-gated ion channels. The transcript level of Bx-daf-11, Bx-tax-2, and Bx-tax-4 increased at low temperature. The survival rates of three gene silenced B. xylophilus revealed a significant decrease at low temperature. This study illustrated that the cGMP pathway plays a key role in low-temperature-induced lifespan extension in B. xylophilus. PMID:29099744

Mg2+ ions: do they bind to nucleobase nitrogens?

PubMed Central

Leonarski, Filip; D'Ascenzo, Luigi; Auffinger, Pascal

2017-01-01

Given the many roles proposed for Mg2+ in nucleic acids, it is essential to accurately determine their binding modes. Here, we surveyed the PDB to classify Mg2+ inner-sphere binding patterns to nucleobase imine N1/N3/N7 atoms. Among those, purine N7 atoms are considered to be the best nucleobase binding sites for divalent metals. Further, Mg2+ coordination to N7 has been implied in several ribozyme catalytic mechanisms. We report that Mg2+ assigned near imine nitrogens derive mostly from poor interpretations of electron density patterns and are most often misidentified Na+, K+, NH4+ ions, water molecules or spurious density peaks. Consequently, apart from few documented exceptions, Mg2+ ions do not bind to N7 atoms. Without much of a surprise, Mn2+, Zn2+ and Cd2+, which have a higher affinity for nitrogens, may contact N7 atoms when present in crystallization buffers. In this respect, we describe for the first time a potential Zn2+ ribosomal binding site involving two purine N7 atoms. Further, we provide a set of guidelines to help in the assignment of Mg2+ in crystallographic, cryo-EM, NMR and model building practices and discuss implications of our findings related to ion substitution experiments. PMID:27923930

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.

Cooperativity and complexity in the binding of anions and cations to a tetratopic ion-pair host.

PubMed

Howe, Ethan N W; Bhadbhade, Mohan; Thordarson, Pall

2014-05-21

Cooperative interactions play a very important role in both natural and synthetic supramolecular systems. We report here on the cooperative binding properties of a tetratopic ion-pair host 1. This host combines two isophthalamide anion recognition sites with two unusual "half-crown/two carbonyl" cation recognition sites as revealed by the combination of single-crystal X-ray analysis of the free host and the 1:2 host:calcium cation complex, together with two-dimensional NMR and computational studies. By systematically comparing all of the binding data to several possible binding models and focusing on four different variants of the 1:2 binding model, it was in most cases possible to quantify these complex cooperative interactions. The data showed strong negative cooperativity (α = 0.01-0.05) of 1 toward chloride and acetate anions, while for cations the results were more variable. Interestingly, in the competitive (CDCl3/CD3OD (9:1, v/v)) solvent, the addition of calcium cations to the tetratopic ion-pair host 1 allosterically switched "on" chloride binding that is otherwise not present in this solvent system. The insight into the complexity of cooperative interactions revealed in this study of the tetratopic ion-pair host 1 can be used to design better cooperative supramolecular systems for information transfer and catalysis.

Engineering of a calcium-ion binding site into the RC-LH1-PufX complex of Rhodobacter sphaeroides to enable ion-dependent spectral red-shifting.

PubMed

Swainsbury, David J K; Martin, Elizabeth C; Vasilev, Cvetelin; Parkes-Loach, Pamela S; Loach, Paul A; Neil Hunter, C

2017-11-01

The reaction centre-light harvesting 1 (RC-LH1) complex of Thermochromatium (Tch.) tepidum has a unique calcium-ion binding site that enhances thermal stability and red-shifts the absorption of LH1 from 880nm to 915nm in the presence of calcium-ions. The LH1 antenna of mesophilic species of phototrophic bacteria such as Rhodobacter (Rba.) sphaeroides does not possess such properties. We have engineered calcium-ion binding into the LH1 antenna of Rba. sphaeroides by progressively modifying the native LH1 polypeptides with sequences from Tch. tepidum. We show that acquisition of the C-terminal domains from LH1 α and β of Tch. tepidum is sufficient to activate calcium-ion binding and the extent of red-shifting increases with the proportion of Tch. tepidum sequence incorporated. However, full exchange of the LH1 polypeptides with those of Tch. tepidum results in misassembled core complexes. Isolated α and β polypeptides from our most successful mutant were reconstituted in vitro with BChl a to form an LH1-type complex, which was stabilised 3-fold by calcium-ions. Additionally, carotenoid specificity was changed from spheroidene found in Rba. sphaeroides to spirilloxanthin found in Tch. tepidum, with the latter enhancing in vitro formation of LH1. These data show that the C-terminal LH1 α/β domains of Tch. tepidum behave autonomously, and are able to transmit calcium-ion induced conformational changes to BChls bound to the rest of a foreign antenna complex. Thus, elements of foreign antenna complexes, such as calcium-ion binding and blue/red switching of absorption, can be ported into Rhodobacter sphaeroides using careful design processes. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

Microstructured Optical Fiber-based Biosensors: Reversible and Nanoliter-Scale Measurement of Zinc Ions.

PubMed

Heng, Sabrina; McDevitt, Christopher A; Kostecki, Roman; Morey, Jacqueline R; Eijkelkamp, Bart A; Ebendorff-Heidepriem, Heike; Monro, Tanya M; Abell, Andrew D

2016-05-25

Sensing platforms that allow rapid and efficient detection of metal ions would have applications in disease diagnosis and study, as well as environmental sensing. Here, we report the first microstructured optical fiber-based biosensor for the reversible and nanoliter-scale measurement of metal ions. Specifically, a photoswitchable spiropyran Zn(2+) sensor is incorporated within the microenvironment of a liposome attached to microstructured optical fibers (exposed-core and suspended-core microstructured optical fibers). Both fiber-based platforms retains high selectivity of ion binding associated with a small molecule sensor, while also allowing nanoliter volume sampling and on/off switching. We have demonstrated that multiple measurements can be made on a single sample without the need to change the sensor. The ability of the new sensing platform to sense Zn(2+) in pleural lavage and nasopharynx of mice was compared to that of established ion sensing methodologies such as inductively coupled plasma mass spectrometry (ICP-MS) and a commercially available fluorophore (Fluozin-3), where the optical-fiber-based sensor provides a significant advantage in that it allows the use of nanoliter (nL) sampling when compared to ICP-MS (mL) and FluoZin-3 (μL). This work paves the way to a generic approach for developing surface-based ion sensors using a range of sensor molecules, which can be attached to a surface without the need for its chemical modification and presents an opportunity for the development of new and highly specific ion sensors for real time sensing applications.

Serum albumin forms a lactoferrin-like soluble iron-binding complex in presence of hydrogen carbonate ions.

PubMed

Ueno, Hiroshi M; Urazono, Hiroshi; Kobayashi, Toshiya

2014-02-15

The iron-lactoferrin complex is a common food ingredient because of its iron-solubilizing capability in the presence of hydrogen carbonate ions. However, it is unclear whether the formation of a stable iron-binding complex is limited to lactoferrin. In this study, we investigated the effects of bovine serum albumin (BSA) on iron solubility and iron-catalyzed lipid oxidation in the presence of hydrogen carbonate ions. BSA could solubilize >100-fold molar equivalents of iron at neutral pH, exceeding the specific metal-binding property of BSA. This iron-solubilizing capability of BSA was impaired by thermally denaturing BSA at ≥ 70 °C for 10 min at pH 8.5. The resulting iron-BSA complex inhibited iron-catalyzed oxidation of soybean oil in a water-in-oil emulsion measured using the Rancimat test. Our study is the first to show that BSA, like lactoferrin, forms a soluble iron-binding complex in the presence of hydrogen carbonate ions. Copyright © 2013 Elsevier Ltd. All rights reserved.

Measurement of Conformational Changes Accompanying Desensitization in an Ionotropic Glutamate Receptor

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

Armstrong,N.; Jasti, J.; Beich-Frandsen, M.

2006-01-01

The canonical conformational states occupied by most ligand-gated ion channels, and many cell-surface receptors, are the resting, activated, and desensitized states. While the resting and activated states of multiple receptors are well characterized, elaboration of the structural properties of the desensitized state, a state that is by definition inactive, has proven difficult. Here we use electrical, chemical, and crystallographic experiments on the AMPA-sensitive GluR2 receptor, defining the conformational rearrangements of the agonist binding cores that occur upon desensitization of this ligand-gated ion channel. These studies demonstrate that desensitization involves the rupture of an extensive interface between domain 1 of 2-foldmore » related glutamate-binding core subunits, compensating for the ca. 21{sup o} of domain closure induced by glutamate binding. The rupture of the domain 1 interface allows the ion channel to close and thereby provides a simple explanation to the long-standing question of how agonist binding is decoupled from ion channel gating upon receptor desensitization.« less

Structural effects of Cu(II)-coordination in the octapeptide region of the human prion protein.

PubMed

Riihimäki, Eva-Stina; Martínez, José Manuel; Kloo, Lars

2008-05-14

The copper-binding ability of the prion protein is thought to be central to its function. The structural effects of copper coordination in the octapeptide region of the human prion protein have been investigated by molecular dynamics simulations. Simulations were performed with the apo state, in order to investigate the behavior of the region without copper ions, as well as with the octapeptide region in the presence of copper ions. While the structure of the apo state is greatly influenced by the interaction between the rings in the histidine, tryptophan and proline residues, the region shows evidence of highly ordered coordination sites in the presence of copper ions. The position of the tryptophan indole ring is stabilized by cation-pi interactions. Two stable orientations of the indole ring with respect to the equatorial coordination plane of copper were observed, which showed that the indole ring can reside on both sides of the coordination plane. The interaction with the indole ring was found to occur without a mediating axial water molecule.

Metal cofactor modulated folding and target recognition of HIV-1 NCp7.

PubMed

Ren, Weitong; Ji, Dongqing; Xu, Xiulian

2018-01-01

The HIV-1 nucleocapsid 7 (NCp7) plays crucial roles in multiple stages of HIV-1 life cycle, and its biological functions rely on the binding of zinc ions. Understanding the molecular mechanism of how the zinc ions modulate the conformational dynamics and functions of the NCp7 is essential for the drug development and HIV-1 treatment. In this work, using a structure-based coarse-grained model, we studied the effects of zinc cofactors on the folding and target RNA(SL3) recognition of the NCp7 by molecular dynamics simulations. After reproducing some key properties of the zinc binding and folding of the NCp7 observed in previous experiments, our simulations revealed several interesting features in the metal ion modulated folding and target recognition. Firstly, we showed that the zinc binding makes the folding transition states of the two zinc fingers less structured, which is in line with the Hammond effect observed typically in mutation, temperature or denaturant induced perturbations to protein structure and stability. Secondly, We showed that there exists mutual interplay between the zinc ion binding and NCp7-target recognition. Binding of zinc ions enhances the affinity between the NCp7 and the target RNA, whereas the formation of the NCp7-RNA complex reshapes the intrinsic energy landscape of the NCp7 and increases the stability and zinc affinity of the two zinc fingers. Thirdly, by characterizing the effects of salt concentrations on the target RNA recognition, we showed that the NCp7 achieves optimal balance between the affinity and binding kinetics near the physiologically relevant salt concentrations. In addition, the effects of zinc binding on the inter-domain conformational flexibility and folding cooperativity of the NCp7 were also discussed.

The nature of cation-pi binding: interactions between tetramethylammonium ion and benzene in aqueous solution.

PubMed Central

Gao, J; Chou, L W; Auerbach, A

1993-01-01

A combined quantum mechanical and molecular mechanical Monte Carlo simulation method was used to determine the free energy of binding between tetramethylammonium ion (TMA+) and benzene in water. The computed free energy as a function of distance (the potential of mean force) has two minima that represent contact and solvent-separated complexes. These species are separated by a broad barrier of about 3 kJ/mol. The results are in good accord with experimental data and suggest that TMA+ binds to benzene more favorably than to chloride ion, with an association constant of about 0.8 M-1. Images FIGURE 2 PMID:8369448

Importance of Diffuse Metal Ion Binding to RNA

PubMed Central

Tan, Zhi-Jie; Chen, Shi-Jie

2016-01-01

RNAs are highly charged polyanionic molecules. RNA structure and function are strongly correlated with the ionic condition of the solution. The primary focus of this article is on the role of diffusive ions in RNA folding. Due to the long-range nature of electrostatic interactions, the diffuse ions can contribute significantly to RNA structural stability and folding kinetics. We present an overview of the experimental findings as well as the theoretical developments on the diffuse ion effects in RNA folding. This review places heavy emphasis on the effect of magnesium ions. Magnesium ions play a highly efficient role in stabilizing RNA tertiary structures and promoting tertiary structural folding. The highly efficient role goes beyond the mean-field effect such as the ionic strength. In addition to the effects of specific ion binding and ion dehydration, ion-ion correlation for the diffuse ions can contribute to the efficient role of the multivalent ions such as the magnesium ions in RNA folding. PMID:22010269

Importance of diffuse metal ion binding to RNA.

PubMed

Tan, Zhi-Jie; Chen, Shi-Jie

2011-01-01

RNAs are highly charged polyanionic molecules. RNA structure and function are strongly correlated with the ionic condition of the solution. The primary focus of this article is on the role of diffusive ions in RNA folding. Due to the long-range nature of electrostatic interactions, the diffuse ions can contribute significantly to RNA structural stability and folding kinetics. We present an overview of the experimental findings as well as the theoretical developments on the diffuse ion effects in RNA folding. This review places heavy emphasis on the effect of magnesium ions. Magnesium ions play a highly efficient role in stabilizing RNA tertiary structures and promoting tertiary structural folding. The highly efficient role goes beyond the mean-field effect such as the ionic strength. In addition to the effects of specific ion binding and ion dehydration, ion-ion correlation for the diffuse ions can contribute to the efficient role of the multivalent ions such as the magnesium ions in RNA folding.

In vivo-folded metal-metallothionein 3 complexes reveal the Cu-thionein rather than Zn-thionein character of this brain-specific mammalian metallothionein.

PubMed

Artells, Ester; Palacios, Oscar; Capdevila, Mercè; Atrian, Sílvia

2014-03-01

Metallothionein-3 (MT3) is one of the four mammalian metallothioneins (MT), and is constitutively synthesized in the brain. MT3 acts both intracellularly and extracellularly in this organ, performing functions related to neuronal growth and physiological metal (Zn and Cu) handling. It appears to be involved in the prevention of neurodegenerative disorders caused by insoluble Cu-peptide aggregates, as it triggers a Zn-Cu swap that may counteract the deleterious presence of copper in neural tissues. The literature data on MT3 coordination come from studies either on apo-MT3 reconstitution or the reaction of Zn-MT3 with Cu(2+) , an ion that is hardly present inside cells. To ascertain the MT3 metal-binding features in a scenario closer to the reductive cell cytoplasm, a study of the recombinant Zn(2+) , Cd(2+) and Cu(+) complexes of MT3, βMT3, and αMT3, as well as the in vitro Zn(2+) -Cd(2+) and Zn(2+) -Cu(+) replacement processes, is presented here. We conclude that MT3 has a Cu-thionein character that is stronger than that of the MT1 and MT2 isoforms - also present in the mammalian brain - which is mainly contributed by its β domain. In contrast, the α domain retains a high capacity to bind Zn(2+) ions, and, consequently, the entire MT3 peptide shows a peculiar dual ability to handle both metal ions. The nature of the formed Cu(+) -MT3 complexes oscillates from heterometallic Cu6 Zn4 -MT3 to homometallic Cu10 -MT3 major species, in a narrow Cu concentration range. Therefore, the entire MT3 peptide shows a high capacity to bind Cu(+) , provided that this occurs in a nonoxidative milieux. This reflects a peculiar property of this MT isoform, which accurately senses different Cu contents in the environment in which it is synthesized. © 2014 FEBS.

Cross-modal working memory binding and word recognition skills: how specific is the link?

PubMed

Wang, Shinmin; Allen, Richard J

2018-04-01

Recent research has suggested that the creation of temporary bound representations of information from different sources within working memory uniquely relates to word recognition abilities in school-age children. However, it is unclear to what extent this link is attributable specifically to the binding ability for cross-modal information. This study examined the performance of Grade 3 (8-9 years old) children on binding tasks requiring either temporary association formation of two visual items (i.e., within-modal binding) or pairs of visually presented abstract shapes and auditorily presented nonwords (i.e., cross-modal binding). Children's word recognition skills were related to performance on the cross-modal binding task but not on the within-modal binding task. Further regression models showed that cross-modal binding memory was a significant predictor of word recognition when memory for its constituent elements, general abilities, and crucially, within-modal binding memory were taken into account. These findings may suggest a specific link between the ability to bind information across modalities within working memory and word recognition skills.

Binding of iron, zinc, and lead ions from aqueous solution by shea butter (Butyrospermun Parkii) seed husks

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

Eromosele, I.C.; Otitolaye, O.O.

1994-08-01

Several workers have reported on the potential use of agricultural products as substrates for the removal of metal ions from aqueous solutions. These studies demonstrated that considerable amounts of metal ions can be removed from aqueous solutions by cellulosic materials. The merit in the use of the latter is their relative abundance and cheapness compared to conventional materials for the removal of toxic metal ions from waste-waters. In some of the studies, chemical modification of cellulosic materials significantly enhanced their ion-binding properties, providing greater flexibility in their applications to a wide range of heavy metal ions. Shea butter plant (Butyrospermunmore » Parkii) normally grows in the wild within the guinea-savana zone of Nigeria. The seeds are a rich source of edible oils and the husks are usually discarded. The husk is thus available in abundance and, hence, there is reason to examine its ion-binding properties for its possible application in the removal of toxic metal ions from industrial waste-waters. This paper reports on preliminary studies of the sorption of iron, zinc and lead ions from aqueous solution by modified and unmodified shea butter seed husks. 8 refs., 5 figs., 1 tab.« less

Calcium ion binding properties and the effect of phosphorylation on the intrinsically disordered Starmaker protein.

PubMed

Wojtas, Magdalena; Hołubowicz, Rafał; Poznar, Monika; Maciejewska, Marta; Ożyhar, Andrzej; Dobryszycki, Piotr

2015-10-27

Starmaker (Stm) is an intrinsically disordered protein (IDP) involved in otolith biomineralization in Danio rerio. Stm controls calcium carbonate crystal formation in vivo and in vitro. Phosphorylation of Stm affects its biomineralization properties. This study examined the effects of calcium ions and phosphorylation on the structure of Stm. We have shown that CK2 kinase phosphorylates 25 or 26 residues in Stm. Furthermore, we have demonstrated that Stm's affinity for calcium binding is dependent on its phosphorylation state. Phosphorylated Stm (StmP) has an estimated 30 ± 1 calcium binding sites per protein molecule with a dissociation constant (KD) of 61 ± 4 μM, while the unphosphorylated protein has 28 ± 3 sites and a KD of 210 ± 22 μM. Calcium ion binding induces a compaction of the Stm molecule, causing a significant decrease in its hydrodynamic radius and the formation of a secondary structure. The screening effect of Na(+) ions on calcium binding was also observed. Analysis of the hydrodynamic properties of Stm and StmP showed that Stm and StmP molecules adopt the structure of native coil-like proteins.

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.

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.

Energetics and kinetics of cooperative cofilin-actin filament interactions.

PubMed

Cao, Wenxiang; Goodarzi, Jim P; De La Cruz, Enrique M

2006-08-11

We have evaluated the thermodynamic parameters associated with cooperative cofilin binding to actin filaments, accounting for contributions of ion-linked equilibria, and determined the kinetic basis of cooperative cofilin binding. Ions weaken non-contiguous (isolated, non-cooperative) cofilin binding to an actin filament without affecting cooperative filament interactions. Non-contiguous cofilin binding is coupled to the dissociation of approximately 1.7 thermodynamically bound counterions. Counterion dissociation contributes approximately 40% of the total cofilin binding free energy (in the presence of 50 mM KCl). The non-contiguous and cooperative binding free energies are driven entirely by large, positive entropy changes, consistent with a cofilin-mediated increase in actin filament structural dynamics. The rate constant for cofilin binding to an isolated site on an actin filament is slow and likely to be limited by filament breathing. Cooperative cofilin binding arises from an approximately tenfold more rapid association rate constant and an approximately twofold slower dissociation rate constant. The more rapid association rate constant is presumably a consequence of cofilin-dependent changes in the average orientation of subdomain 2, subunit angular disorder and filament twist, which increase the accessibility of a neighboring cofilin-binding site on an actin filament. Cooperative association is more rapid than binding to an isolated site, but still slow for a second-order reaction, suggesting that cooperative binding is limited also by binding site accessibility. We suggest that the dissociation of actin-associated ions weakens intersubunit interactions in the actin filament lattice that enhance cofilin-binding site accessibility, favor cooperative binding and promote filament severing.

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).

Comparison of structural, thermodynamic, kinetic and mass transport properties of Mg(2+) ion models commonly used in biomolecular simulations.

PubMed

Panteva, Maria T; Giambaşu, George M; York, Darrin M

2015-05-15

The prevalence of Mg(2+) ions in biology and their essential role in nucleic acid structure and function has motivated the development of various Mg(2+) ion models for use in molecular simulations. Currently, the most widely used models in biomolecular simulations represent a nonbonded metal ion as an ion-centered point charge surrounded by a nonelectrostatic pairwise potential that takes into account dispersion interactions and exchange effects that give rise to the ion's excluded volume. One strategy toward developing improved models for biomolecular simulations is to first identify a Mg(2+) model that is consistent with the simulation force fields that closely reproduces a range of properties in aqueous solution, and then, in a second step, balance the ion-water and ion-solute interactions by tuning parameters in a pairwise fashion where necessary. The present work addresses the first step in which we compare 17 different nonbonded single-site Mg(2+) ion models with respect to their ability to simultaneously reproduce structural, thermodynamic, kinetic and mass transport properties in aqueous solution. None of the models based on a 12-6 nonelectrostatic nonbonded potential was able to reproduce the experimental radial distribution function, solvation free energy, exchange barrier and diffusion constant. The models based on a 12-6-4 potential offered improvement, and one model in particular, in conjunction with the SPC/E water model, performed exceptionally well for all properties. The results reported here establish useful benchmark calculations for Mg(2+) ion models that provide insight into the origin of the behavior in aqueous solution, and may aid in the development of next-generation models that target specific binding sites in biomolecules. © 2015 Wiley Periodicals, Inc.

Antioxidant mechanism of milk mineral-high-affinity iron binding.

PubMed

Allen, K; Cornforth, D

2007-01-01

Milk mineral (MM), a by-product of whey processing, is an effective antioxidant in meat systems, but the antioxidant mechanism has not been established. MM has been postulated to chelate iron and prevent iron-catalysis of lipid oxidation. The objective of this research was to examine this putative mechanism. MM was compared to sodium tripolyphosphate (STPP), calcium phosphate monobasic (CPM), and calcium pyrophosphate (CPP) to determine iron-binding capacity, sample solubility, and eluate soluble phosphorus after treating samples with a ferrous chloride standard. Scanning electron microscopy with energy-dispersive X-ray analysis was used to localize minerals on iron-treated MM particle surfaces. Histochemical staining for calcium was performed on raw and cooked ground beef samples with added MM. MM bound more iron per gram (P < 0.05) than the other compounds, and was much less soluble (P < 0.05) than either STPP or CPM. Mineral localization showed an even distribution of calcium, phosphorus, oxygen, and iron across the MM particle surface, directly demonstrating iron binding to MM particles. Unlike other common chelating agents, such as STPP and citrate, histochemical staining demonstrated that MM remained insoluble in ground beef, even after cooking. The ability of MM to bind iron and remain insoluble may enhance its antioxidant effect by removing iron ions from solution. However, MM particles must be small and well distributed in order to adequately bind iron throughout the food system.

Structure of the large FK506-binding protein FKBP51, an Hsp90-binding protein and a component of steroid receptor complexes

PubMed Central

Sinars, Cindy R.; Cheung-Flynn, Joyce; Rimerman, Ronald A.; Scammell, Jonathan G.; Smith, David F.; Clardy, Jon

2003-01-01

The ability to bind immunosuppressive drugs such as cyclosporin and FK506 defines the immunophilin family of proteins, and the FK506-binding proteins form the FKBP subfamily of immunophilins. Some FKBPs, notably FKBP12 (the 12-kDa FK506-binding protein), have defined roles in regulating ion channels or cell signaling, and well established structures. Other FKBPs, especially the larger ones, participate in important biological processes, but their exact roles and the structural bases for these roles are poorly defined. FKBP51 (the 51-kDa FKBP) associates with heat shock protein 90 (Hsp90) and appears in functionally mature steroid receptor complexes. In New World monkeys, FKBP51 has been implicated in cortisol resistance. We report here the x-ray structures of human FKBP51, to 2.7 Å, and squirrel monkey FKBP51, to 2.8 Å, by using multiwavelength anomalous dispersion phasing. FKBP51 is composed of three domains: two consecutive FKBP domains and a three-unit repeat of the TPR (tetratricopeptide repeat) domain. This structure of a multi-FKBP domain protein clarifies the arrangement of these domains and their possible interactions with other proteins. The two FKBP domains differ by an insertion in the second that affects the formation of the progesterone receptor complex. PMID:12538866

Simultaneous addition of two ligands: a potential strategy for estimating divalent ion affinities in EF-hand proteins by isothermal titration calorimetry.

PubMed

Henzl, Michael T; Markus, Lindsey A; Davis, Meredith E; McMillan, Andrew T

2013-03-01

Capable of providing a detailed thermodynamic picture of noncovalent association reactions, isothermal titration calorimetry (ITC) has become a popular method for studying protein-ligand interactions. We routinely employ the technique to study divalent ion-binding by two-site EF-hand proteins from the parvalbumin- and polcalcin lineages. The combination of high Ca(2+) affinity and relatively low Mg(2+) affinity, and the attendant complication of parameter correlation, conspire to make the simultaneous extraction of binding constants and -enthalpies for both ions challenging. Although global analysis of multiple ITC experiments can overcome these hurdles, our current experimental protocol includes upwards of 10 titrations - requiring a substantial investment in labor, machine time, and material. This paper explores the potential for using a smaller suite of experiments that includes simultaneous titrations with Ca(2+) and Mg(2+) at different ratios of the two ions. The results obtained for four proteins, differing substantially in their divalent ion-binding properties, suggest that the approach has merit. The Ca(2+)- and Mg(2+)-binding constants afforded by the streamlined analysis are in reasonable agreement with those obtained from the standard analysis protocol. Likewise, the abbreviated analysis provides comparable values for the Ca(2+)-binding enthalpies. However, the streamlined analysis can yield divergent values for the Mg(2+)-binding enthalpies - particularly those for lower affinity sites. This shortcoming can be remedied, in large measure, by including data from a direct Ca(2+) titration in the presence of a high, fixed Mg(2+) concentration. Copyright © 2013. Published by Elsevier Inc.

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.

Binding of Dissolved Strontium by Micrococcus luteus

PubMed Central

Faison, Brendlyn D.; Cancel, Carmen A.; Lewis, Susan N.; Adler, Howard I.

1990-01-01

Resting cells of Micrococcus luteus have been shown to remove strontium (Sr) from dilute aqueous solutions of SrCl2 at pH 7. Loadings of 25 mg of Sr per g of cell dry weight were achieved by cells exposed to a solution containing 50 ppm (mg/liter) of Sr. Sr binding occurred in the absence of nutrients and did not require metabolic activity. Initial binding was quite rapid (<0.5 h), although a slow, spontaneous release of Sr was observed over time. Sr binding was inhibited in the presence of polyvalent cations but not monovalent cations. Ca and Sr were bound preferentially over all other cations tested. Sr-binding activity was localized on the cell envelope and was sensitive to various chemical and physical pretreatments. Bound Sr was displaced by divalent ions or by H+. Other monovalent ions were less effective. Bound Sr was also removed by various chelating agents. It was concluded that Sr binding by M. luteus is a reversible equilibrium process. Both ion exchange mediated by acidic cell surface components and intracellular uptake may be involved in this activity. PMID:16348370

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

Copper nanocluster coupling europium as an off-to-on fluorescence probe for the determination of phosphate ion in water samples.

PubMed

Cao, Haiyan; Chen, Zhaohui; Huang, Yuming

2015-10-01

This paper reports an "off-to-on" fluorescence (FL) probe for sensitively and selectively detecting phosphate ions (Pi's). Fabrication of the probe was based on the competition between Pi's and tannic acid-stabilized copper nanoclusters (TA-Cu NCs) for Eu(3+) binding. The addition of Eu(3+) ions to TA-Cu NCs triggered the aggregation of TA-Cu NCs, which quenched the FL of TA-Cu NCs. After Pi addition, the aggregated TA-Cu NCs solubilized into the aqueous solution to facilitate the Pi-triggered dispersion of TA-Cu NCs. This phenomenon was due to the stronger binding ability between Pi's and Eu(3+) than that between TA and Eu(3+), leading to FL recovery of Cu NCs. The degree of redispersion of TA-Cu NCs was directly related to Pi concentration. Thus, Pi concentration can be quantitatively determined by the change in FL of the TA-Cu NCs dispersion. Under the optimized conditions, the change in FL presented a linear relationship with Pi concentration from 0.07 μmol L(-1) to 80 μmol L(-1). The limit of detection for Pi was 9.6×10(-3) μmol L(-1) at a signal-to-noise ratio of 3. For Pi determination in real samples, only 1 mL water sample was needed. The proposed probe was highly sensitive, free from the interference of other common species in aqueous media, and particularly useful for the fast and simple diagnosis of water-eutrophication extent. Copyright © 2015 Elsevier B.V. All rights reserved.

Effects of rare earth doping on multi-core iron oxide nanoparticles properties

NASA Astrophysics Data System (ADS)

Petran, Anca; Radu, Teodora; Borodi, Gheorghe; Nan, Alexandrina; Suciu, Maria; Turcu, Rodica

2018-01-01

New multi-core iron oxide magnetic nanoparticles doped with rare earth metals (Gd, Eu) were obtained by a one step synthesis procedure using a solvothermal method for potential biomedical applications. The obtained clusters were characterized by X-ray diffraction (XRD), transmission electron microscopy (TEM), energy-dispersive X-ray microanalysis (EDX), X-ray photoelectron spectroscopy (XPS) and magnetization measurements. They possess high colloidal stability, a saturation magnetization of up to 52 emu/g, and nearly spherical shape. The presence of rare earth ions in the obtained samples was confirmed by EDX and XPS. XRD analysis proved the homogeneous distribution of the trivalent rare earth ions in the inverse-spinel structure of magnetite and the increase of crystal strain upon doping the samples. XPS study reveals the valence state and the cation distribution on the octahedral and tetrahedral sites of the analysed samples. The observed shift of the XPS valence band spectra maximum in the direction of higher binding energies after rare earth doping, as well as theoretical valence band calculations prove the presence of Gd and Eu ions in octahedral sites. The blood protein adsorption ability of the obtained samples surface, the most important factor of the interaction between biomaterials and body fluids, was assessed by interaction with bovine serum albumin (BSA). The rare earth doped clusters surface show higher afinity for binding BSA. In vitro cytotoxicity test results for the studied samples showed no cytotoxicity in low and medium doses, establishing a potential perspective for rare earth doped MNC to facilitate multiple therapies in a single formulation for cancer theranostics.

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

Stochastic steps in secondary active sugar transport

PubMed Central

Adelman, Joshua L.; Ghezzi, Chiara; Bisignano, Paola; Loo, Donald D. F.; Choe, Seungho; Abramson, Jeff; Rosenberg, John M.; Wright, Ernest M.; Grabe, Michael

2016-01-01

Secondary active transporters, such as those that adopt the leucine-transporter fold, are found in all domains of life, and they have the unique capability of harnessing the energy stored in ion gradients to accumulate small molecules essential for life as well as expel toxic and harmful compounds. How these proteins couple ion binding and transport to the concomitant flow of substrates is a fundamental structural and biophysical question that is beginning to be answered at the atomistic level with the advent of high-resolution structures of transporters in different structural states. Nonetheless, the dynamic character of the transporters, such as ion/substrate binding order and how binding triggers conformational change, is not revealed from static structures, yet it is critical to understanding their function. Here, we report a series of molecular simulations carried out on the sugar transporter vSGLT that lend insight into how substrate and ions are released from the inward-facing state of the transporter. Our simulations reveal that the order of release is stochastic. Functional experiments were designed to test this prediction on the human homolog, hSGLT1, and we also found that cytoplasmic release is not ordered, but we confirmed that substrate and ion binding from the extracellular space is ordered. Our findings unify conflicting published results concerning cytoplasmic release of ions and substrate and hint at the possibility that other transporters in the superfamily may lack coordination between ions and substrate in the inward-facing state. PMID:27325773

Stochastic steps in secondary active sugar transport.

PubMed

Adelman, Joshua L; Ghezzi, Chiara; Bisignano, Paola; Loo, Donald D F; Choe, Seungho; Abramson, Jeff; Rosenberg, John M; Wright, Ernest M; Grabe, Michael

2016-07-05

Secondary active transporters, such as those that adopt the leucine-transporter fold, are found in all domains of life, and they have the unique capability of harnessing the energy stored in ion gradients to accumulate small molecules essential for life as well as expel toxic and harmful compounds. How these proteins couple ion binding and transport to the concomitant flow of substrates is a fundamental structural and biophysical question that is beginning to be answered at the atomistic level with the advent of high-resolution structures of transporters in different structural states. Nonetheless, the dynamic character of the transporters, such as ion/substrate binding order and how binding triggers conformational change, is not revealed from static structures, yet it is critical to understanding their function. Here, we report a series of molecular simulations carried out on the sugar transporter vSGLT that lend insight into how substrate and ions are released from the inward-facing state of the transporter. Our simulations reveal that the order of release is stochastic. Functional experiments were designed to test this prediction on the human homolog, hSGLT1, and we also found that cytoplasmic release is not ordered, but we confirmed that substrate and ion binding from the extracellular space is ordered. Our findings unify conflicting published results concerning cytoplasmic release of ions and substrate and hint at the possibility that other transporters in the superfamily may lack coordination between ions and substrate in the inward-facing state.

Radioactive ion detector

DOEpatents

Bower, Kenneth E.; Weeks, Donald R.

1997-01-01

Apparatus for detecting the presence, in aqueous media, of substances which emit alpha and/or beta radiation and determining the oxidation state of these radioactive substances, that is, whether they are in cationic or anionic form. In one embodiment, a sensor assembly has two elements, one comprised of an ion-exchange material which binds cations and the other comprised of an ion-exchange material which binds anions. Each ion-exchange element is further comprised of a scintillation plastic and a photocurrent generator. When a radioactive substance to which the sensor is exposed binds to either element and emits alpha or beta particles, photons produced in the scintillation plastic illuminate the photocurrent generator of that element. Sensing apparatus senses generator output and thereby indicates whether cationic species or anionic species or both are present and also provides an indication of species quantity.

Radioactive ion detector

DOEpatents

Bower, K.E.; Weeks, D.R.

1997-08-12

Apparatus for detecting the presence, in aqueous media, of substances which emit alpha and/or beta radiation and determining the oxidation state of these radioactive substances, that is, whether they are in cationic or anionic form. In one embodiment, a sensor assembly has two elements, one comprised of an ion-exchange material which binds cations and the other comprised of an ion-exchange material which binds anions. Each ion-exchange element is further comprised of a scintillation plastic and a photocurrent generator. When a radioactive substance to which the sensor is exposed binds to either element and emits alpha or beta particles, photons produced in the scintillation plastic illuminate the photocurrent generator of that element. Sensing apparatus senses generator output and thereby indicates whether cationic species or anionic species or both are present and also provides an indication of species quantity. 2 figs.

Dynamic ion-ion and water-ion interactions in ion channels.

PubMed Central

Wu, J V

1992-01-01

The dynamic interactions among ions and water molecules in ion channels are treated based on an assumption that ions at binding sites can be knocked off by both transient entering ions and local water molecules. The theory, when applied to a single-site model K+ channel, provides solutions for super- and subsaturations, flux-ratio exponent (n') greater than 1, osmotic streaming current, activity-dependent reversal potentials, and anomalous mole-fraction behavior. The analysis predicts that: (a) the saturation may but, in general, does not follow the Michaelis-Menten relation; (b) streaming current results from imbalanced water-ion knock-off interactions; (c) n' greater than 1 even for single-site channels, but it is unlikely to exceed 1.4 unless the pore is occupied by one or more ion(s); (d) in the calculation involving two permeant ion species with similar radii, the heavier ions show higher affinity; the ion-ion knock-off dissociation from the site is more effective when two interacting ions are identical. Therefore, the "multi-ion behaviors" found in most ion channels are the consequences of dynamic ion-ion and water-ion interactions. The presence of these interactions does not require two or more binding sites in channels. PMID:1376158

Use of Methacrylic Acid-Containing Hydrogels to Increase Protein Transport Across the Intestinal Epithelium

NASA Astrophysics Data System (ADS)

Blanchette, James; Lopez, Jennifer; Park, Kinam; Peppas, Nicholas

2002-03-01

Oral protein delivery requires protection from the harsh environment of the stomach, release in the small intestine and passage from the intestinal lumen into the circulation. Hydrogels that swell in response to the pH change when passing from the stomach to the small intestine can accomplish the first two points. The ability to enhance the permeability of intestinal epithelial cells is currently under investigation. Methacrylic acid-containing hydrogels have shown the ability to bind calcium ions that decreases the concentration of free extracellular calcium for these epithelial cells. This change triggers a number of intracellular events including rearrangement of the cytoskeleton leading to increased permeability. Studies done on Caco-2 cells (human colon adenocarcinoma) measuring changes in transepithelial resistance are used to assess the effect of the polymer-cell interactions on the integrity of intestinal epithelial cell monolayers.

Application of differential scanning calorimetry to measure the differential binding of ions, water and protons in the unfolding of DNA molecules.

PubMed

Olsen, Chris M; Shikiya, Ronald; Ganugula, Rajkumar; Reiling-Steffensmeier, Calliste; Khutsishvili, Irine; Johnson, Sarah E; Marky, Luis A

2016-05-01

The overall stability of DNA molecules globally depends on base-pair stacking, base-pairing, polyelectrolyte effect and hydration contributions. In order to understand how they carry out their biological roles, it is essential to have a complete physical description of how the folding of nucleic acids takes place, including their ion and water binding. To investigate the role of ions, water and protons in the stability and melting behavior of DNA structures, we report here an experimental approach i.e., mainly differential scanning calorimetry (DSC), to determine linking numbers: the differential binding of ions (Δnion), water (ΔnW) and protons (ΔnH(+)) in the helix-coil transition of DNA molecules. We use DSC and temperature-dependent UV spectroscopic techniques to measure the differential binding of ions, water, and protons for the unfolding of a variety of DNA molecules: salmon testes DNA (ST-DNA), one dodecamer, one undecamer and one decamer duplexes, nine hairpin loops, and two triplexes. These methods can be applied to any conformational transition of a biomolecule. We determined complete thermodynamic profiles, including all three linking numbers, for the unfolding of each molecule. The favorable folding of a DNA helix results from a favorable enthalpy-unfavorable entropy compensation. DSC thermograms and UV melts as a function of salt, osmolyte and proton concentrations yielded releases of ions and water. Therefore, the favorable folding of each DNA molecule results from the formation of base-pair stacks and uptake of both counterions and water molecules. In addition, the triplex with C(+)GC base triplets yielded an uptake of protons. Furthermore, the folding of a DNA duplex is accompanied by a lower uptake of ions and a similar uptake of four water molecules as the DNA helix gets shorter. In addition, the oligomer duplexes and hairpin thermodynamic data suggest ion and water binding depends on the DNA sequence rather than DNA composition. Copyright © 2015. Published by Elsevier B.V.

Elucidation of the Molecular Basis for Arabinoxylan-Debranching Activity of a Thermostable Family GH62 α-l-Arabinofuranosidase from Streptomyces thermoviolaceus

PubMed Central

Wang, Weijun; Mai-Gisondi, Galina; Stogios, Peter J.; Kaur, Amrit; Xu, Xiaohui; Cui, Hong; Turunen, Ossi; Savchenko, Alexei

2014-01-01

Xylan-debranching enzymes facilitate the complete hydrolysis of xylan and can be used to alter xylan chemistry. Here, the family GH62 α-l-arabinofuranosidase from Streptomyces thermoviolaceus (SthAbf62A) was shown to have a half-life of 60 min at 60°C and the ability to cleave α-1,3 l-arabinofuranose (l-Araf) from singly substituted xylopyranosyl (Xylp) backbone residues in wheat arabinoxylan; low levels of activity on arabinan as well as 4-nitrophenyl α-l-arabinofuranoside were also detected. After selective removal of α-1,3 l-Araf substituents from disubstituted Xylp residues present in wheat arabinoxylan, SthAbf62A could also cleave the remaining α-1,2 l-Araf substituents, confirming the ability of SthAbf62A to remove α-l-Araf residues that are (1→2) and (1→3) linked to monosubstituted β-d-Xylp sugars. Three-dimensional structures of SthAbf62A and its complex with xylotetraose and l-arabinose confirmed a five-bladed β-propeller fold and revealed a molecular Velcro in blade V between the β1 and β21 strands, a disulfide bond between Cys27 and Cys297, and a calcium ion coordinated in the central channel of the fold. The enzyme-arabinose complex structure further revealed a narrow and seemingly rigid l-arabinose binding pocket situated at the center of one side of the β propeller, which stabilized the arabinofuranosyl substituent through several hydrogen-bonding and hydrophobic interactions. The predicted catalytic amino acids were oriented toward this binding pocket, and the catalytic essentiality of Asp53 and Glu213 was confirmed by site-specific mutagenesis. Complex structures with xylotetraose revealed a shallow cleft for xylan backbone binding that is open at both ends and comprises multiple binding subsites above and flanking the l-arabinose binding pocket. PMID:24951792

A Hexahomotrioxacalix[3]arene-Based Ditopic Receptor for Alkylammonium Ions Controlled by Ag⁺ Ions.

PubMed

Jiang, Xue-Kai; Ikejiri, Yusuke; Wu, Chong; Rahman, Shofiur; Georghiou, Paris E; Zeng, Xi; Elsegood, Mark R J; Redshaw, Carl; Teat, Simon J; Yamato, Takehiko

2018-02-21

A receptor cone-1 based on a hexahomotrioxacalix[3]arene bearing three pyridyl groups was successfully synthesized, which has a C₃-symmetric conformation and is capable of binding alkylammonium and metal ions simultaneously in a cooperative fashion. It can bind alkylammonium ions through the -cavity formed by three aryl rings. This behaviour is consistent with the cone-in/cone-out conformational rearrangement needed to reorganize the cavity for endo-complexation. As a C₃-symmetrical pyridyl-substituted calixarene, receptor cone-1 can also bind an Ag⁺ ion, and the nitrogen atoms are turned towards the inside of the cavity and interact with Ag⁺. After complexation of tris(2-pyridylamide) derivative receptor cone-1 with Ag⁺, the original C₃-symmetry was retained and higher complexation selectivity for n-BuNH₃⁺ versus t-BuNH₃⁺ was observed. Thus, it is believed that this receptor will have a role to play in the sensing, detection, and recognition of Ag⁺ and n-BuNH₃ + ions.

Adsorption of surfactant ions and binding of their counterions at an air/water interface.

PubMed

Tagashira, Hiroaki; Takata, Youichi; Hyono, Atsushi; Ohshima, Hiroyuki

2009-01-01

An expression for the surface tension of an aqueous mixed solution of surfactants and electrolyte ions in the presence of the common ions was derived from the Helmholtz free energy of an air/water surface. By applying the equation to experimental data for the surface tension, the adsorption constant of surfactant ions onto the air/water interface, the binding constant of counterions on the surfactants, and the surface potential and surface charge density of the interface were estimated. The adsorption constant and binding constant were dependent on the species of surfactant ion and counterion, respectively. Taking account of the dependence of surface potential and surface charge density on the concentration of electrolyte, it was suggested that the addition of electrolyte to the aqueous surfactant solution brings about the decrease in the surface potential, the increase in the surface density of surfactant ions, and consequently, the decrease in the surface tension. Furthermore, it was found that the configurational entropy plays a predominant role for the surface tension, compared to the electrical work.

A Hexahomotrioxacalix[3]arene-Based Ditopic Receptor for Alkylammonium Ions Controlled by Ag + Ions

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

Jiang, Xue-Kai; Ikejiri, Yusuke; Wu, Chong

A receptor cone-1 based on a hexahomotrioxacalix[3]arene bearing three pyridyl groups was successfully synthesized, which has a C 3-symmetric conformation and is capable of binding alkylammonium and metal ions simultaneously in a cooperative fashion. It can bind alkylammonium ions through the π-cavity formed by three aryl rings. This behaviour is consistent with the cone-in/cone-out conformational rearrangement needed to reorganize the cavity for endo-complexation. As a C 3-symmetrical pyridyl-substituted calixarene, receptor cone-1 can also bind an Ag + ion, and the nitrogen atoms are turned towards the inside of the cavity and interact with Ag +. After complexation of tris(2-pyridylamide) derivativemore » receptor cone-1 with Ag +, the original C 3-symmetry was retained and higher complexation selectivity for n-BuNH 3 + versus t-BuNH 3 + was observed. Thus, it is believed that this receptor will have a role to play in the sensing, detection, and recognition of Ag + and n-BuNH 3 + ions.« less

A Hexahomotrioxacalix[3]arene-Based Ditopic Receptor for Alkylammonium Ions Controlled by Ag + Ions

DOE PAGES

Jiang, Xue-Kai; Ikejiri, Yusuke; Wu, Chong; ...

2018-02-21

A receptor cone-1 based on a hexahomotrioxacalix[3]arene bearing three pyridyl groups was successfully synthesized, which has a C 3-symmetric conformation and is capable of binding alkylammonium and metal ions simultaneously in a cooperative fashion. It can bind alkylammonium ions through the π-cavity formed by three aryl rings. This behaviour is consistent with the cone-in/cone-out conformational rearrangement needed to reorganize the cavity for endo-complexation. As a C 3-symmetrical pyridyl-substituted calixarene, receptor cone-1 can also bind an Ag + ion, and the nitrogen atoms are turned towards the inside of the cavity and interact with Ag +. After complexation of tris(2-pyridylamide) derivativemore » receptor cone-1 with Ag +, the original C 3-symmetry was retained and higher complexation selectivity for n-BuNH 3 + versus t-BuNH 3 + was observed. Thus, it is believed that this receptor will have a role to play in the sensing, detection, and recognition of Ag + and n-BuNH 3 + ions.« less

Unexpected Interactions of the Cyanobacterial Metallothionein SmtA with Uranium.

PubMed

Acharya, Celin; Blindauer, Claudia A

2016-02-15

Molecules for remediating or recovering uranium from contaminated environmental resources are of high current interest, with protein-based ligands coming into focus recently. Metallothioneins either bind or redox-silence a range of heavy metals, conferring protection against metal stress in many organisms. Here, we report that the cyanobacterial metallothionein SmtA competes with carbonate for uranyl binding, leading to formation of heterometallic (UO2)(n)Zn4SmtA species, without thiol oxidation, zinc loss, or compromising secondary or tertiary structure of SmtA. In turn, only metalated and folded SmtA species were found to be capable of uranyl binding. (1)H NMR studies and molecular modeling identified Glu34/Asp38 and Glu12/C-terminus as likely adventitious, but surprisingly strong, bidentate binding sites. While it is unlikely that these interactions correspond to an evolved biological function of this metallothionein, their occurrence may offer new possibilities for designing novel multipurpose bacterial metallothioneins with dual ability to sequester both soft metal ions including Cu(+), Zn(2+), Cd(2+), Hg(2+), and Pb(2+) and hard, high-oxidation state heavy metals such as U(VI). The concomitant protection from the chemical toxicity of uranium may be valuable for the development of bacterial strains for bio-remediation.

Analysis of Multiplexed Nanosensor Arrays Based on Near-Infrared Fluorescent Single-Walled Carbon Nanotubes.

PubMed

Dong, Juyao; Salem, Daniel P; Sun, Jessica H; Strano, Michael S

2018-04-24

The high-throughput, label-free detection of biomolecules remains an important challenge in analytical chemistry with the potential of nanosensors to significantly increase the ability to multiplex such assays. In this work, we develop an optical sensor array, printable from a single-walled carbon nanotube/chitosan ink and functionalized to enable a divalent ion-based proximity quenching mechanism for transducing binding between a capture protein or an antibody with the target analyte. Arrays of 5 × 6, 200 μm near-infrared (nIR) spots at a density of ≈300 spots/cm 2 are conjugated with immunoglobulin-binding proteins (proteins A, G, and L) for the detection of human IgG, mouse IgM, rat IgG2a, and human IgD. Binding kinetics are measured in a parallel, multiplexed fashion from each sensor spot using a custom laser scanning imaging configuration with an nIR photomultiplier tube detector. These arrays are used to examine cross-reactivity, competitive and nonspecific binding of analyte mixtures. We find that protein G and protein L functionalized sensors report selective responses to mouse IgM on the latter, as anticipated. Optically addressable platforms such as the one examined in this work have potential to significantly advance the real-time, multiplexed biomolecular detection of complex mixtures.

[Propranolol beta-blocker decrease in the concentration of high-affinity binding sites for calcium ions by sarcolemma membranes of the rat heart].

PubMed

Seleznev, Iu M; Martynov, A V; Smirnov, V N

1982-05-01

In vivo administration of propranolol considerably inhibits the isoproterenol-stimulated increase in 45Ca accumulation by the myocardium and completely eliminates the potentiation of isoproterenol effect by hydrocortisone. A significant lowering of the concentration of high affinity binding sites for calcium in the sarcolemmal membranes can be produced by propranolol in vitro. Under these conditions, the glucocorticoids do not change the sarcolemmal Ca2+-binding parameters or modulate the propranolol effect. Therefore, for the manifestation of glucocorticoid action to be brought about, the integrity of the cells is apparently required, while propranolol seems to change calcium binding by direct interaction with the sarcolemmal membranes. It is suggested that in vivo propranolol inhibition of catecholamine effect on calcium ion accumulation by the myocardium depends on the interaction with the beta-receptors and direct modulation of the concentration of high affinity binding sites for calcium ions on the surface of the sarcolemma.

Binding of trivalent chromium to serum transferrin is sufficiently rapid to be physiologically relevant.

PubMed

Deng, Ge; Wu, Kristi; Cruce, Alex A; Bowman, Michael K; Vincent, John B

2015-02-01

Transferrin, the major iron transport protein in the blood, also transports trivalent chromium in vivo. Recent in vitro studies have, however, suggested that the binding of chromic ions to apotransferrin is too slow to be biologically relevant. Nevertheless, the in vitro studies have generally failed to adequately take physiological bicarbonate concentrations into account. In aqueous buffer (with ambient (bi)carbonate concentrations), the binding of chromium to transferrin is too slow to be physiologically relevant, taking days to reach equilibrium with the protein's associated conformational changes. However, in the presence of 25mM (bi)carbonate, the concentration in human blood, chromic ions bind rapidly and tightly to transferrin. Details of the kinetics of chromium binding to human serum transferrin and conalbumin (egg white transferrin) in the presence of bicarbonate and other major potential chromium ligands are described and are consistent with transferrin being the major chromic ion transporter from the blood to tissues. Copyright © 2014 Elsevier Inc. All rights reserved.

A Secondary Structural Transition in the C-helix Promotes Gating of Cyclic Nucleotide-regulated Ion Channels*

PubMed Central

Puljung, Michael C.; Zagotta, William N.

2013-01-01

Cyclic nucleotide-regulated ion channels bind second messengers like cAMP to a C-terminal domain, consisting of a β-roll, followed by two α-helices (B- and C-helices). We monitored the cAMP-dependent changes in the structure of the C-helix of a C-terminal fragment of HCN2 channels using transition metal ion FRET between fluorophores on the C-helix and metal ions bound between histidine pairs on the same helix. cAMP induced a change in the dimensions of the C-helix and an increase in the metal binding affinity of the histidine pair. cAMP also caused an increase in the distance between a fluorophore on the C-helix and metal ions bound to the B-helix. Stabilizing the C-helix of intact CNGA1 channels by metal binding to a pair of histidines promoted channel opening. These data suggest that ordering of the C-helix is part of the gating conformational change in cyclic nucleotide-regulated channels. PMID:23525108

Colorimetric detection of copper in water using a Schiff base derivative

NASA Astrophysics Data System (ADS)

Peralta Domínguez, D.; Ramos-Ortiz, G.; Maldonado, J. L.; Rodriguez, M.; Meneses-Nava, M. A.; Barbosa-Garcia, O.; Santillan, R.; Farfán, N.

2013-09-01

Organic molecular sensors have the advantage of being used through an easy, fast, economical and reliable optical method for detecting toxic metal ions in our environment. In this work, we present a simple but highly specific organic ligand compound 5-Chloro-2-((E)-((E)-3-(4-(dimethylamino)phenyl)allylidene)amino)phenol (L1) that acts as a colorimetric sensor for ions in a mixture of acetonitrile/water (ratio 10:1, v:v). Binding interaction between L1 and various metal-ions has been established by ultraviolet-visible spectroscopic measurements that indicate favorable coordination of the ligand with selective metal ions, particularly, with copper. These results showed that the electronic transition band shape of L1 change after binding with copper in aqueous solution. L1 exhibited binding-induced color changes from yellow to pink one detected by the naked eye. This new sensor presented 2.5 × 10-6 M as limit detection, even under the presence of other metal ions.

HD Exchange and PLIMSTEX Determine the Affinities and Order of Binding of Ca2+ with Troponin C

PubMed Central

Huang, Richard Y-C.; Rempel, Don L.; Gross, Michael L.

2011-01-01

Troponin C (TnC), present in all striated muscle, is the Ca2+-activated trigger that initiates myocyte contraction. The binding of Ca2+ to TnC initiates a cascade of conformational changes involving the constituent proteins of the thin filament. The functional properties of TnC and its ability to bind Ca2+ have significant regulatory influence on the contractile reaction of muscle. Changes in TnC may also correlate with cardiac and various other muscle-related diseases. We report here the implementation of the PLIMSTEX strategy (Protein Ligand Interaction by Mass Spectrometry, Titration and H/D Exchange) to elucidate the binding affinity of TnC with Ca2+ and, more importantly, to determine the order of Ca2+ binding of the four EF hands of the protein. The four equilibrium constants, K1 = (5 ± 5) × 10 M-1, K2 = (1.8 ± 0.8) × 107 M-1, K3 = (4.2 ± 0.9) × 106 M-1, and K4 = (1.6 ± 0.6) × 106 M-1, agree well with determinations by other methods and serve to increase our confidence in the PLIMSTEX approach. We determined the order of binding to the four EF hands to be III, IV, II, and I by extracting from the H/DX results the deuterium patterns for each EF hand for each state of the protein (apo through fully Ca2+ bound). This approach, demonstrated for the first time, may be general for determining binding orders of metal ions and other ligands to proteins. PMID:21574565

Modulation of FadR Binding Capacity for Acyl-CoA Fatty Acids Through Structure-Guided Mutagenesis

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

Bacik, John-Paul; Yeager, Chris M.; Twary, Scott N.

FadR is a versatile global regulator in Escherichia coli that controls fatty acid metabolism and thereby modulates the ability of this bacterium to grow using fatty acids or acetate as the sole carbon source. FadR regulates fatty acid metabolism in response to intra-cellular concentrations of acyl-CoA lipids. The ability of FadR to bind acyl-CoA fatty acids is hence of significant interest for the engineering of biosynthetic pathways for the production of lipid-based biofuels and commodity chemicals. Based on the available crystal structure of E. coli bound to myristoyl- CoA, we predicted amino acid positions within the effector binding pocket thatmore » would alter the ability of FadR to bind acyl-CoA fatty acids without affecting DNA binding. We utilized fluorescence polarization to characterize the in-vitro binding properties of wild type and mutant FadR. We found that a Leu102Ala mutant enhanced binding of the effector, likely by increasing the size of the binding pocket for the acyl moiety of the molecule. Conversely, the elimination of the guanidine side chain (Arg213Ala and Arg213Met mutants) of the CoA moiety binding site severely diminished the ability of FadR to bind the acyl-CoA effector. These results demonstrate the ability to fine tune FadR binding capacity. The validation of an efficient method to fully characterize all the binding events involved in the specific activity (effector and DNA operator binding) of FadR has allowed us to increase our understanding of the role of specific amino acids in the binding and recognition of acyl-CoA fatty acids and will greatly facilitate efforts aimed at engineering tunable FadR regulators for synthetic biology.« less

Modulation of FadR Binding Capacity for Acyl-CoA Fatty Acids Through Structure-Guided Mutagenesis

DOE PAGES

Bacik, John-Paul; Yeager, Chris M.; Twary, Scott N.; ...

2015-09-18

FadR is a versatile global regulator in Escherichia coli that controls fatty acid metabolism and thereby modulates the ability of this bacterium to grow using fatty acids or acetate as the sole carbon source. FadR regulates fatty acid metabolism in response to intra-cellular concentrations of acyl-CoA lipids. The ability of FadR to bind acyl-CoA fatty acids is hence of significant interest for the engineering of biosynthetic pathways for the production of lipid-based biofuels and commodity chemicals. Based on the available crystal structure of E. coli bound to myristoyl- CoA, we predicted amino acid positions within the effector binding pocket thatmore » would alter the ability of FadR to bind acyl-CoA fatty acids without affecting DNA binding. We utilized fluorescence polarization to characterize the in-vitro binding properties of wild type and mutant FadR. We found that a Leu102Ala mutant enhanced binding of the effector, likely by increasing the size of the binding pocket for the acyl moiety of the molecule. Conversely, the elimination of the guanidine side chain (Arg213Ala and Arg213Met mutants) of the CoA moiety binding site severely diminished the ability of FadR to bind the acyl-CoA effector. These results demonstrate the ability to fine tune FadR binding capacity. The validation of an efficient method to fully characterize all the binding events involved in the specific activity (effector and DNA operator binding) of FadR has allowed us to increase our understanding of the role of specific amino acids in the binding and recognition of acyl-CoA fatty acids and will greatly facilitate efforts aimed at engineering tunable FadR regulators for synthetic biology.« less

Ice restructuring inhibition activities in antifreeze proteins with distinct differences in thermal hysteresis.

PubMed

Yu, Sally O; Brown, Alan; Middleton, Adam J; Tomczak, Melanie M; Walker, Virginia K; Davies, Peter L

2010-12-01

Antifreeze proteins (AFPs) share two related properties: the ability to depress the freezing temperature below the melting point of ice (thermal hysteresis; TH); and the ability to inhibit the restructuring of ice into larger crystals. Since the 'hyperactive' AFPs, which have been more recently discovered, show an order of magnitude more TH than previously characterized AFPs, we have now determined their activities in ice restructuring inhibition (IrI) assays. IrI activities of three TH-hyperactive AFPs and three less TH-active AFPs varied over an 8-fold range. There was no obvious correlation between high TH activity and high IrI activity. However, the use of mutant AFPs demonstrated that severe disruption of ice-binding residues diminished both TH and IrI similarly, revealing that that the same ice-binding residues are crucial for both activities. In addition, bicarbonate ions, which are known to enhance the TH activity of AFPs, also enhanced their IrI activity. We suggest that these seemingly contradictory observations can be partially explained by differences in the coverage of ice by TH-hyperactive and non-hyperactive AFPs, and by differences in the stability of AFP-bound ice under supercooled and recrystallization conditions. Copyright © 2010 Elsevier Inc. All rights reserved.

Fluorescence lifetime analysis and effect of magnesium ions on binding of NADH to human aldehyde dehydrogenase 1

USDA-ARS?s Scientific Manuscript database

Aldehyde dehydrogenase 1 (ALDH1) catalyzes oxidation of toxic aldehydes to carboxylic acids. Physiologic levels of Mg2+ ions influence ALDH1 activity in part by increasing NADH binding affinity to the enzyme thus reducing activity. By using time-resolved fluorescence spectroscopy, we have resolved t...

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.

Tetrodotoxin- and tributyltin-binding abilities of recombinant pufferfish saxitoxin and tetrodotoxin binding proteins of Takifugu rubripes.

PubMed

Satone, Hina; Nonaka, Shohei; Lee, Jae Man; Shimasaki, Yohei; Kusakabe, Takahiro; Kawabata, Shun-Ichiro; Oshima, Yuji

2017-01-01

We investigated the ability of recombinant pufferfish saxitoxin and tetrodotoxin binding protein types 1 and 2 of Takifugu rubripes (rTrub.PSTBP1 and rTrub.PSTBP2) to bind to tetrodotoxin (TTX) and tributyltin. Both rTrub.PSTBPs bound to tributyltin in an ultrafiltration binding assay but lost this ability on heat denaturation. In contrast, only rTrub.PSTBP2 bound to TTX even heat denaturation. This result suggests that the amino acid sequence of PSTBP2 may be contributed for its affinity for TTX. Copyright © 2016 Elsevier Ltd. All rights reserved.

[Effect of calcium and magnesium ions on the interaction of corticosterone with the cytosol receptor(s) in the rat brain].

PubMed

Ueda, M

1981-01-01

The effects of calcium and magnesium ions on the corticosterone binding to rat brain cytosol receptor protein(s) were investigated. The increasing amounts of CaCl2 or MgCl2 up to 5.0 mM were added, the specific [3H] corticosterone binding increased 1.3-fold and 1.5 respectively. The addition of MnCl2 and KCl did not affect this binding. The binding of corticosterone with rat brain cytosol receptor(s) were decreased by increasing amounts of EDTA and complete inhibition was observed at concentration equal to and greater than 2.5 mM. Inhibition of this binding by EDTA was less than by EGTA. Either theophylline or dibutyryl cyclic AMP had no effect on this binding.

Energetics of Glutamate Binding to an Ionotropic Glutamate Receptor.

PubMed

Yu, Alvin; Lau, Albert Y

2017-11-22

Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that are responsible for the majority of excitatory transmission at the synaptic cleft. Mechanically speaking, agonist binding to the ligand binding domain (LBD) activates the receptor by triggering a conformational change that is transmitted to the transmembrane region, opening the ion channel pore. We use fully atomistic molecular dynamics simulations to investigate the binding process in the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, an iGluR subtype. The string method with swarms of trajectories was applied to calculate the possible pathways glutamate traverses during ligand binding. Residues peripheral to the binding cleft are found to metastably bind the ligand prior to ligand entry into the binding pocket. Umbrella sampling simulations were performed to compute the free energy barriers along the binding pathways. The calculated free energy profiles demonstrate that metastable interactions contribute substantially to the energetics of ligand binding and form local minima in the overall free energy landscape. Protein-ligand interactions at sites outside of the orthosteric agonist-binding site may serve to lower the transition barriers of the binding process.

Relation between heat of vaporization, ion transport, molar volume, and cation-anion binding energy for ionic liquids.

PubMed

Borodin, Oleg

2009-09-10

A number of correlations between heat of vaporization (H(vap)), cation-anion binding energy (E(+/-)), molar volume (V(m)), self-diffusion coefficient (D), and ionic conductivity for 29 ionic liquids have been investigated using molecular dynamics (MD) simulations that employed accurate and validated many-body polarizable force fields. A significant correlation between D and H(vap) has been found, while the best correlation was found for -log(DV(m)) vs H(vap) + 0.28E(+/-). A combination of enthalpy of vaporization and a fraction of the cation-anion binding energy was suggested as a measure of the effective cohesive energy for ionic liquids. A deviation of some ILs from the reported master curve is explained based upon ion packing and proposed diffusion pathways. No general correlations were found between the ion diffusion coefficient and molecular volume or the diffusion coefficient and cation/anion binding energy.

Self-consistent field theory of polymer-ionic molecule complexation.

PubMed

Nakamura, Issei; Shi, An-Chang

2010-05-21

A self-consistent field theory is developed for polymers that are capable of binding small ionic molecules (adsorbates). The polymer-ionic molecule association is described by Ising-like binding variables, C(i) ((a))(kDelta)(=0 or 1), whose average determines the number of adsorbed molecules, n(BI). Polymer gelation can occur through polymer-ionic molecule complexation in our model. For polymer-polymer cross-links through the ionic molecules, three types of solutions for n(BI) are obtained, depending on the equilibrium constant of single-ion binding. Spinodal lines calculated from the mean-field free energy exhibit closed-loop regions where the homogeneous phase becomes unstable. This phase instability is driven by the excluded-volume interaction due to the single occupancy of ion-binding sites on the polymers. Moreover, sol-gel transitions are examined using a critical degree of conversion. A gel phase is induced when the concentration of adsorbates is increased. At a higher concentration of the adsorbates, however, a re-entrance from a gel phase into a sol phase arises from the correlation between unoccupied and occupied ion-binding sites. The theory is applied to a model system, poly(vinyl alcohol) and borate ion in aqueous solution with sodium chloride. Good agreement between theory and experiment is obtained.

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

Binding site in eag voltage sensor accommodates a variety of ions and is accessible in closed channel.

PubMed

Silverman, William R; Bannister, John P A; Papazian, Diane M

2004-11-01

In ether-a-go-go K+ channels, voltage-dependent activation is modulated by ion binding to a site located in an extracellular-facing crevice between transmembrane segments S2 and S3 in the voltage sensor. We find that acidic residues D278 in S2 and D327 in S3 are able to coordinate a variety of divalent cations, including Mg2+, Mn2+, and Ni2+, which have qualitatively similar functional effects, but different half-maximal effective concentrations. Our data indicate that ions binding to individual voltage sensors in the tetrameric channel act without cooperativity to modulate activation gating. We have taken advantage of the unique phenotype of Ni2+ in the D274A channel, which contains a mutation of a nonbinding site residue, to demonstrate that ions can access the binding site from the extracellular solution when the voltage sensor is in the resting conformation. Our results are difficult to reconcile with the x-ray structure of the KvAP K+ channel, in which the binding site residues are widely separated, and with the hydrophobic paddle model for voltage-dependent activation, in which the voltage sensor domain, including the S3-S4 loop, is near the cytoplasmic side of the membrane in the closed channel.

Tracing Cytoplasmic Ca2+ Ion and Water Access Points in the Ca2+-ATPase

PubMed Central

Musgaard, Maria; Thøgersen, Lea; Schiøtt, Birgit; Tajkhorshid, Emad

2012-01-01

Sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) transports two Ca2+ ions across the membrane of the sarco(endo)plasmic reticulum against the concentration gradient, harvesting the required energy by hydrolyzing one ATP molecule during each transport cycle. Although SERCA is one of the best structurally characterized membrane transporters, it is still largely unknown how the transported Ca2+ ions reach their transmembrane binding sites in SERCA from the cytoplasmic side. Here, we performed extended all-atom molecular dynamics simulations of SERCA. The calculated electrostatic potential of the protein reveals a putative mechanism by which cations may be attracted to and bind to the Ca2+-free state of the transporter. Additional molecular dynamics simulations performed on a Ca2+-bound state of SERCA reveal a water-filled pathway that may be used by the Ca2+ ions to reach their buried binding sites from the cytoplasm. Finally, several residues that are involved in attracting and guiding the cations toward the possible entry channel are identified. The results point to a single Ca2+ entry site close to the kinked part of the first transmembrane helix, in a region loaded with negatively charged residues. From this point, a water pathway outlines a putative Ca2+ translocation pathway toward the transmembrane ion-binding sites. PMID:22339863

A novel biphenolic ligand for selective Mg2+ and Zn2+ ions sensing followed by colorimetric, spectroscopic and cell imaging methods.

PubMed

Maheswari, Palanisamy Uma; Renuga, Duraisamy; Henry, Linda Jeeva Kumari; Ruckmani, Kandasamy

2018-04-30

The (E)-2-((2-hydrohy-5-methylphenylimino) methyl) phenol ligand was synthesized. The receptor was characterized by IR, 1 H and 13 C NMR and CHN analysis. The ligand exhibits colorimetric and fluorometric sensing of Zn 2+ and Mg 2+ ions in semi-aqueous medium (DMSO-H2O). The receptor was tested with series of transition metal ions (Cr 2+ , Fe 2+ , Ni 2+ , Co 2+ , Cu 2+ , Zn 2+ ) and heavy metal ions (Sn 2+ , Pd 2+ , Ce 2+ , Hg 2+ , Cd 2+ ) and the essential human body elements like Ca 2+ , Mg 2+ , Na + and K + ions. The naked eye colorimetric sensing was absorbed only for Zn 2+ and Mg 2+ . Both ions (ZnCl 2 and MgCl 2 in H 2 O), when added to the colorless solutions of the receptor of about 1 equivalence in incremental additions turn the solution into bright turmeric yellow. All other ions remain inactive, in colorimetric sensing. Further the Zn 2+ and Mg 2+ ions were probed by absorption and emission spectroscopy through incremental addition of respective metal ions. The in-situ deprotonation of the ligand on both Mg 2+ and Zn 2+ ions binding was confirmed by 1 H NMR titration studies. The imino nitrogen of the receptor is not coordinated to the metal ions. The Job's plot studies reveal the 1:2 binding ratio of metal ions to the receptor. The high fold fluorescence output on metal ions binding was positively used to sense the Zn 2+ and Mg 2+ ions, separately and together in HeLa cancer cells through cell imaging. Copyright © 2018 Elsevier B.V. All rights reserved.

The second Cu(II)-binding site in a proton-rich environment interferes with the aggregation of amyloid-beta(1-40) into amyloid fibrils.

PubMed

Jun, Sangmi; Gillespie, Joel R; Shin, Byong-kyu; Saxena, Sunil

2009-11-17

The overall morphology and Cu(II) ion coordination for the aggregated amyloid-beta(1-40) [Abeta(1-40)] in N-ethylmorpholine (NEM) buffer are affected by Cu(II) ion concentration. This effect is investigated by transmission electron microscopy (TEM), atomic force microscopy (AFM), and electron spin echo envelope modulation (ESEEM) spectroscopy. At lower than equimolar concentrations of Cu(II) ions, fibrillar aggregates of Abeta(1-40) are observed. At these concentrations of Cu(II), the monomeric and fibrillar Abeta(1-40) ESEEM data indicate that the Cu(II) ion is coordinated by histidine residues. For aggregated Abeta(1-40) at a Cu(II):Abeta molar ratio of 2:1, TEM and AFM images show both linear fibrils and granular amorphous aggregates. The ESEEM spectra show that the multi-histidine coordination for Cu(II) ion partially breaks up and becomes exposed to water or exchangeable protons of the peptide at a higher Cu(II) concentration. Since the continuous-wave electron spin resonance results also suggest two copper-binding sites in Abeta(1-40), the proton ESEEM peak may arise from the second copper-binding site, which may be significantly involved in the formation of granular amorphous aggregates. Thioflavin T fluorescence and circular dichroism experiments also show that Cu(II) inhibits the formation of fibrils and induces a nonfibrillar beta-sheet conformation. Therefore, we propose that Abeta(1-40) has a second copper-binding site in a proton-rich environment and the second binding Cu(II) ion interferes with a conformational transition into amyloid fibrils, inducing the formation of granular amorphous aggregates.

Stimulation of eryptosis by aluminium ions

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

Niemoeller, Olivier M.; Kiedaisch, Valentin; Dreischer, Peter

2006-12-01

Aluminium salts are utilized to impede intestinal phosphate absorption in chronic renal failure. Toxic side effects include anemia, which could result from impaired formation or accelerated clearance of circulating erythrocytes. Erythrocytes may be cleared secondary to suicidal erythrocyte death or eryptosis, which is characterized by cell shrinkage and exposure of phosphatidylserine (PS) at the erythrocyte surface. As macrophages are equipped with PS receptors, they bind, engulf and degrade PS-exposing cells. The present experiments have been performed to explore whether Al{sup 3+} ions trigger eryptosis. The PS exposure was estimated from annexin binding and cell volume from forward scatter in FACSmore » analysis. Exposure to Al{sup 3+} ions ({>=} 10 {mu}M Al{sup 3+} for 24 h) indeed significantly increased annexin binding, an effect paralleled by decrease of forward scatter at higher concentrations ({>=} 30 {mu}M Al{sup 3+}). According to Fluo3 fluorescence Al{sup 3+} ions ({>=} 30 {mu}M for 3 h) increased cytosolic Ca{sup 2+} activity. Al{sup 3+} ions ({>=} 10 {mu}M for 24 h) further decreased cytosolic ATP concentrations. Energy depletion by removal of glucose similarly triggered annexin binding, an effect not further enhanced by Al{sup 3+} ions. The eryptosis was paralleled by release of hemoglobin, pointing to loss of cell membrane integrity. In conclusion, Al{sup 3+} ions decrease cytosolic ATP leading to activation of Ca{sup 2+}-permeable cation channels, Ca{sup 2+} entry, stimulation of cell membrane scrambling and cell shrinkage. Moreover, Al{sup 3+} ions lead to loss of cellular hemoglobin, a feature of hemolysis. Both effects are expected to decrease the life span of circulating erythrocytes and presumably contribute to the development of anemia during Al{sup 3+} intoxication.« less

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.

Does alpha 1-acid glycoprotein act as a non-functional receptor for alpha 1-adrenergic antagonists?

PubMed

Qin, M; Oie, S

1994-11-01

The ability of a variety of alpha 1-acid glycoproteins (AAG) to affect the intrinsic activity of the alpha 1-adrenergic antagonist prazosin was studied in rabbit aortic strip preparations. From these studies, the activity of AAG appears to be linked to their ability to bind the antagonist. However, a capability to bind prazosin was not the only requirement for this effect. The removal of sialic acid and partial removal of the galactose and mannose residues by periodate oxidation of human AAG all but eliminated the ability of AAG to affect the intrinsic pharmacologic activity of prazosin, although the binding of prazosin was not significantly affected. The presence of bovine AAG, a protein that has a low ability to bind prazosin, reduced the effect of human AAG on prazosin activity. Based upon these results, we propose that AAG is able to bind in the vicinity of the alpha 1-adrenoceptors, therefore extending the binding region for antagonists in such a way as to decrease the ability of the antagonist to interact with the receptor. The carbohydrate side-chains are important for the binding of AAG in the region of the adrenoceptor.

Determination of copper binding in Pseudomonas putida CZ1 by chemical modifications and X-ray absorption spectroscopy.

PubMed

Chen, XinCai; Shi, JiYan; Chen, YingXu; Xu, XiangHua; Chen, LiTao; Wang, Hui; Hu, TianDou

2007-03-01

Previously performed studies have shown that Pseudomonas putida CZ1 biomass can bind an appreciable amount of Cu(II) and Zn(II) ions from aqueous solutions. The mechanisms of Cu- and Zn-binding by P. putida CZ1 were ascertained by chemical modifications of the biomass followed by Fourier transform infrared and X-ray absorption spectroscopic analyses of the living or nonliving cells. A dramatic decrease in Cu(II)- and Zn(II)-binding resulted after acidic methanol esterification of the nonliving cells, indicating that carboxyl functional groups play an important role in the binding of metal to the biomaterial. X-ray absorption spectroscopy was used to determine the speciation of Cu ions bound by living and nonliving cells, as well as to elucidate which functional groups were involved in binding of the Cu ions. The X-ray absorption near-edge structure spectra analysis showed that the majority of the Cu was bound in both samples as Cu(II). The fitting results of Cu K-edge extended X-ray absorption fine structure spectra showed that N/O ligands dominated in living and nonliving cells. Therefore, by combining different techniques, our results indicate that carboxyl functional groups are the major ligands responsible for the metal binding in P. putida CZ1.

Evaluation of Nonferrous Metals as Potential In Vivo Tracers of Transferrin-Based Therapeutics

NASA Astrophysics Data System (ADS)

Zhao, Hanwei; Wang, Shunhai; Nguyen, Son N.; Elci, S. Gokhan; Kaltashov, Igor A.

2016-02-01

Transferrin (Tf) is a promising candidate for targeted drug delivery. While development of such products is impossible without the ability to monitor biodistribution of Tf-drug conjugates in tissues and reliable measurements of their levels in blood and other biological fluids, the presence of very abundant endogenous Tf presents a significant impediment to such efforts. Several noncognate metals have been evaluated in this work as possible tracers of exogenous transferrin in complex biological matrices using inductively coupled plasma mass spectrometry (ICP MS) as a detection tool. Placing Ni(II) on a His-tag of recombinant Tf resulted in formation of a marginally stable protein-metal complex, which readily transfers the metal to ubiquitous physiological scavengers, such as serum albumin. An alternative strategy targeted iron-binding pockets of Tf, where cognate Fe(III) was replaced by metal ions known to bind this protein. Both Ga(III) and In(III) were evaluated, with the latter being vastly superior as a tracer (stronger binding to Tf unaffected by the presence of metal scavengers and the retained ability to associate with Tf receptor). Spiking serum with indium-loaded Tf followed by ICP MS detection demonstrated that protein quantities as low as 0.04 nM can be readily detected in animal blood. Combining laser ablation with ICP MS detection allows distribution of exogenous Tf to be mapped within animal tissue cross-sections with spatial resolution exceeding 100 μm. The method can be readily extended to a range of other therapeutics where metalloproteins are used as either carriers or payloads.

Identification of Escherichia coli enterotoxin inhibitors from traditional medicinal herbs by in silico, in vitro, and in vivo analyses.

PubMed

Chen, Jaw-Chyun; Ho, Tin-Yun; Chang, Yuan-Shiun; Wu, Shih-Lu; Li, Chia-Cheng; Hsiang, Chien-Yun

2009-01-30

Glycyrrhiza uralensis has been used for the treatment of gastrointestinal disorders, such as diarrhea, in several ancient cultures. Glycyrrhizin is the principal component of liquorice and lots of pharmacological effects have been demonstrated. Heat-labile enterotoxin (LT), the virulence factor of enterotoxigenic Escherichia coli, induces diarrhea by initially binding to the GM1 on the surfaces of intestinal epithelial cells and consequently leading to the massive loss of fluid and ions from cells. Therefore, we evaluated the inhibitory effects of traditional medicinal herbs (TMH) on the B subunit of LT (LTB) and GM1 interaction. The inhibitory effects of TMH on LTB-GM1 interaction were evaluated by GM1-enzyme-linked immunosorbent assay (ELISA). The likely active phytochemicals of these TMH were then predicted by in silico model (docking) and analyzed by in vitro (GM1-ELISA) and in vivo (patent mouse gut assay) models. We found that various TMH, which have been ethnomedically used for the treatment of diarrhea, inhibited the LTB-GM1 interaction. Docking data showed that triterpenoids were the most active phytochemicals and the oleanane-type triterpenoids presented better LTB-binding abilities than other types of triterpenoids. Moreover, by in vitro and in vivo models, we demonstrated that glycyrrhizin was the most effective oleanane-type triterpenoid that significantly suppressed both the LTB-binding ability (IC50=3.26+/-0.17 mM) and the LT-induced fluid accumulation in mice. We found an LT inhibitor, glycyrrhizin, from TMH by in silico, in vitro, and in vivo analyses.

Direct sensing of fluoride in aqueous solutions using a boronic acid based sensor.

PubMed

Wu, Xin; Chen, Xuan-Xuan; Song, Bing-Nan; Huang, Yan-Jun; Ouyang, Wen-Juan; Li, Zhao; James, Tony D; Jiang, Yun-Bao

2014-11-21

Binding of the fluoride ion triggers aggregation of a pyreneboronic acid-catechol ensemble in acidic aqueous solutions, giving rise to intense excimer emission, allowing for sensitive fluoride ion sensing at ppm levels, with an apparent fluoride binding constant higher than 10(3) M(-1) which is unprecedented for boronic acid sensors in water.

Binding patterns of vanadium ions with different valence states to human serum transferrin studied by HPLC/high-resolution ICP-MS.

PubMed

Nagaoka, Megumi Hamano; Yamazaki, Takeshi; Maitani, Tamio

2002-09-06

Vanadium (V) is an essential metal for mammals and has different valence states. In blood, V is bound to serum transferrin (Tf), a glycoprotein which has two metal-binding sites, and carbonate is generally required for the binding. In this study, the binding patterns of V(III), V(IV), and V(V) to human serum Tf (hTf) were analyzed using an HPLC system equipped with an anion-exchange column and directly connected to a high-resolution inductively coupled plasma-mass spectrometer for metal detection (51V). In affinity to hTf, the three ions were ranked V(III)>V(IV)>V(V) in the presence of bicarbonate and V(III) reverse congruent V(IV)>V(V) in the absence. Intermediates in the "open forms" binding to the respective sites were detected at the initial stage. V(IV) and V(V) were bound to the N-lobe site in the "closed form" and "open form," respectively. In the absence of bicarbonate, V ions with respective valence states were bound to hTf in the "open form." In terms of binding to hTf, tri-valent V was most favorable in the presence of bicarbonate.

Structural, energetic, spectroscopic and QTAIM analyses of cation-π interactions involving mono- and bi-cyclic ring fused benzene systems.

PubMed

Hassan, Ayorinde; Dinadayalane, Tandabany C; Grabowski, Sławomir J; Leszczynski, Jerzy

2013-12-28

The effect of increasing the number of monocyclic six-membered rings or bicyclic rings of bicyclo[2.1.1]hexenyl fused to benzene on cation-π interactions involving alkali metal ions (Li(+), Na(+), and K(+)) has been investigated. The binding energy data at the B3LYP/6-311+G(2d,2p) level clearly indicate that the binding affinity of the metal ion with benzene is enhanced by increasing the number of rings fused irrespective of a monocyclic or a bicyclic ring. Calculated binding energies are in good agreement with the available experimental results. The binding strength of cations with ligands decreases in the order Li(+) > Na(+) > K(+). Our study establishes that trisannelation of bicyclo[2.1.1]hexene to benzene facilitates a very strong interaction between benzene and cations. Infrared (IR) frequencies and nuclear magnetic resonance (NMR) chemical shifts are shown to be valuable in characterizing cation-π interactions. The C-C bonds of the central six-membered rings are weakened due to metal ion binding. Based on the Quantum Theory of Atoms in Molecules (QTAIM), we have observed the presence of stabilizing H∙∙∙H interactions in two of the considered systems as opposed to the frequent description of these interactions as non-bonded repulsive interactions. Alkali metal ion binding with those two ligands slightly reduces the strength of such H∙∙∙H interactions.

Quantum dot nanocrystals having guanosine imprinted nanoshell for DNA recognition.

PubMed

Diltemiz, Sibel Emir; Say, Ridvan; Büyüktiryaki, Sibel; Hür, Deniz; Denizli, Adil; Ersöz, Arzu

2008-05-30

Molecular imprinted polymers (MIPs) as a recognition element for sensors are increasingly of interest and MIP nanoparticles have started to appear in the literature. In this study, we have proposed a novel thiol ligand-capping method with polymerizable methacryloylamido-cysteine (MAC) attached to CdS quantum dots (QDs), reminiscent of a self-assembled monolayer and have reconstructed surface shell by synthetic host polymers based on molecular imprinting method for DNA recognition. In this method, methacryloylamidohistidine-platinium (MAH-Pt(II)) is used as a new metal-chelating monomer via metal coordination-chelation interactions and guanosine templates of DNA. Nanoshell sensors with guanosine templates give a cavity that is selective for guanosine and its analogues. The guanosine can simultaneously chelate to Pt(II) metal ion and fit into the shape-selective cavity. Thus, the interaction between Pt(II) ion and free coordination spheres has an effect on the binding ability of the CdS QD nanosensor. The binding affinity of the guanosine imprinted nanocrystals has investigated by using the Langmuir and Scatchard methods, and experiments have shown the shape-selective cavity formation with O6 and N7 of a guanosine nucleotide (K(a) = 4.841x10(6) mol L(-1)) and a free guanine base (K(a) = 0.894x10(6) mol L(-1)). Additionally, the guanosine template of the nanocrystals is more favored for single stranded DNA compared to double stranded DNA.

Metal dealing at the origin of the Chordata phylum: the metallothionein system and metal overload response in amphioxus.

PubMed

Guirola, Maria; Pérez-Rafael, Sílvia; Capdevila, Mercè; Palacios, Oscar; Atrian, Sílvia

2012-01-01

Non-vertebrate chordates, specifically amphioxus, are considered of the utmost interest for gaining insight into the evolutionary trends, i.e. differentiation and specialization, of gene/protein systems. In this work, MTs (metallothioneins), the most important metal binding proteins, are characterized for the first time in the cephalochordate subphylum at both gene and protein level, together with the main features defining the amphioxus response to cadmium and copper overload. Two MT genes (BfMT1 and BfMT2) have been identified in a contiguous region of the genome, as well as several ARE (antioxidant response element) and MRE (metal response element) located upstream the transcribed region. Their corresponding cDNAs exhibit identical sequence in the two lancelet species (B. floridae and B. lanceolatum), BfMT2 cDNA resulting from an alternative splicing event. BfMT1 is a polyvalent metal binding peptide that coordinates any of the studied metal ions (Zn, Cd or Cu) rendering complexes stable enough to last in physiological environments, which is fully concordant with the constitutive expression of its gene, and therefore, with a metal homeostasis housekeeping role. On the contrary, BfMT2 exhibits a clear ability to coordinate Cd(II) ions, while it is absolutely unable to fold into stable Cu (I) complexes, even as mixed species. This identifies it as an essential detoxification agent, which is consequently only induced in emergency situations. The cephalochordate MTs are not directly related to vertebrate MTs, neither by gene structure, protein similarity nor metal-binding behavior of the encoded peptides. The closest relative is the echinoderm MT, which confirm proposed phylogenetic relationships between these two groups. The current findings support the existence in most organisms of two types of MTs as for their metal binding preferences, devoted to different biological functions: multivalent MTs for housekeeping roles, and specialized MTs that evolve either as Cd-thioneins or Cu-thioneins, according to the ecophysiological needs of each kind of organisms.

Metal Dealing at the Origin of the Chordata Phylum: The Metallothionein System and Metal Overload Response in Amphioxus

PubMed Central

Capdevila, Mercè; Palacios, Òscar; Atrian, Sílvia

2012-01-01

Non-vertebrate chordates, specifically amphioxus, are considered of the utmost interest for gaining insight into the evolutionary trends, i.e. differentiation and specialization, of gene/protein systems. In this work, MTs (metallothioneins), the most important metal binding proteins, are characterized for the first time in the cephalochordate subphylum at both gene and protein level, together with the main features defining the amphioxus response to cadmium and copper overload. Two MT genes (BfMT1 and BfMT2) have been identified in a contiguous region of the genome, as well as several ARE (antioxidant response element) and MRE (metal response element) located upstream the transcribed region. Their corresponding cDNAs exhibit identical sequence in the two lancelet species (B. floridae and B. lanceolatum), BfMT2 cDNA resulting from an alternative splicing event. BfMT1 is a polyvalent metal binding peptide that coordinates any of the studied metal ions (Zn, Cd or Cu) rendering complexes stable enough to last in physiological environments, which is fully concordant with the constitutive expression of its gene, and therefore, with a metal homeostasis housekeeping role. On the contrary, BfMT2 exhibits a clear ability to coordinate Cd(II) ions, while it is absolutely unable to fold into stable Cu (I) complexes, even as mixed species. This identifies it as an essential detoxification agent, which is consequently only induced in emergency situations. The cephalochordate MTs are not directly related to vertebrate MTs, neither by gene structure, protein similarity nor metal-binding behavior of the encoded peptides. The closest relative is the echinoderm MT, which confirm proposed phylogenetic relationships between these two groups. The current findings support the existence in most organisms of two types of MTs as for their metal binding preferences, devoted to different biological functions: multivalent MTs for housekeeping roles, and specialized MTs that evolve either as Cd-thioneins or Cu-thioneins, according to the ecophysiological needs of each kind of organisms. PMID:22905252

Analysis of the actions of the novel dopamine receptor-directed compounds (S)-OSU6162 and ACR16 at the D2 dopamine receptor

PubMed Central

Kara, Elodie; Lin, Hong; Svensson, Kjell; Johansson, Anette M; Strange, Philip G

2010-01-01

BACKGROUND AND PURPOSE The two phenylpiperidines, OSU6162 and ACR16, have been proposed as novel drugs for the treatment of brain disorders, including schizophrenia and Huntington's disease, because of their putative dopamine stabilizing effects. Here we evaluated the activities of these compounds in a range of assays for the D2 dopamine receptor in vitro. EXPERIMENTAL APPROACH The affinities of these compounds for the D2 dopamine receptor were evaluated in competition with [3H]spiperone and [3H]NPA. Agonist activity of these compounds was evaluated in terms of their ability to stimulate [35S]GTPγS binding. KEY RESULTS Both compounds had low affinities for inhibition of [3H]spiperone binding (pKi vs. [3H]spiperone, ACR16: <5, OSU6162: 5.36). Neither compound was able to stimulate [35S]GTPγS binding when assayed in the presence of Na+ ions, but if the Na+ ions were removed, both compounds were low-affinity, partial agonists (Emax relative to dopamine: ACR16: 10.2%, OSU6162:54.3%). Schild analysis of the effects of OSU6162 to inhibit dopamine-stimulated [35S]GTPγS binding indicated Schild slopes of ∼0.9, suggesting little deviation from competitive inhibition. OSU6162 was, however, able to accelerate [3H]NPA dissociation from D2 dopamine receptors, indicating some allosteric effects of this compound. CONCLUSIONS AND IMPLICATIONS The two phenylpiperidines were low-affinity, low-efficacy partial agonists at the D2 dopamine receptor in vitro, possibly exhibiting some allosteric effects. Comparing their in vitro and in vivo effects, the in vitro affinities were a reasonable guide to potencies in vivo. However, the lack of in vitro–in vivo correlation for agonist efficacy needs to be further addressed. PMID:20804495

Calcium-dependent antigen binding as a novel modality for antibody recycling by endosomal antigen dissociation

PubMed Central

Hironiwa, N; Ishii, S; Kadono, S; Iwayanagi, Y; Mimoto, F; Habu, K; Igawa, T; Hattori, K

2016-01-01

The pH-dependent antigen binding antibody, termed a recycling antibody, has recently been reported as an attractive type of second-generation engineered therapeutic antibody. A recycling antibody can dissociate antigen in the acidic endosome, and thus bind to its antigen multiple times. As a consequence, a recycling antibody can neutralize large amounts of antigen in plasma. Because this approach relies on histidine residues to achieve pH-dependent antigen binding, which could limit the epitopes that can be targeted and affect the rate of antigen dissociation in the endosome, we explored an alternative approach for generating recycling antibodies. Since calcium ion concentration is known to be lower in endosome than in plasma, we hypothesized that an antibody with antigen-binding properties that are calcium-dependent could be used as recycling antibody. Here, we report a novel anti-interleukin-6 receptor (IL-6R) antibody, identified from a phage library that binds to IL-6R only in the presence of a calcium ion. Thermal dynamics and a crystal structure study revealed that the calcium ion binds to the heavy chain CDR3 region (HCDR3), which changes and possibly stabilizes the structure of HCDR3 to make it bind to antigen calcium dependently (PDB 5AZE). In vitro and in vivo studies confirmed that this calcium-dependent antigen-binding antibody can dissociate its antigen in the endosome and accelerate antigen clearance from plasma, making it a novel approach for generating recycling antibody. PMID:26496237

Molecular recognition of organic ammonium ions in solution using synthetic receptors

PubMed Central

Späth, Andreas

2010-01-01

Summary Ammonium ions are ubiquitous in chemistry and molecular biology. Considerable efforts have been undertaken to develop synthetic receptors for their selective molecular recognition. The type of host compounds for organic ammonium ion binding span a wide range from crown ethers to calixarenes to metal complexes. Typical intermolecular interactions are hydrogen bonds, electrostatic and cation–π interactions, hydrophobic interactions or reversible covalent bond formation. In this review we discuss the different classes of synthetic receptors for organic ammonium ion recognition and illustrate the scope and limitations of each class with selected examples from the recent literature. The molecular recognition of ammonium ions in amino acids is included and the enantioselective binding of chiral ammonium ions by synthetic receptors is also covered. In our conclusion we compare the strengths and weaknesses of the different types of ammonium ion receptors which may help to select the best approach for specific applications. PMID:20502608

Initial evaluation of 18F-GE-179, a putative PET Tracer for activated N-methyl D-aspartate receptors.

PubMed

McGinnity, Colm J; Hammers, Alexander; Riaño Barros, Daniela A; Luthra, Sajinder K; Jones, Paul A; Trigg, William; Micallef, Caroline; Symms, Mark R; Brooks, David J; Koepp, Matthias J; Duncan, John S

2014-03-01

N-methyl D-aspartate (NMDA) ion channels play a key role in a wide range of physiologic (e.g., memory and learning tasks) and pathologic processes (e.g., excitotoxicity). To date, suitable PET markers of NMDA ion channel activity have not been available. (18)F-GE-179 is a novel radioligand that selectively binds to the open/active state of the NMDA receptor ion channel, displacing the binding of (3)H-tenocyclidine from the intrachannel binding site with an affinity of 2.4 nM. No significant binding was observed with 10 nM GE-179 at 60 other neuroreceptors, channels, or transporters. We describe the kinetic behavior of the radioligand in vivo in humans. Nine healthy participants (6 men, 3 women; median age, 37 y) each underwent a 90-min PET scan after an intravenous injection of (18)F-GE-179. Continuous arterial blood sampling over the first 15 min was followed by discrete blood sampling over the duration of the scan. Brain radioactivity (KBq/mL) was measured in summation images created from the attenuation- and motion-corrected dynamic images. Metabolite-corrected parent plasma input functions were generated. We assessed the abilities of 1-, 2-, and 3-compartment models to kinetically describe cerebral time-activity curves using 6 bilateral regions of interest. Parametric volume-of-distribution (V(T)) images were generated by voxelwise rank-shaping regularization of exponential spectral analysis (RS-ESA). A 2-brain-compartment, 4-rate-constant model best described the radioligand's kinetics in normal gray matter of subjects at rest. At 30 min after injection, 37% of plasma radioactivity represented unmetabolized (18)F-GE-179. The highest mean levels of gray matter radioactivity were seen in the putamina and peaked at 7.5 min. A significant positive correlation was observed between K1 and V(T) (Spearman ρ = 0.398; P = 0.003). Between-subject coefficients of variation of V(T) ranged between 12% and 16%. Voxelwise RS-ESA yielded similar V(T)s and coefficients of variation. (18)F-GE-179 exhibits high and rapid brain extraction, with a relatively homogeneous distribution in gray matter and acceptable between-subject variability. Despite its rapid peripheral metabolism, quantification of (18)F-GE-179 VT is feasible both within regions of interest and at the voxel level. The specificity of (18)F-GE-179 binding, however, requires further characterization with in vivo studies using activation and disease models.

Mutagenesis of the C2 domain of protein kinase C-alpha. Differential roles of Ca2+ ligands and membrane binding residues.

PubMed

Medkova, M; Cho, W

1998-07-10

The C2 domains of conventional protein kinase C (PKC) have been implicated in their Ca2+-dependent membrane binding. The C2 domain of PKC-alpha contains several Ca2+ ligands that bind multiple Ca2+ ions and other putative membrane binding residues. To understand the roles of individual Ca2+ ligands and protein-bound Ca2+ ions in the membrane binding and activation of PKC-alpha, we mutated five putative Ca2+ ligands (D187N, D193N, D246N, D248N, and D254N) and measured the effects of mutations on vesicle binding, enzyme activity, and monolayer penetration of PKC-alpha. Altered properties of these mutants indicate that individual Ca2+ ions and their ligands have different roles in the membrane binding and activation of PKC-alpha. The binding of Ca2+ to Asp187, Asp193, and Asp246 of PKC-alpha is important for the initial binding of protein to membrane surfaces. On the other hand, the binding of another Ca2+ to Asp187, Asp246, Asp248, and Asp254 induces the conformational change of PKC-alpha, which in turn triggers its membrane penetration and activation. Among these Ca2+ ligands, Asp246 was shown to be most essential for both membrane binding and activation of PKC-alpha, presumably due to its coordination to multiple Ca2+ ions. Furthermore, to identify the residues in the C2 domain that are involved in membrane binding of PKC-alpha, we mutated four putative membrane binding residues (Trp245, Trp247, Arg249, and Arg252). Membrane binding and enzymatic properties of two double-site mutants (W245A/W247A and R249A/R252A) indicate that Arg249 and Arg252 are involved in electrostatic interactions of PKC-alpha with anionic membranes, whereas Trp245 and Trp247 participate in its penetration into membranes and resulting hydrophobic interactions. Taken together, these studies provide the first experimental evidence for the role of C2 domain of conventional PKC as a membrane docking unit as well as a module that triggers conformational changes to activate the protein.

Ni2+-binding RNA motifs with an asymmetric purine-rich internal loop and a G-A base pair.

PubMed Central

Hofmann, H P; Limmer, S; Hornung, V; Sprinzl, M

1997-01-01

RNA molecules with high affinity for immobilized Ni2+ were isolated from an RNA pool with 50 randomized positions by in vitro selection-amplification. The selected RNAs preferentially bind Ni2+ and Co2+ over other cations from first series transition metals. Conserved structure motifs, comprising about 15 nt, were identified that are likely to represent the Ni2+ binding sites. Two conserved motifs contain an asymmetric purine-rich internal loop and probably a mismatch G-A base pair. The structure of one of these motifs was studied with proton NMR spectroscopy and formation of the G-A pair at the junction of helix and internal loop was demonstrated. Using Ni2+ as a paramagnetic probe, a divalent metal ion binding site near this G-A base pair was identified. Ni2+ ions bound to this motif exert a specific stabilization effect. We propose that small asymmetric purine-rich loops that contain a G-A interaction may represent a divalent metal ion binding site in RNA. PMID:9409620

The study of zinc ions binding to casein.

PubMed

Pomastowski, P; Sprynskyy, M; Buszewski, B

2014-08-01

The presented research was focused on physicochemical study of casein properties and the kinetics of zinc ions binding to the protein. Moreover, a fast and simple method of casein extraction from cow's milk has been proposed. Casein isoforms, zeta potential (ζ) and particle size of the separated caseins were characterized with the use of capillary electrophoresis, zeta potential analysis and field flow fractionation (FFF) technique, respectively. The kinetics of the metal-binding process was investigated in batch adsorption experiments. Intraparticle diffusion model, first-order and zero-order kinetic models were applied to test the kinetic experimental data. Analysis of changes in infrared bands registered for casein before and after zinc binding was also performed. The obtained results showed that the kinetic process of zinc binding to casein is not homogeneous but is expressed with an initial rapid stage with about 70% of zinc ions immobilized by casein and with a much slower second step. Maximum amount of bound zinc in the experimental conditions was 30.04mgZn/g casein. Copyright © 2014 Elsevier B.V. All rights reserved.

A calix[4]arene strapped calix[4]pyrrole: an ion-pair receptor displaying three different cesium cation recognition modes.

PubMed

Kim, Sung Kuk; Sessler, Jonathan L; Gross, Dustin E; Lee, Chang-Hee; Kim, Jong Seung; Lynch, Vincent M; Delmau, Laetitia H; Hay, Benjamin P

2010-04-28

An ion-pair receptor, the calix[4]pyrrole-calix[4]arene pseudodimer 2, bearing a strong anion-recognition site but not a weak cation-recognition site, has been synthesized and characterized by standard spectroscopic means and via single-crystal X-ray diffraction analysis. In 10% CD(3)OD in CDCl(3) (v/v), this new receptor binds neither the Cs(+) cation nor the F(-) anion when exposed to these species in the presence of other counterions; however, it forms a stable 1:1 solvent-separated CsF complex when exposed to these two ions in concert with one another in this same solvent mixture. In contrast to what is seen in the case of a previously reported crown ether "strapped" calixarene-calixpyrrole ion-pair receptor 1 (J. Am. Chem. Soc. 2008, 130, 13162-13166), where Cs(+) cation recognition takes place within the crown, in 2.CsF cation recognition takes place within the receptor cavity itself, as inferred from both single-crystal X-ray diffraction analyses and (1)H NMR spectroscopic studies. This binding mode is supported by calculations carried out using the MMFF94 force field model. In 10% CD(3)OD in CDCl(3) (v/v), receptor 2 shows selectivity for CsF over the Cs(+) salts of Cl(-), Br(-), and NO(3)(-) but will bind these other cesium salts in the absence of fluoride, both in solution and in the solid state. In the case of CsCl, an unprecedented 2:2 complex is observed in the solid state that is characterized by two different ion-pair binding modes. One of these consists of a contact ion pair with the cesium cation and chloride anion both being bound within the central binding pocket and in direct contact with one another. The other mode involves a chloride anion bound to the pyrrole NH protons of a calixpyrrole subunit and a cesium cation sandwiched between two cone shaped calix[4]pyrroles originating from separate receptor units. In contrast to what is seen for CsF and CsCl, single-crystal X-ray structural analyses and (1)H NMR spectroscopic studies reveal that receptor 2 forms a 1:1 complex with CsNO(3), with the ions bound in the form of a contact ion pair. Thus, depending on the counteranion, receptor 2 is able to stabilize three different ion-pair binding modes with Cs(+), namely solvent-bridged, contact, and host-separated.

Binding of ATP by pertussis toxin and isolated toxin subunits

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

Hausman, S.Z.; Manclark, C.R.; Burns, D.L.

1990-07-03

The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of ({sup 3}H)ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of ({sup 3}H)ATP to pertussis toxin in a competitive manner;more » however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of ({sup 3}H)ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site.« less

Thermodynamic evidence of flexibility in H2O and CO2 absorption of transition metal ion exchanged zeolite LTA.

PubMed

Guo, Xin; Wu, Lili; Navrotsky, Alexandra

2018-02-07

Gas absorption calorimetry has been employed to probe the intercation of water and carbon dioxide with transition metal ion (TM = Mn 2+ , Fe 2+ , Co 2+ , Cu 2+ , and Zn 2+ ) exchanged zeolite A samples. There appears to be a two-phase region, indicative of a guest-induced flexibility transition, separating hydrated zeolite A and its dehydrated form, both of which have variable water content in the single phase region. The differential enthalpy of absorption as a function of water loading directly identifies different strengths of multiple interactions along with possible binding mechanisms of Zn-A and Mn-A exhibiting the highest water absorption with most exothermic initial enthalpies of -125.28 ± 4.82 and -115.30 ± 2.56 kJ mol -1 . Zn-A and Mn-A also show moderately good capture ability for CO 2 with zero-coverage negative enthalpies of -55.59 ± 2.48 and -44.07 ± 1.53 kJ mol -1 . The thermodynamic information derived from differential enthalpy, chemical potential and differential entropy elucidated the multistage interactive behavior of small guest molecules (H 2 O/CO 2 ) and ion-exchanged frameworks.

Impact of sodium caseinate concentration and location on magnesium release from multiple W/O/W emulsions.

PubMed

Bonnet, Marie; Cansell, Maud; Placin, Frédéric; Anton, Marc; Leal-Calderon, Fernando

2010-06-15

Water-in-oil-in-water (W/O/W) double emulsions were prepared and the rate of release of magnesium ions from the internal to the external aqueous phase was followed. Sodium caseinate was used not only as a hydrophilic surface-active species but also as a chelating agent able to bind magnesium ions. The release occurred without film rupturing (no coalescence). The kinetics of the release process depended on the location (in only one or in both aqueous compartments) and on the concentration of sodium caseinate. The rate of release increased with the concentration of sodium caseinate in the external phase and decreased when sodium caseinate was present in the inner droplets. The experiments were interpreted within the frame of a mean-field model based on diffusion, integrating the effect of ion binding. The data could be adequately fitted by considering a time-dependent permeation coefficient of the magnesium ions across the oil phase. Our results suggested that ion permeability was influenced by the state of the protein interfacial layers which itself depended on the extent of magnesium binding.

Chronic Beryllium Disease: Revealing the Role of Beryllium Ion and Small Peptides Binding to HLA-DP2

PubMed Central

Petukh, Marharyta; Wu, Bohua; Stefl, Shannon; Smith, Nick; Hyde-Volpe, David; Wang, Li; Alexov, Emil

2014-01-01

Chronic Beryllium (Be) Disease (CBD) is a granulomatous disorder that predominantly affects the lung. The CBD is caused by Be exposure of individuals carrying the HLA-DP2 protein of the major histocompatibility complex class II (MHCII). While the involvement of Be in the development of CBD is obvious and the binding site and the sequence of Be and peptide binding were recently experimentally revealed [1], the interplay between induced conformational changes and the changes of the peptide binding affinity in presence of Be were not investigated. Here we carry out in silico modeling and predict the Be binding to be within the acidic pocket (Glu26, Glu68 and Glu69) present on the HLA-DP2 protein in accordance with the experimental work [1]. In addition, the modeling indicates that the Be ion binds to the HLA-DP2 before the corresponding peptide is able to bind to it. Further analysis of the MD generated trajectories reveals that in the presence of the Be ion in the binding pocket of HLA-DP2, all the different types of peptides induce very similar conformational changes, but their binding affinities are quite different. Since these conformational changes are distinctly different from the changes caused by peptides normally found in the cell in the absence of Be, it can be speculated that CBD can be caused by any peptide in presence of Be ion. However, the affinities of peptides for Be loaded HLA-DP2 were found to depend of their amino acid composition and the peptides carrying acidic group at positions 4 and 7 are among the strongest binders. Thus, it is proposed that CBD is caused by the exposure of Be of an individual carrying the HLA-DP2*0201 allele and that the binding of Be to HLA-DP2 protein alters the conformational and ionization properties of HLA-DP2 such that the binding of a peptide triggers a wrong signaling cascade. PMID:25369028

Effects of heat treatment on oil-binding ability of rice flour.

PubMed

Tabara, Aya; Nakagawa, Mariko; Ushijima, Yuki; Matsunaga, Kotaro; Seguchi, Masaharu

2015-01-01

Heat-treated (120 °C for 120 min) rice flour showed high affinity to oil (oil-binding ability). This oil-binding ability could be observed by shaking the heat-treated rice flour (2.0 g), oil (4.0 mL), and water (20 mL) vigorously in a test tube, and the oil bound to the rice flour sank into the water. To examine the time-dependent levels of the oil-binding ability, rice flour was heat-treated at 120 °C for 10, 20, 40, 60, and 120 min, and the precipitated volume of oil/rice flour complex increased with an increase of the heating time. The oil-binding ability of the rice flour was not affected by the treatments with diethyl ether or boiled chloroform/methanol (2:1) solutions, which suggested no relationship to the oil in the rice flour, but was lost upon alkali (0.2% NaOH solution) or pepsin treatment, which suggested its relationship to the rice proteins.

Binding of Pseudomonas aeruginosa Apo-Bacterioferritin Associated Ferredoxin to Bacterioferritin B Promotes Heme Mediation of Electron Delivery and Mobilization of Core Mineral Iron†

PubMed Central

Weeratunga, Saroja K.; Gee, Casey E.; Lovell, Scott; Zeng, Yuhong; Woodin, Carrie L.; Rivera, Mario

2009-01-01

The bfrB gene from Pseudomonas aeruginosa was cloned and expressed in E. coli. The resultant protein (BfrB), which assembles into a 445.3 kDa complex0020from 24 identical subunits, binds 12 molecules of heme axially coordinated by two Met residues. BfrB, isolated with 5–10 iron atoms per protein molecule, was reconstituted with ferrous ions to prepare samples with a core mineral containing 600 ± 40 ferric ions per BfrB molecule and approximately one phosphate molecule per iron atom. In the presence of sodium dithionite or in the presence of P. aeruginosa ferredoxin NADP reductase (FPR) and NADPH the heme in BfrB remains oxidized and the core iron mineral is mobilized sluggishly. In stark contrast, addition of NADPH to a solution containing BfrB, FPR and the apo-form of P. aeruginosa bacterioferritin associated ferredoxin (apo-Bfd) results in rapid reduction of the heme in BfrB and in the efficient mobilization of the core iron mineral. Results from additional experimentation indicate that Bfd must bind to BfrB to promote heme mediation of electrons from the surface to the core to support the efficient mobilization of ferrous ions from BfrB. In this context, the thus far mysterious role of heme in bacterioferritins has been brought to the front by reconstituting BfrB with its physiological partner, apo-Bfd. These findings are discussed in the context of a model for the utilization of stored iron in which the significant upregulation of the bfd gene under low-iron conditions [Ochsner, U.A., Wilderman, P.J., Vasil, A.I., and Vasil, M.L. (2002) Mol. Microbiol. 45, 1277–1287] ensures sufficient concentrations of apo-Bfd to bind BfrB and unlock the iron stored in its core. Although these findings are in contrast to previous speculations suggesting redox mediation of electron transfer by holo-Bfd, the ability of apo-Bfd to promote iron mobilization is an economical strategy used by the cell because it obviates the need to further deplete cellular iron levels to assemble iron sulfur clusters in Bfd before the iron stored in BfrB can be mobilized and utilized. PMID:19575528

Antidepressant Binding Site in a Bacterial Homologue of Neurotransmitter Transporters

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

Singh,S.; Yamashita, A.; Gouaux, E.

Sodium-coupled transporters are ubiquitous pumps that harness pre-existing sodium gradients to catalyse the thermodynamically unfavourable uptake of essential nutrients, neurotransmitters and inorganic ions across the lipid bilayer. Dysfunction of these integral membrane proteins has been implicated in glucose/galactose malabsorption, congenital hypothyroidism, Bartter's syndrome, epilepsy, depression, autism and obsessive-compulsive disorder. Sodium-coupled transporters are blocked by a number of therapeutically important compounds, including diuretics, anticonvulsants and antidepressants, many of which have also become indispensable tools in biochemical experiments designed to probe antagonist binding sites and to elucidate transport mechanisms. Steady-state kinetic data have revealed that both competitive and noncompetitive modes of inhibitionmore » exist. Antagonist dissociation experiments on the serotonin transporter (SERT) have also unveiled the existence of a low-affinity allosteric site that slows the dissociation of inhibitors from a separate high-affinity site. Despite these strides, atomic-level insights into inhibitor action have remained elusive. Here we screen a panel of molecules for their ability to inhibit LeuT, a prokaryotic homologue of mammalian neurotransmitter sodium symporters, and show that the tricyclic antidepressant (TCA) clomipramine noncompetitively inhibits substrate uptake. Cocrystal structures show that clomipramine, along with two other TCAs, binds in an extracellular-facing vestibule about 11 {angstrom} above the substrate and two sodium ions, apparently stabilizing the extracellular gate in a closed conformation. Off-rate assays establish that clomipramine reduces the rate at which leucine dissociates from LeuT and reinforce our contention that this TCA inhibits LeuT by slowing substrate release. Our results represent a molecular view into noncompetitive inhibition of a sodium-coupled transporter and define principles for the rational design of new inhibitors.« less

Role of urea on recombinant Apo A-I stability and its utilization in anion exchange chromatography.

PubMed

Angarita, Monica; Arosio, Paolo; Müller-Späth, Thomas; Baur, Daniel; Falkenstein, Roberto; Kuhne, Wolfgang; Morbidelli, Massimo

2014-08-08

Apolipoprotein A-I (Apo A-I) is an important lipid-binding protein involved in the transport and metabolism of cholesterol. High protein purity, in particular with respect to endotoxins is required for therapeutic applications. The use of urea during the purification process of recombinant Apo A-I produced in Escherichia coli has been suggested so as to provide high endotoxin clearance. In this work, we show that urea can be used as a sole modifier during the ion exchange chromatographic purification of Apo A-I and we investigate the molecular mechanism of elution by correlating the effect of urea on self-association, conformation and adsorption equilibrium properties of a modified model Apo A-I. In the absence of urea the protein was found to be present as a population of oligomers represented mainly by trimers, hexamers and nonamers. The addition of urea induced oligomer dissociation and protein structure unfolding. We correlated the changes in protein association and conformation with variations of the adsorption equilibrium of the protein on a strong anion exchanger. It was confirmed that the adsorption isotherms, described by a Langmuir model, were dependent on both protein and urea concentrations. Monomers, observed at low urea concentration (0.5M), were characterized by larger binding affinity and adsorption capacity compared to both protein oligomers (0M) and unfolded monomers (2-8M). The reduction of both the binding strength and maximum adsorption capacity at urea concentrations larger than 0.5M explains the ability of urea of inducing elution of the protein from the ion exchange resin. The dissociation of the protein complexes occurring during the elution could likely be the origin of the effective clearance of endotoxins originally trapped inside the oligomers. Copyright © 2014 Elsevier B.V. All rights reserved.

The Hexahistidine Motif of Host-Defense Protein Human Calprotectin Contributes to Zinc Withholding and Its Functional Versatility.

PubMed

Nakashige, Toshiki G; Stephan, Jules R; Cunden, Lisa S; Brophy, Megan Brunjes; Wommack, Andrew J; Keegan, Brenna C; Shearer, Jason M; Nolan, Elizabeth M

2016-09-21

Human calprotectin (CP, S100A8/S100A9 oligomer, MRP-8/MRP-14 oligomer) is an abundant host-defense protein that is involved in the metal-withholding innate immune response. CP coordinates a variety of divalent first-row transition metal ions, which is implicated in its antimicrobial function, and its ability to sequester nutrient Zn(II) ions from microbial pathogens has been recognized for over two decades. CP has two distinct transition-metal-binding sites formed at the S100A8/S100A9 dimer interface, including a histidine-rich site composed of S100A8 residues His17 and His27 and S100A9 residues His91 and His95. In this study, we report that CP binds Zn(II) at this site using a hexahistidine motif, completed by His103 and His105 of the S100A9 C-terminal tail and previously identified as the high-affinity Mn(II) and Fe(II) coordination site. Zn(II) binding at this unique site shields the S100A9 C-terminal tail from proteolytic degradation by proteinase K. X-ray absorption spectroscopy and Zn(II) competition titrations support the formation of a Zn(II)-His6 motif. Microbial growth studies indicate that the hexahistidine motif is important for preventing microbial Zn(II) acquisition from CP by the probiotic Lactobacillus plantarum and the opportunistic human pathogen Candida albicans. The Zn(II)-His6 site of CP expands the known biological coordination chemistry of Zn(II) and provides new insight into how the human innate immune system starves microbes of essential metal nutrients.

Exopolysaccharides regulate calcium flow in cariogenic biofilms

PubMed Central

Varenganayil, Muth M.; Decho, Alan W.

2017-01-01

Caries-associated biofilms induce loss of calcium from tooth surfaces in the presence of dietary carbohydrates. Exopolysaccharides (EPS) provide a matrix scaffold and an abundance of primary binding sites within biofilms. The role of EPS in binding calcium in cariogenic biofilms is only partially understood. Thus, the aim of the present study is to investigate the relationship between the calcium dissolution rates and calcium tolerance of caries-associated bacteria and yeast as well as to examine the properties of EPS to quantify its binding affinity for dissolved calcium. Calcium dissolution was measured by dissolution zones on Pikovskaya’s agar. Calcium tolerance was assessed by isothermal microcalorimetry (IMC) by adding CaCl2 to the bacterial cultures. Acid-base titration and Fourier transform infrared (FTIR) spectroscopy were used to identify possible functional groups responsible for calcium binding, which was assessed by isothermal titration calorimetry (ITC). Lactobacillus spp. and mutans streptococci demonstrated calcium dissolution in the presence of different carbohydrates. All strains that demonstrated high dissolution rates also revealed higher rates of calcium tolerance by IMC. In addition, acidic functional groups were predominantly identified as possible binding sites for calcium ions by acid-base titration and FTIR. Finally, ITC revealed EPS to have a higher binding affinity for calcium compared, for example, to lactic acid. In conclusion, this study illustrates the role of EPS in terms of the calcium tolerance of cariogenic microbiota by determining the ability of EPS to control free calcium concentrations within the biofilms as a self-regulating mode of action in the pathogenesis of dental caries. PMID:29023506

Stoichiometry for activation of neuronal α7 nicotinic receptors

PubMed Central

Andersen, Natalia; Corradi, Jeremías; Sine, Steven M.; Bouzat, Cecilia

2013-01-01

Neuronal α7 nicotinic receptors elicit rapid cation influx in response to acetylcholine (ACh) or its hydrolysis product choline. They contribute to cognition, synaptic plasticity, and neuroprotection and have been implicated in neurodegenerative and neuropsychiatric disorders. α7, however, often localizes distal to sites of nerve-released ACh and binds ACh with low affinity, and thus elicits its biological response with low agonist occupancy. To assess the function of α7 when ACh occupies fewer than five of its identical binding sites, we measured the open-channel lifetime of individual receptors in which four of the five ACh binding sites were disabled. To improve the time resolution of the inherently brief α7 channel openings, background mutations or a potentiator was used to increase open duration. We find that, in receptors with only one intact binding site, the open-channel lifetime is indistinguishable from receptors with five intact binding sites, counter to expectations from prototypical neurotransmitter-gated ion channels where the open-channel lifetime increases with the number of binding sites occupied by agonist. Replacing the membrane-embedded domain of α7 by that of the related 5-HT3A receptor increases the number of sites that need to be occupied to achieve the maximal open-channel lifetime, thus revealing a unique interdependence between the detector and actuator domains of these receptors. The distinctive ability of a single occupancy to elicit a full biological response adapts α7 to volume transmission, a prevalent mechanism of ACh-mediated signaling in the nervous system and nonneuronal cells. PMID:24297903

Coordination ability determined transition metal ions substitution of Tb in Tb-Asp fluorescent nanocrystals and a facile ions-detection approach.

PubMed

Duan, Jiazhi; Ma, Baojin; Liu, Feng; Zhang, Shan; Wang, Shicai; Kong, Ying; Du, Min; Han, Lin; Wang, Jianjun; Sang, Yuanhua; Liu, Hong

2018-04-26

Although the synthesis and fluorescent properties of lanthanide-amino acid complex nanostructures have been investigated extensively, limited studies have been reported on metal ions' substitution ability for the lanthanide ions in the complex and their effect on the fluorescent property. In this study, taking biocompatible Tb-aspartic acid (Tb-Asp) complex nanocrystals as a model, the substitution mechanism of metal ions, particularly transition metals, for Tb ions in Tb-Asp nanocrystals and the change in the fluorescent property of the Tb-Asp nanocrystals after substitution were systematically investigated. The experimental results illustrated that metal ions with higher electronegativity, higher valence, and smaller radius possess stronger ability for Tb ions' substitution in Tb-Asp nanocrystals. Based on the effect of substituting ions' concentration on the fluorescent property of Tb-Asp, a facile method for copper ions detection with high sensitivity was proposed by measuring the fluorescent intensity of Tb-Asp nanocrystals' suspensions containing different concentrations of copper ions. The good biocompatibility, great convenience of synthesis and sensitive detection ability make Tb-Asp nanocrystals a very low cost and effective material for metal ions detection, which also opens a new door for practical applications of metal-Asp coordinated nanocrystals.

Conformational changes in the M2 muscarinic receptor induced by membrane voltage and agonist binding

PubMed Central

Navarro-Polanco, Ricardo A; Galindo, Eloy G Moreno; Ferrer-Villada, Tania; Arias, Marcelo; Rigby, J Ryan; Sánchez-Chapula, José A; Tristani-Firouzi, Martin

2011-01-01

Abstract The ability to sense transmembrane voltage is a central feature of many membrane proteins, most notably voltage-gated ion channels. Gating current measurements provide valuable information on protein conformational changes induced by voltage. The recent observation that muscarinic G-protein-coupled receptors (GPCRs) generate gating currents confirms their intrinsic capacity to sense the membrane electrical field. Here, we studied the effect of voltage on agonist activation of M2 muscarinic receptors (M2R) in atrial myocytes and how agonist binding alters M2R gating currents. Membrane depolarization decreased the potency of acetylcholine (ACh), but increased the potency and efficacy of pilocarpine (Pilo), as measured by ACh-activated K+ current, IKACh. Voltage-induced conformational changes in M2R were modified in a ligand-selective manner: ACh reduced gating charge displacement while Pilo increased the amount of charge displaced. Thus, these ligands manifest opposite voltage-dependent IKACh modulation and exert opposite effects on M2R gating charge displacement. Finally, mutations in the putative ligand binding site perturbed the movement of the M2R voltage sensor. Our data suggest that changes in voltage induce conformational changes in the ligand binding site that alter the agonist–receptor interaction in a ligand-dependent manner. Voltage-dependent GPCR modulation has important implications for cellular signalling in excitable tissues. Gating current measurement allows for the tracking of subtle conformational changes in the receptor that accompany agonist binding and changes in membrane voltage. PMID:21282291

The crystal structure of the Rv0301-Rv0300 VapBC-3 toxin-antitoxin complex from M. tuberculosis reveals a Mg 2+ ion in the active site and a putative RNA-binding site

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

Min, Andrew B; Miallau, Linda; Sawaya, Michael R

VapBC pairs account for 45 out of 88 identified toxin-antitoxin (TA) pairs in the Mycobacterium tuberculosis (Mtb) H37Rv genome. A working model suggests that under times of stress, antitoxin molecules are degraded, releasing the toxins to slow the metabolism of the cell, which in the case of VapC toxins is via their RNase activity. Otherwise the TA pairs remain bound to their promoters, autoinhibiting transcription. The crystal structure of Rv0301-Rv0300, an Mtb VapBC TA complex determined at 1.49 Å resolution, suggests a mechanism for these three functions: RNase activity, its inhibition by antitoxin, and its ability to bind promoter DNA.more » The Rv0301 toxin consists of a core of five parallel beta strands flanked by alpha helices. Three proximal aspartates coordinate a Mg2+ ion forming the putative RNase active site. The Rv0300 antitoxin monomer is extended in structure, consisting of an N-terminal beta strand followed by four helices. The last two helices wrap around the toxin and terminate near the putative RNase active site, but with different conformations. In one conformation, the C-terminal arginine interferes with Mg2+ ion coordination, suggesting a mechanism by which the antitoxin can inhibit toxin activity. At the N-terminus of the antitoxin, two pairs of Ribbon-Helix-Helix (RHH) motifs are related by crystallographic twofold symmetry. The resulting hetero-octameric complex is similar to the FitAB system, but the two RHH motifs are about 30 Å closer together in the Rv0301-Rv0300 complex, suggesting either a different span of the DNA recognition sequence or a conformational change.« less

Real-Time Study of the Interaction between G-Rich DNA Oligonucleotides and Lead Ion on DNA Tetrahedron-Functionalized Sensing Platform by Dual Polarization Interferometry.

PubMed

Wang, Shuang; Lu, Shasha; Zhao, Jiahui; Huang, Jianshe; Yang, Xiurong

2017-11-29

G-quadruplex plays roles in numerous physiological and pathological processes of organisms. Due to the unique properties of G-quadruplex (e.g., forming G4/hemin complexes with catalytic activity and electron acceptability, binding with metal ions, proteins, fluorescent ligands, and so on), it has been widely applied in biosensing. But the formation process of G-quadruplex is not yet fully understood. Here, a DNA tetrahedron platform with higher reproducibility, regenerative ability, and time-saving building process was coupled with dual polarization interferometry technique for the real-time and label-free investigation of the specific interaction process of guanine-rich singled-stranded DNA (G-rich ssDNA) and Pb 2+ . The oriented immobilization of probes greatly decreased the spatial hindrance effect and improved the accessibility of the probes to the Pb 2+ ions. Through real-time monitoring of the whole formation process of the G-quadruplex, we speculated that the probes on the tetrahedron platform initially stood on the sensing surface with a random coil conformation, then the G-rich ssDNA preliminarily formed unstable G-quartets by H-bonding and cation binding, subsequently forming a completely folded and stable quadruplex structure through relatively slow strand rearrangements. On the basis of these studies, we also developed a novel sensing platform for the specific and sensitive determination of Pb 2+ and its chelating agent ethylenediaminetetraacetic acid. This study not only provides a proof-of-concept for conformational dynamics of G-quadruplex-related drugs and pathogenes, but also enriches the biosensor tools by combining nanomaterial with interfaces technique.

Structure of the Dominant Negative S17N Mutant of Ras

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

Nassar, N.; Singh, K; Garcia-Diaz, M

2010-01-01

The use of the dominant negative mutant of Ras has been crucial in elucidating the cellular signaling of Ras in response to the activation of various membrane-bound receptors. Although several point mutants of Ras exhibit a dominant negative effect, the asparagine to serine mutation at position 17 (S17N) remains the most popular and the most effective at inhibiting the activation of endogenous Ras. It is now widely accepted that the dominant negative effect is due to the ability of the mutant to sequester upstream activators and its inability to activate downstream effectors. Here, we present the crystal structure of RasS17Nmore » in the GDP-bound form. In the three molecules that populate the asymmetric unit, the Mg{sup 2+} ion that normally coordinates the {beta}-phosphate is absent because of steric hindrance from the Asn17 side chain. Instead, a Ca{sup 2+} ion is coordinating the {alpha}-phosphate. Also absent from one molecule is electron density for Phe28, a conserved residue that normally stabilizes the nucleotide's guanine base. Except for Phe28, the nucleotide makes conserved interactions with Ras. Combined, the inability of Phe28 to stabilize the guanine base and the absence of a Mg{sup 2+} ion to neutralize the negative charges on the phosphates explain the weaker affinity of GDP for Ras. Our data suggest that the absence of the Mg{sup 2+} should also dramatically affect GTP binding to Ras and the proper positioning of Thr35 necessary for the activation of switch 1 and the binding to downstream effectors, a prerequisite for the triggering of signaling pathways.« less

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.

Mechanistic Insight from Calorimetric Measurements of the Assembly of the Binuclear Metal Active Site of Glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes.

PubMed

Pedroso, Marcelo M; Ely, Fernanda; Carpenter, Margaret C; Mitić, Nataša; Gahan, Lawrence R; Ollis, David L; Wilcox, Dean E; Schenk, Gerhard

2017-07-05

Glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes is a binuclear metallohydrolase with a high affinity for metal ions at its α site but a lower affinity at its β site in the absence of a substrate. Isothermal titration calorimetry (ITC) has been used to quantify the Co(II) and Mn(II) binding affinities and thermodynamics of the two sites in wild-type GpdQ and two mutants, both in the absence and in the presence of phosphate. Metal ions bind to the six-coordinate α site in an entropically driven process with loss of a proton, while binding at the β site is not detected by ITC. Phosphate enhances the metal affinity of the α site by increasing the binding entropy and the metal affinity of the β site by enthalpic (Co) or entropic (Mn) contributions, but no additional loss of protons. Mutations of first- and second-coordination sphere residues at the β site increase the metal affinity of both sites by enhancing the binding enthalpy. In particular, loss of the hydrogen bond from second-sphere Ser127 to the metal-coordinating Asn80 has a significant effect on the metal binding thermodynamics that result in a resting binuclear active site with high catalytic activity. While structural and spectroscopic data with excess metal ions have indicated a bridging hydroxide in the binuclear GpdQ site, analysis of ITC data here reveals the loss of a single proton in the assembly of this site, indicating that the metal-bound hydroxide nucleophile is formed in the resting inactive mononuclear form, which becomes catalytically competent upon binding the second metal ion.

Influence of water chemistry and natural organic matter on active and passive uptake of inorganic mercury by gills of rainbow trout (Oncorhynchus mykiss).

PubMed

Klinck, Joel; Dunbar, Michael; Brown, Stephanie; Nichols, Joel; Winter, Anna; Hughes, Christopher; Playle, Richard C

2005-03-25

To distinguish physiologically regulated uptake from passive uptake of inorganic Hg in fish, rainbow trout (Oncorhynchus mykiss) were exposed to inorganic Hg (0.5, 1, or 2 microM total Hg) in ion-poor water with various treatments. Addition of ions to the water (mM concentrations of Ca, K, Cl) did not consistently alter Hg accumulation by trout gills, although there was a trend to higher Hg accumulation at higher ion concentrations. The apical Ca channel blockers Verapamil and lanthanum also did not consistently affect Hg accumulation by trout gills. Pre-treatment of trout with the Na channel blocker Phenamil decreased Hg uptake by about half. These results suggest a combination of physiologically regulated and passive uptake of Hg by trout gills. Strong complexing agents of Hg (EDTA, NTA, ethylenediamine, cysteine) decreased Hg-binding by trout gills in a dose-dependent manner. From these data, a conditional equilibrium binding constant for Hg to the gills was estimated as logK(Hg-gill) = 18.0, representing very strong binding of Hg to the gills. This value is a first step in creating a biotic ligand model (BLM) for inorganic Hg and fish. Natural organic matter (2-10 mg C/L) also decreased Hg-binding by trout gills, although mM concentrations of Na, K, and Cl interfered with this effect. At low concentrations of these ions, natural organic matter samples isolated from various sources bound Hg to similar degrees, as judged by Hg accumulation by trout gills. A conditional binding constant to natural organic matter (NOM) was estimated as logK(Hg-NOM) = 18.0 with about 0.5 micromol binding sites per mg C, representing strong binding of Hg to NOM.

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

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

Defects in the calcium-binding region drastically affect the cadherin-like domains of RET tyrosine kinase.

PubMed

Gao, Chunxia; Grøtli, Morten; Eriksson, Leif A

2016-03-28

Mutations in the rearranged during transfection (RET) tyrosine kinase gene leading to gain or loss of function have been associated with the development of several human cancers and Hirschsprung's disease (HSCR). However, to what extent these mutations affect individual bio-molecular functions remains unclear. In this article, the functionally significant mutations in the RET CLD1-4 calcium-binding site which lead to HSCR, and depletion of calcium ions in the RET CLD1-4 calcium binding site, were investigated by molecular dynamics simulations--to understand the mechanistic action of the mutations or loss of calcium ions in altering the protein kinase structure, dynamics, and stability. The mutations or loss of calcium ions change the local conformation and change the free energy landscape. Specifically, the mutations and loss of calcium ions decrease the radius of gyration of the whole structure, leading to improper protein folding and GFL-GFRα contact site reduction. Furthermore, based on the most populated conformation in the wildtype MD simulations, a pharmacophore was generated by fragment docking to identify key features of the possible inhibitors targeting the calcium binding site. Overall, the findings may provide useful structural insights into the molecular mechanism underlying RET calcium-binding site mutations and assist in development of novel drugs targeting the extracellular ligand contact site of wildtype RET.

Crucial role of dynamic linker histone binding and divalent ions for DNA accessibility and gene regulation revealed by mesoscale modeling of oligonucleosomes

PubMed Central

Collepardo-Guevara, Rosana; Schlick, Tamar

2012-01-01

Monte Carlo simulations of a mesoscale model of oligonucleosomes are analyzed to examine the role of dynamic-linker histone (LH) binding/unbinding in high monovalent salt with divalent ions, and to further interpret noted chromatin fiber softening by dynamic LH in monovalent salt conditions. We find that divalent ions produce a fiber stiffening effect that competes with, but does not overshadow, the dramatic softening triggered by dynamic-LH behavior. Indeed, we find that in typical in vivo conditions, dynamic-LH binding/unbinding reduces fiber stiffening dramatically (by a factor of almost 5, as measured by the elasticity modulus) compared with rigidly fixed LH, and also the force needed to initiate chromatin unfolding, making it consistent with those of molecular motors. Our data also show that, during unfolding, divalent ions together with LHs induce linker-DNA bending and DNA–DNA repulsion screening, which guarantee formation of heteromorphic superbeads-on-a-string structures that combine regions of loose and compact fiber independently of the characteristics of the LH–core bond. These structures might be important for gene regulation as they expose regions of the DNA selectively. Dynamic control of LH binding/unbinding, either globally or locally, in the presence of divalent ions, might constitute a mechanism for regulation of gene expression. PMID:22790986

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

Effect of Ion Binding in Palmitoyl-Oleoyl Phosphatidylserine Monolayers

NASA Astrophysics Data System (ADS)

Eckler, Matthew; Matysiak, Silvina

2013-03-01

Molecular dynamics simulations of palmitoyl-oleoyl phosphatidylserine (POPS) monolayers at the air-water interface were performed with different ionic strengths with the aim of determining the specific organization and dynamics of counterion binding events. Na + ions penetrated the monolayers into both the ester carbonyl and carboxylate regions of the phospholipids. The binding events increase with the addition of salt. Differences in lipid order parameter, headgroup orientation, and prevalence of inter- and intramolecular hydrogen bonding events between the amine group of the lipid and oxygen groups are observed depending on whether the Na + is binding near the carboxylate or ester region of the lipid. The observed changes are explained in terms of the salting-out effect.

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.

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(+).

Highly sensitive and selective detection of Al(III) ions in aqueous buffered solution with fluorescent peptide-based sensor.

PubMed

In, Byunggyu; Hwang, Gi Won; Lee, Keun-Hyeung

2016-09-15

A fluorescent sensor based on a tripeptide (SerGluGlu) with a dansyl fluorophore detected selectively Al(III) among 16 metal ions in aqueous buffered solutions without any organic cosolvent. The peptide-based sensor showed a highly sensitive turn on response to aluminium ion with high binding affinity (1.84×10(4)M(-1)) in aqueous buffered solutions. The detection limit (230nM, 5.98ppb) of the peptide-based sensor was much lower than the maximum allowable level (7.41μM) of aluminium ions in drinking water demanded by EPA. The binding mode of the peptide sensor with aluminium ions was characterized using ESI mass spectrometry, NMR titration, and pH titration experiments. Copyright © 2016 Elsevier Ltd. All rights reserved.

Anions mediate ligand binding in Adineta vaga glutamate receptor ion channels

PubMed Central

Lomash, Suvendu; Chittori, Sagar; Brown, Patrick; Mayer, Mark L.

2014-01-01

SUMMARY AvGluR1, a glutamate receptor ion channel from the primitive eukaryote Adineta vaga, is activated by alanine, cysteine, methionine and phenylalanine which produce lectin-sensitive desensitizing responses like those to glutamate, aspartate and serine. AvGluR1 LBD crystal structures reveal a novel scheme for binding dissimilar ligands that may be utilized by distantly related odorant/chemosensory receptors. Arginine residues in domain 2 coordinate the γ-carboxyl group of glutamate, while in the alanine, methionine and serine complexes a chloride ion acts as a surrogate ligand, replacing the γ-carboxyl group. Removal of Cl− lowers affinity for these ligands, but not for glutamate, aspartate or for phenylalanine which occludes the anion binding site and binds with low affinity. AvGluR1 LBD crystal structures and sedimentation analysis also provide insights into the evolutionary link between prokaryotic and eukaryotic iGluRs and reveal features unique to both classes, emphasizing the need for additional structure based studies on iGluR-ligand interactions. PMID:23434404

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

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

ELECTRON STAINS

PubMed Central

Zobel, C. Richard; Beer, Michael

1961-01-01

Chemical studies have been carried out on the interaction of DNA with uranyl salts. The effect of variations in pH, salt concentration, and structural integrity of the DNA on the stoichiometry of the salt-substrate complex have been investigated. At pH 3.5 DNA interacts with uranyl ions in low concentration yielding a substrate metal ion complex with a UO2++/P mole ratio of about ½ and having a large association constant. At low pH's (about 2.3) the mole ratio decreases to about ⅓. Destruction of the structural integrity of the DNA by heating in HCHO solutions leads to a similar drop in the amount of metal ion bound. Raising the pH above 3.5 leads to an apparent increase in binding as does increasing the concentration of the salt solution. This additional binding has a lower association constant. Under similar conditions DNA binds about seven times more uranyl ion than bovine serum albumin, indicating useful selectivity in staining for electron microscopy. PMID:13788706

Application of Sargassum biomass to remove heavy metal ions from synthetic multi-metal solutions and urban storm water runoff.

PubMed

Vijayaraghavan, K; Teo, Ting Ting; Balasubramanian, R; Joshi, Umid Man

2009-05-30

The ability of Sargassum sp. to biosorb four metal ions, namely lead, copper, zinc, and manganese from a synthetic multi-solute system and real storm water runoff has been investigated for the first time. Experiments on synthetic multi-solute systems revealed that Sargassum performed well in the biosorption of all four metal ions, with preference towards Pb, followed by Cu, Zn, and Mn. The solution pH strongly affected the metal biosorption, with pH 6 being identified as the optimal condition for achieving maximum biosorption. Experiments at different biosorbent dosages revealed that good biosorption capacity as well as high metal removal efficiency was observed at 3g/L. The biosorption kinetics was found to be fast with equilibrium being attained within 50 min. According to the Langmuir isotherm model, Sargassum exhibited maximum uptakes of 214, 67.5, 24.2 and 20.2mg/g for lead, copper, zinc, and manganese, respectively in single-solute systems. In multi-metal systems, strong competition between four metal ions in terms of occupancy binding sites was observed, and Sargassum showed preference in the order of Pb>Cu>Zn>Mn. The application of Sargassum to remove four heavy metal ions in real storm water runoff revealed that the biomass was capable of removing the heavy metal ions. However, the biosorption performance was slightly lower compared to that of synthetic metal solutions. Several factors were responsible for this difference, and the most important factor is the presence of other contaminants such as anions, organics, and other trace metals in the runoff.

Formation of crystalline nanoparticles by iron binding to pentapeptide (Asp-His-Thr-Lys-Glu) from egg white hydrolysates.

PubMed

Sun, Na; Cui, Pengbo; Li, Dongmei; Jin, Ziqi; Zhang, Shuyu; Lin, Songyi

2017-09-20

A novel peptide from egg white, Asp-His-Thr-Lys-Glu (DHTKE), contains specific amino acids associated with iron binding. The present study aims to better understand the molecular basis of interactions between the DHTKE peptide and iron ions. The ultraviolet-visible and fluorescence spectra indicate an interaction between the DHTKE peptide and iron ions, which leads to the formation of a DHTKE-iron complex. Notably, Asp, Glu, His, and Lys in the DHTKE peptide play crucial roles in the formation of the DHTKE-iron complex, and the iron-binding site of the DHTKE peptide corresponds primarily to the amide and carboxyl groups. The DHTKE peptide can bind iron ions in a 1 : 2 ratio with a binding constant of 1.312 × 10 5 M -1 . Moreover, the DHTKE-iron complex belongs to thermodynamically stable nanoparticles that are present in the crystalline structure, which might be attributed to peptide folding induced by iron binding. Meanwhile, the DHTKE-iron complex exhibits a relatively high iron-releasing percentage and exerts excellent solubility in the human gastrointestinal tract in vitro. This suggests a potential application of peptides containing Asp, Glu, His, or Lys residues as potential iron supplements.

Using affinity capillary electrophoresis and computational models for binding studies of heparinoids with p-selectin and other proteins.

PubMed

Mozafari, Mona; Balasupramaniam, Shantheya; Preu, Lutz; El Deeb, Sami; Reiter, Christian G; Wätzig, Hermann

2017-06-01

A fast and precise affinity capillary electrophoresis (ACE) method has been developed and applied for the investigation of the binding interactions between P-selectin and heparinoids as potential P-selectin inhibitors in the presence and absence of calcium ions. Furthermore, model proteins and vitronectin were used to appraise the binding behavior of P-selectin. The normalized mobility ratios (∆R/R f ), which provided information about the binding strength and the overall charge of the protein-ligand complex, were used to evaluate the binding affinities. It was found that P-selectin interacts more strongly with heparinoids in the presence of calcium ions. P-selectin was affected by heparinoids at the concentration of 3 mg/L. In addition, the results of the ACE experiments showed that among other investigated proteins, albumins and vitronectin exhibited strong interactions with heparinoids. Especially with P-selectin and vitronectin, the interaction may additionally induce conformational changes. Subsequently, computational models were applied to interpret the ACE experiments. Docking experiments explained that the binding of heparinoids on P-selectin is promoted by calcium ions. These docking models proved to be particularly well suited to investigate the interaction of charged compounds, and are therefore complementary to ACE experiments. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Computational membrane biophysics: From ion channel interactions with drugs to cellular function.

PubMed

Miranda, Williams E; Ngo, Van A; Perissinotti, Laura L; Noskov, Sergei Yu

2017-11-01

The rapid development of experimental and computational techniques has changed fundamentally our understanding of cellular-membrane transport. The advent of powerful computers and refined force-fields for proteins, ions, and lipids has expanded the applicability of Molecular Dynamics (MD) simulations. A myriad of cellular responses is modulated through the binding of endogenous and exogenous ligands (e.g. neurotransmitters and drugs, respectively) to ion channels. Deciphering the thermodynamics and kinetics of the ligand binding processes to these membrane proteins is at the heart of modern drug development. The ever-increasing computational power has already provided insightful data on the thermodynamics and kinetics of drug-target interactions, free energies of solvation, and partitioning into lipid bilayers for drugs. This review aims to provide a brief summary about modeling approaches to map out crucial binding pathways with intermediate conformations and free-energy surfaces for drug-ion channel binding mechanisms that are responsible for multiple effects on cellular functions. We will discuss post-processing analysis of simulation-generated data, which are then transformed to kinetic models to better understand the molecular underpinning of the experimental observables under the influence of drugs or mutations in ion channels. This review highlights crucial mathematical frameworks and perspectives on bridging different well-established computational techniques to connect the dynamics and timescales from all-atom MD and free energy simulations of ion channels to the physiology of action potentials in cellular models. This article is part of a Special Issue entitled: Biophysics in Canada, edited by Lewis Kay, John Baenziger, Albert Berghuis and Peter Tieleman. Copyright © 2017 Elsevier B.V. All rights reserved.

Selective detection of Mg2+ ions via enhanced fluorescence emission using Au–DNA nanocomposites

PubMed Central

Basu, Tanushree; Rana, Khyati; Das, Niranjan

2017-01-01

The biophysical properties of DNA-modified Au nanoparticles (AuNPs) have attracted a great deal of research interest for various applications in biosensing. AuNPs have strong binding capability to the phosphate and sugar groups in DNA, rendering unique physicochemical properties for detection of metal ions. The formation of Au–DNA nanocomposites is evident from the observed changes in the optical absorption, plasmon band, zeta potential, DLS particle size distribution, as well as TEM and AFM surface morphology analysis. Circular dichroism studies also revealed that DNA-functionalized AuNP binding caused a conformational change in the DNA structure. Due to the size and shape dependent plasmonic interactions of AuNPs (33–78 nm) with DNA, the resultant Au–DNA nanocomposites (NCs) exhibit superior fluorescence emission due to chemical binding with Ca2+, Fe2+ and Mg2+ ions. A significant increase in fluorescence emission (λex = 260 nm) of Au–DNA NCs was observed after selectively binding with Mg2+ ions (20–800 ppm) in an aqueous solution where a minimum of 100 ppm Mg2+ ions was detected based on the linearity of concentration versus fluorescence intensity curve (λem = 400 nm). The effectiveness of Au–DNA nanocomposites was further verified by comparing the known concentration (50–120 ppm) of Mg2+ ions in synthetic tap water and a real life sample of Gelusil (300–360 ppm Mg2+), a widely used antacid medicine. Therefore, this method could be a sensitive tool for the estimation of water hardness after careful preparation of a suitably designed Au–DNA nanostructure. PMID:28487819

Ion-binding properties of a K+ channel selectivity filter in different conformations.

PubMed

Liu, Shian; Focke, Paul J; Matulef, Kimberly; Bian, Xuelin; Moënne-Loccoz, Pierre; Valiyaveetil, Francis I; Lockless, Steve W

2015-12-08

K(+) channels are membrane proteins that selectively conduct K(+) ions across lipid bilayers. Many voltage-gated K(+) (KV) channels contain two gates, one at the bundle crossing on the intracellular side of the membrane and another in the selectivity filter. The gate at the bundle crossing is responsible for channel opening in response to a voltage stimulus, whereas the gate at the selectivity filter is responsible for C-type inactivation. Together, these regions determine when the channel conducts ions. The K(+) channel from Streptomyces lividians (KcsA) undergoes an inactivation process that is functionally similar to KV channels, which has led to its use as a practical system to study inactivation. Crystal structures of KcsA channels with an open intracellular gate revealed a selectivity filter in a constricted conformation similar to the structure observed in closed KcsA containing only Na(+) or low [K(+)]. However, recent work using a semisynthetic channel that is unable to adopt a constricted filter but inactivates like WT channels challenges this idea. In this study, we measured the equilibrium ion-binding properties of channels with conductive, inactivated, and constricted filters using isothermal titration calorimetry (ITC). EPR spectroscopy was used to determine the state of the intracellular gate of the channel, which we found can depend on the presence or absence of a lipid bilayer. Overall, we discovered that K(+) ion binding to channels with an inactivated or conductive selectivity filter is different from K(+) ion binding to channels with a constricted filter, suggesting that the structures of these channels are different.

Ratiometric and turn-on monitoring for heavy and transition metal ions in aqueous solution with a fluorescent peptide sensor.

PubMed

Joshi, Bishnu Prasad; Park, Junwon; Lee, Wan In; Lee, Keun-Hyeung

2009-05-15

A novel fluorescent peptide sensor containing tryptophan (donor) and dansyl fluorophore (acceptor) was synthesized for monitoring heavy and transition metal (HTM) ions on the basis of metal ion binding motif (Cys-X-X-X-Cys). The peptide probe successfully exhibited a turn on and ratiometric response for several heavy metal ions such as Hg(2+), Cd(2+), Pb(2+), Zn(2+), and Ag(+) in aqueous solution. The enhancements of emission intensity were achieved in the presence of the HTM ions by fluorescent resonance energy transfer (FRET) and chelation enhanced fluorescence (CHEF) effects. The detection limits of the sensor for Cd(2+), Pb(2+), Zn(2+), and Ag(+) were lower than the EPA's drinking water maximum contaminant levels (MCL). We described the fluorescent enhancement, binding affinity, and detection limit of the peptide probe for HTM ions.

Cross-Modal Binding in Developmental Dyslexia

ERIC Educational Resources Information Center

Jones, Manon W.; Branigan, Holly P.; Parra, Mario A.; Logie, Robert H.

2013-01-01

The ability to learn visual-phonological associations is a unique predictor of word reading, and individuals with developmental dyslexia show impaired ability in learning these associations. In this study, we compared developmentally dyslexic and nondyslexic adults on their ability to form cross-modal associations (or "bindings") based…

Molecular dynamics and principal components of potassium binding with human telomeric intra-molecular G-quadruplex.

PubMed

Wang, Zhiguo; Chen, Ruping; Hou, Ling; Li, Jianfeng; Liu, Jun-Ping

2015-06-01

Telomere assumes intra-molecular G-quadruplex that is a significant drug target for inhibiting telomerase maintenance of telomeres in cancer. Metal cations have been recognized as playing important roles in stabilizing G-quadruplex, but their binding processes to human telomeric G-quadruplex remain uncharacterized. To investigate the detailed binding procedures, molecular dynamics simulations were conducted on the hybrid [3 + 1] form-one human telomeric intra-molecular G-quadruplex. We show here that the binding of a potassium ion to a G-tetrad core is mediated by two alternative pathways. Principal component analysis illustrated the dominant concerted motions of G-quadruplex occurred at the loop domains. MM-PBSA calculations revealed that binding was energetically favorable and driven by the electrostatic interactions. The lower binding site was found more constructive favorable for binding. Our data provide useful information on a potassium-mediated stable structure of human telomeric intra-molecular G-quadruplex, implicating in ion disorder associated conformational changes and targeted drug design.

Estimation of the binding ability of main transport proteins of blood plasma with liver cirrhosis by the fluorescent probe method

NASA Astrophysics Data System (ADS)

Korolenko, E. A.; Korolik, E. V.; Korolik, A. K.; Kirkovskii, V. V.

2007-07-01

We present results from an investigation of the binding ability of the main transport proteins (albumin, lipoproteins, and α-1-acid glycoprotein) of blood plasma from patients at different stages of liver cirrhosis by the fluorescent probe method. We used the hydrophobic fluorescent probes anionic 8-anilinonaphthalene-1-sulfonate, which interacts in blood plasma mainly with albumin; cationic Quinaldine red, which interacts with α-1-acid glycoprotein; and neutral Nile red, which redistributes between lipoproteins and albumin in whole blood plasma. We show that the binding ability of albumin and α-1-acid glycoprotein to negatively charged and positively charged hydrophobic metabolites, respectively, increases in the compensation stage of liver cirrhosis. As the pathology process deepens and transitions into the decompensation stage, the transport abilities of albumin and α-1-acid glycoprotein decrease whereas the binding ability of lipoproteins remains high.

Novel selective and sensitive optical chemosensor based on phenylfluorone derivative for detection of Ge(IV) ion in aqueous solution.

PubMed

Keawwangchai, Somchai; Morakot, Nongnit; Keawwangchai, Tasawan

2018-09-05

A water soluble chemosensor for Ge 4+ ion based on fluorone derivative containing 3,4-bis(2-(diethylamino)-2-oxoethoxy)phenyl (R8) has been synthesized. The binding abilities between R8 and 10 equiv. of Na + , K + , Ca 2+ , Fe 2+ , Cu 2+ , Cd 2+ , Hg 2+ , Pb 2+ , Al 3+ , Cr 3+ , Fe 3+ and Ge 4+ ions in 1% v/v EtOH-water (tris-buffer pH 7.0) were studied using UV-vis and fluorescence spectrophotometry. When observed by naked-eyes, the color of R8 changed from yellow-orange to pink and the fluorescent color changed from green to non-fluorescence when complexed with Ge 4+ ion. The spectral analysis showed that UV-vis absorption and fluorescence emission intensity of R8 decreased dramatically when Ge 4+ ion was added comparing with other ions. To explain this behavior, the quantum calculation was performed using the hybrid density functional at B3LYP /LanL2DZ level of theory. The calculated orbital energies indicated that the decreasing of UV-vis absorption and the quenching of fluorescence were due to the complexation induced metal to ligand charge transfer. The association constants (K a ) of R8-Ge 4+ complexes calculated from Benesi-Hildebrand equation was 6.21 × 10 5  M -1 . The UV-vis detection limit for Ge 4+ was 4.40 × 10 -7  M which was three orders of magnitude lower than those of Al 3+ , Cd 2+ , Cu 2+ and Na + ion. Copyright © 2018 Elsevier B.V. All rights reserved.

Effects of magnesium ions on the stabilization of RNA oligomers of defined structures.

PubMed Central

Serra, Martin J; Baird, John D; Dale, Taraka; Fey, Bridget L; Retatagos, Kimberly; Westhof, Eric

2002-01-01

Optical melting was used to determine the stabilities of 11 small RNA oligomers of defined secondary structure as a function of magnesium ion concentration. The oligomers included helices composed of Watson-Crick base pairs, GA tandem base pairs, GU tandem base pairs, and loop E motifs (both eubacterial and eukaryotic). The effect of magnesium ion concentration on stability was interpreted in terms of two simple models. The first assumes an uptake of metal ion upon duplex formation. The second assumes nonspecific electrostatic attraction of metal ions to the RNA oligomer. For all oligomers, except the eubacterial loop E, the data could best be interpreted as nonspecific binding of metal ions to the RNAs. The effect of magnesium ions on the stability of the eubacterial loop E was distinct from that seen with the other oligomers in two ways. First, the extent of stabilization by magnesium ions (as measured by either change in melting temperature or free energy) was three times greater than that observed for the other helical oligomers. Second, the presence of magnesium ions produces a doubling of the enthalpy for the melting transition. These results indicate that magnesium ion stabilizes the eubacterial loop E sequence by chelating the RNA specifically. Further, these results on a rather small system shed light on the large enthalpy changes observed upon thermal unfolding of large RNAs like group I introns. It is suggested that parts of those large enthalpy changes observed in the folding of RNAs may be assigned to variations in the hydration states and types of coordinating atoms in some specifically bound magnesium ions and to an increase in the observed cooperativity of the folding transition due to the binding of those magnesium ions coupling the two stems together. Brownian dynamic simulations, carried out to visualize the metal ion binding sites, reveal rather delocalized ionic densities in all oligomers, except for the eubacterial loop E, in which precisely located ion densities were previously calculated. PMID:12003491

Method Development for Binding Media Analysis in Painting Cross-Sections by Desorption Electrospray Ionization-Mass Spectrometry (DESI-MS).

PubMed

Watts, Kristen; Lagalante, Anthony

2018-06-06

Art conservation science is in need of a relatively nondestructive way of rapidly identifying the binding media within a painting cross-section and isolating binding media to specific layers within the cross-section. Knowledge of the stratigraphy of cross-sections can be helpful for removing possible unoriginal paint layers on the artistic work. Desorption electrospray ionization-mass spectrometry (DESI-MS) was used in ambient mode to study cross-sections from mock-up layered paint samples and samples from a 17th century baroque painting. The DESI spray was raster scanned perpendicular to the cross-section layers to maximize lateral resolution then analyzed with a triple quadrupole mass analyzer in linear ion trap mode. From these scans, isobaric mass maps were created to map the locations of masses indicative of particular binding media onto the cross-sections. Line paint-outs of pigments in different binding media showed specific and unique ions to distinguish between the modern acrylic media and the lipid containing binding media. This included: OP (EO) 9 surfactant in positive ESI for acrylic (m/z 621), and oleic (m/z 281), stearic (m/z 283), and azelaic (m/z 187) acids in negative ESI for oil and egg tempera. DESI-MS maps of mock-up cross-sections of layered pigmented binding media showed correlation between these ions and the layers with a spatial resolution of 100 μm. DESI-MS is effective in monitoring binding media within an intact painting cross-section via mass spectrometric methods. This includes distinguishing between lipid-containing and modern binding materials present in a known mockup cross section matrix as well as identifying lipid binding media in a 17th century baroque era painting. This article is protected by copyright. All rights reserved.

Understanding Ion Binding Affinity and Selectivity in β-Parvalbumin Using Molecular Dynamics and Mean Spherical Approximation Theory.

PubMed

Kucharski, Amir N; Scott, Caitlin E; Davis, Jonathan P; Kekenes-Huskey, Peter M

2016-08-25

Parvalbumin (PV) is a globular calcium (Ca(2+))-selective protein expressed in a variety of biological tissues. Our computational studies of the rat β-parvalbumin (β-PV) isoform seek to elucidate the molecular thermodynamics of Ca(2+) versus magnesium (Mg(2+)) binding at the protein's two EF-hand motifs. Specifically, we have utilized molecular dynamics (MD) simulations and a mean-field electrolyte model (mean spherical approximation (MSA) theory) to delineate how the EF-hand scaffold controls the "local" thermodynamics of Ca(2+) binding selectivity over Mg(2+). Our MD simulations provide the probability density of metal-chelating oxygens within the EF-hand scaffolds for both Ca(2+) and Mg(2+), as well the conformational strain induced by Mg(2+) relative to Ca(2+) binding. MSA theory utilizes the binding domain oxygen and charge distributions to predict the chemical potential of ion binding, as well as their corresponding concentrations within the binding domain. We find that the electrostatic and steric contributions toward ion binding were similar for Mg(2+) and Ca(2+), yet the latter was 5.5 kcal/mol lower in enthalpy when internal strain within the EF hand was considered. We therefore speculate that beyond differences in dehydration energies for the Ca(2+) versus Mg(2+), strain induced in the β-PV EF hand by cation binding significantly contributes to the nearly 10,000-fold difference in binding affinity reported in the literature. We further complemented our analyses of local factors governing cation binding selectivity with whole-protein (global) contributions, such as interhelical residue-residue contacts and solvent exposure of hydrophobic surface. These contributions were found to be comparable for both Ca(2+)- and Mg(2+)-bound β-PV, which may implicate local factors, EF-hand strain, and dehydration, in providing the primary means of selectivity. We anticipate these methods could be used to estimate metal binding thermodynamics across a broad range of PV sequence homologues and EF-hand-containing, Ca(2+) binding proteins.

Multi-stimuli-responsive organometallic gels based on ferrocene-linked poly(aryl ether) dendrons: reversible redox switching and Pb2+-ion sensing.

PubMed

Lakshmi, Neelakandan Vidhya; Mandal, Dipendu; Ghosh, Sundargopal; Prasad, Edamana

2014-07-14

We describe the design, synthesis, and "stimuli-responsive" study of ferrocene-linked Fréchet-type [poly(aryl ether)]-dendron-based organometallic gels, in which the ferrocene moiety is attached to the dendron framework through an acyl hydrazone linkage. The low-molecular-weight gelators (LMWGs) form robust gels in both polar and non-polar solvent/solvent mixtures. The organometallic gels undergo stimuli-responsive behavior through 1) thermal, 2) chemical, and 3) electrochemical methods. Among them, conditions 1 and 3 lead to seamlessly reversible with repeated cycles of identical efficiency. Results indicate that the flexible nature of the poly(aryl ether) dendron framework plays a key role in retaining the reversible electrochemical behavior of ferrocene moiety in the LMWGs. Further, the organometallic gelators have exhibited unique selectivity towards Pb(2+) ions (detection limit ≈10(-8)  M). The metal ion-sensing results in a gel-sol phase transition associated with a color change visible to the naked eye. Most importantly, decomplexing the metal ion from the system leads to the regeneration of the initial gel morphology, indicating the restoring ability of the organometallic gel. The metal-ligand binding nature has been analyzed by using (1)H NMR spectroscopy, mass spectrometry, and DFT calculations. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

Actions and Mechanisms of Polyunsaturated Fatty Acids on Voltage-Gated Ion Channels.

PubMed

Elinder, Fredrik; Liin, Sara I

2017-01-01

Polyunsaturated fatty acids (PUFAs) act on most ion channels, thereby having significant physiological and pharmacological effects. In this review we summarize data from numerous PUFAs on voltage-gated ion channels containing one or several voltage-sensor domains, such as voltage-gated sodium (Na V ), potassium (K V ), calcium (Ca V ), and proton (H V ) channels, as well as calcium-activated potassium (K Ca ), and transient receptor potential (TRP) channels. Some effects of fatty acids appear to be channel specific, whereas others seem to be more general. Common features for the fatty acids to act on the ion channels are at least two double bonds in cis geometry and a charged carboxyl group. In total we identify and label five different sites for the PUFAs. PUFA site 1 : The intracellular cavity. Binding of PUFA reduces the current, sometimes as a time-dependent block, inducing an apparent inactivation. PUFA site 2 : The extracellular entrance to the pore. Binding leads to a block of the channel. PUFA site 3 : The intracellular gate. Binding to this site can bend the gate open and increase the current. PUFA site 4 : The interface between the extracellular leaflet of the lipid bilayer and the voltage-sensor domain. Binding to this site leads to an opening of the channel via an electrostatic attraction between the negatively charged PUFA and the positively charged voltage sensor. PUFA site 5 : The interface between the extracellular leaflet of the lipid bilayer and the pore domain. Binding to this site affects slow inactivation. This mapping of functional PUFA sites can form the basis for physiological and pharmacological modifications of voltage-gated ion channels.

Actions and Mechanisms of Polyunsaturated Fatty Acids on Voltage-Gated Ion Channels

PubMed Central

Elinder, Fredrik; Liin, Sara I.

2017-01-01

Polyunsaturated fatty acids (PUFAs) act on most ion channels, thereby having significant physiological and pharmacological effects. In this review we summarize data from numerous PUFAs on voltage-gated ion channels containing one or several voltage-sensor domains, such as voltage-gated sodium (NaV), potassium (KV), calcium (CaV), and proton (HV) channels, as well as calcium-activated potassium (KCa), and transient receptor potential (TRP) channels. Some effects of fatty acids appear to be channel specific, whereas others seem to be more general. Common features for the fatty acids to act on the ion channels are at least two double bonds in cis geometry and a charged carboxyl group. In total we identify and label five different sites for the PUFAs. PUFA site 1: The intracellular cavity. Binding of PUFA reduces the current, sometimes as a time-dependent block, inducing an apparent inactivation. PUFA site 2: The extracellular entrance to the pore. Binding leads to a block of the channel. PUFA site 3: The intracellular gate. Binding to this site can bend the gate open and increase the current. PUFA site 4: The interface between the extracellular leaflet of the lipid bilayer and the voltage-sensor domain. Binding to this site leads to an opening of the channel via an electrostatic attraction between the negatively charged PUFA and the positively charged voltage sensor. PUFA site 5: The interface between the extracellular leaflet of the lipid bilayer and the pore domain. Binding to this site affects slow inactivation. This mapping of functional PUFA sites can form the basis for physiological and pharmacological modifications of voltage-gated ion channels. PMID:28220076

Binding of Alkali Metal Ions with 1,3,5-Tri(phenyl)benzene and 1,3,5-Tri(naphthyl)benzene: The Effect of Phenyl and Naphthyl Ring Substitution on Cation-π Interactions Revealed by DFT Study.

PubMed

Mirchi, Ali; Sizochenko, Natalia; Dinadayalane, Tandabany; Leszczynski, Jerzy

2017-11-22

The effect of substitution of phenyl and naphthyl rings to benzene was examined to elucidate the cation-π interactions involving alkali metal ions with 1,3,5-tri(phenyl)benzene (TPB) and 1,3,5-tri(naphthyl)benzene (TNB). Benzene, TPB, and four TNB isomers (with ααα, ααβ, αββ, and βββ types of fusion) and their complexes with Li + , Na + , K + , Rb + , and Cs + were optimized using DFT approach with B3LYP and M06-2X functionals in conjunction with the def2-QZVP basis set. Higher relative stability of β,β,β-TNB over α,α,α-TNB can be attributed to peri repulsion, which is defined as the nonbonding repulsive interaction between substituents in the 1- and the 8-positions on the naphthalene core. Binding energies, distances between ring centroid and the metal ions, and the distance to metal ions from the center of other six-membered rings were compared for all complexes. Our computational study reveals that the binding affinity of alkali metal cations increases significantly with the 1,3,5-trisubstitution of phenyl and naphthyl rings to benzene. The detailed computational analyses of geometries, partial charges, binding energies, and ligand organization energies reveal the possibility of favorable C-H···M + interactions when a α-naphthyl group exists in complexes of TNB structures. Like benzene-alkali metal ion complexes, the binding affinity of metal ions follows the order: Li + > Na + > K + > Rb + > Cs + for any considered 1,3,5-trisubstituted benzene systems. In case of TNB, we found that the strength of interactions increases as the fusion point changes from α to β position of naphthalene.

Crystal structures reveal metal-binding plasticity at the metallo-β-lactamase active site of PqqB from Pseudomonas putida

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

Tu, Xiongying; Latham, John A.; Klema, Valerie J.

PqqB is an enzyme involved in the biosynthesis of pyrroloquinoline quinone and a distal member of the metallo-β-lactamase (MBL) superfamily. PqqB lacks two residues in the conserved signature motif HxHxDH that makes up the key metal-chelating elements that can bind up to two metal ions at the active site of MBLs and other members of its superfamily. Here, we report crystal structures of PqqB bound to Mn2+, Mg2+, Cu2+, and Zn2+. These structures demonstrate that PqqB can still bind metal ions at the canonical MBL active site. The fact that PqqB can adapt its side chains to chelate a widemore » spectrum of metal ions with different coordination features on a uniform main chain scaffold demonstrates its metal-binding plasticity. This plasticity may provide insights into the structural basis of promiscuous activities found in ensembles of metal complexes within this superfamily. Furthermore, PqqB belongs to a small subclass of MBLs that contain an additional CxCxxC motif that binds a structural Zn2+. Our data support a key role for this motif in dimerization.« less

Barbiturates Bind in the GLIC Ion Channel Pore and Cause Inhibition by Stabilizing a Closed State*♦

PubMed Central

Fourati, Zaineb; Ruza, Reinis Reinholds; Laverty, Duncan; Drège, Emmanuelle; Delarue-Cochin, Sandrine; Joseph, Delphine; Koehl, Patrice; Smart, Trevor; Delarue, Marc

2017-01-01

Barbiturates induce anesthesia by modulating the activity of anionic and cationic pentameric ligand-gated ion channels (pLGICs). Despite more than a century of use in clinical practice, the prototypic binding site for this class of drugs within pLGICs is yet to be described. In this study, we present the first X-ray structures of barbiturates bound to GLIC, a cationic prokaryotic pLGIC with excellent structural homology to other relevant channels sensitive to general anesthetics and, as shown here, to barbiturates, at clinically relevant concentrations. Several derivatives of barbiturates containing anomalous scatterers were synthesized, and these derivatives helped us unambiguously identify a unique barbiturate binding site within the central ion channel pore in a closed conformation. In addition, docking calculations around the observed binding site for all three states of the receptor, including a model of the desensitized state, showed that barbiturates preferentially stabilize the closed state. The identification of this pore binding site sheds light on the mechanism of barbiturate inhibition of cationic pLGICs and allows the rationalization of several structural and functional features previously observed for barbiturates. PMID:27986812

How Cations Can Assist DNase I in DNA Binding and Hydrolysis

PubMed Central

Guéroult, Marc; Picot, Daniel; Abi-Ghanem, Joséphine; Hartmann, Brigitte; Baaden, Marc

2010-01-01

DNase I requires Ca2+ and Mg2+ for hydrolyzing double-stranded DNA. However, the number and the location of DNase I ion-binding sites remain unclear, as well as the role of these counter-ions. Using molecular dynamics simulations, we show that bovine pancreatic (bp) DNase I contains four ion-binding pockets. Two of them strongly bind Ca2+ while the other two sites coordinate Mg2+. These theoretical results are strongly supported by revisiting crystallographic structures that contain bpDNase I. One Ca2+ stabilizes the functional DNase I structure. The presence of Mg2+ in close vicinity to the catalytic pocket of bpDNase I reinforces the idea of a cation-assisted hydrolytic mechanism. Importantly, Poisson-Boltzmann-type electrostatic potential calculations demonstrate that the divalent cations collectively control the electrostatic fit between bpDNase I and DNA. These results improve our understanding of the essential role of cations in the biological function of bpDNase I. The high degree of conservation of the amino acids involved in the identified cation-binding sites across DNase I and DNase I-like proteins from various species suggests that our findings generally apply to all DNase I-DNA interactions. PMID:21124947

Synthesis of rigidified flavin–guanidinium ion conjugates and investigation of their photocatalytic properties

PubMed Central

Schmaderer, Harald; Bhuyan, Mouchumi

2009-01-01

Summary Flavin chromophores can mediate redox reactions upon irradiation by blue light. In an attempt to increase their catalytic efficacy, flavin derivatives bearing a guanidinium ion as oxoanion binding site were prepared. Chromophore and substrate binding site are linked by a rigid Kemp’s acid structure. The molecular structure of the new flavins was confirmed by an X-ray structure analysis and their photocatalytic activity was investigated in benzyl ester cleavage, nitroarene reduction and a Diels–Alder reaction. The modified flavins photocatalyze the reactions, but the introduced substrate binding site does not enhance their performance. PMID:19590745

Synthesis of rigidified flavin-guanidinium ion conjugates and investigation of their photocatalytic properties.

PubMed

Schmaderer, Harald; Bhuyan, Mouchumi; König, Burkhard

2009-05-28

Flavin chromophores can mediate redox reactions upon irradiation by blue light. In an attempt to increase their catalytic efficacy, flavin derivatives bearing a guanidinium ion as oxoanion binding site were prepared. Chromophore and substrate binding site are linked by a rigid Kemp's acid structure. The molecular structure of the new flavins was confirmed by an X-ray structure analysis and their photocatalytic activity was investigated in benzyl ester cleavage, nitroarene reduction and a Diels-Alder reaction. The modified flavins photocatalyze the reactions, but the introduced substrate binding site does not enhance their performance.

Reversible ion transportation switch by a ligand-gated synthetic supramolecular ion channel.

PubMed

Muraoka, Takahiro; Endo, Takahiro; Tabata, Kazuhito V; Noji, Hiroyuki; Nagatoishi, Satoru; Tsumoto, Kouhei; Li, Rui; Kinbara, Kazushi

2014-11-05

Inspired by the regulation of cellular activities found in the ion channel proteins, here we developed membrane-embedded synthetic chiral receptors 1 and 2 with different terminal structures, where receptor 1 has hydrophobic triisopropylsilyl (TIPS) groups and receptor 2 has hydrophilic hydroxy groups. The receptors have ligand-binding units that interact with cationic amphiphiles such as 2-phenethylamine (PA). Conductance study revealed that the receptors hardly show ion transportation at the ligand-free state. After ligand binding involving a conformational change, receptor 1 bearing TIPS termini displays a significant current enhancement due to ion transportation. The current substantially diminishes upon addition of β-cyclodextrin (βCD) that scavenges the ligand from the receptor. Importantly, the receptor again turns into the conductive state by the second addition of PA, and the activation/deactivation of the ion transportation can be repeated. In contrast, receptor 2 bearing the hydroxy terminal groups hardly exhibits ion transportation, suggesting the importance of terminal TIPS groups of 1 that likely anchor the receptor in the membrane.

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

Moving Fe2+ from ferritin ion channels to catalytic OH centers depends on conserved protein cage carboxylates.

PubMed

Behera, Rabindra K; Theil, Elizabeth C

2014-06-03

Ferritin biominerals are protein-caged metabolic iron concentrates used for iron-protein cofactors and oxidant protection (Fe(2+) and O2 sequestration). Fe(2+) passage through ion channels in the protein cages, like membrane ion channels, required for ferritin biomineral synthesis, is followed by Fe(2+) substrate movement to ferritin enzyme (Fox) sites. Fe(2+) and O2 substrates are coupled via a diferric peroxo (DFP) intermediate, λmax 650 nm, which decays to [Fe(3+)-O-Fe(3+)] precursors of caged ferritin biominerals. Structural studies show multiple conformations for conserved, carboxylate residues E136 and E57, which are between ferritin ion channel exits and enzymatic sites, suggesting functional connections. Here we show that E136 and E57 are required for ferritin enzyme activity and thus are functional links between ferritin ion channels and enzymatic sites. DFP formation (Kcat and kcat/Km), DFP decay, and protein-caged hydrated ferric oxide accumulation decreased in ferritin E57A and E136A; saturation required higher Fe(2+) concentrations. Divalent cations (both ion channel and intracage binding) selectively inhibit ferritin enzyme activity (block Fe(2+) access), Mn(2+) < Co(2+) < Cu(2+) < Zn(2+), reflecting metal ion-protein binding stabilities. Fe(2+)-Cys126 binding in ferritin ion channels, observed as Cu(2+)-S-Cys126 charge-transfer bands in ferritin E130D UV-vis spectra and resistance to Cu(2+) inhibition in ferritin C126S, was unpredicted. Identifying E57 and E136 links in Fe(2+) movement from ferritin ion channels to ferritin enzyme sites completes a bucket brigade that moves external Fe(2+) into ferritin enzymatic sites. The results clarify Fe(2+) transport within ferritin and model molecular links between membrane ion channels and cytoplasmic destinations.

The binding energies of one and two water molecules to the first transition-row metal positive ions. II

NASA Technical Reports Server (NTRS)

Rosi, Marzio; Bauschlicher, Charles W., Jr.

1990-01-01

The present investigation of H2O's binding energy to transition-metal ions proceeds from the D(2h) structure and bends the two water molecules out of plane. The molecule is constrained to have C(2v) symmetry, so that each water molecule and metal ion lies on a plane. The ground states are bent only for Mn(H2O)2(+) and Zn(H2O)2(+), where only 4s4p hybridization is energetically favorable; 4s4p hybridization reduces repulsion.

Electronic and Spatial Structures of Water-Soluble Dinitrosyl Iron Complexes with Thiol-Containing Ligands Underlying Their Ability to Act as Nitric Oxide and Nitrosonium Ion Donors

PubMed Central

Vanin, Anatoly F.; Burbaev, Dosymzhan Sh.

2011-01-01

The ability of mononuclear dinitrosyl iron commplexes (M-DNICs) with thiolate ligands to act as NO donors and to trigger S-nitrosation of thiols can be explain only in the paradigm of the model of the [Fe+(NO+)2] core ({Fe(NO)2}7 according to the Enemark-Feltham classification). Similarly, the {(RS−)2Fe+(NO+)2}+ structure describing the distribution of unpaired electron density in M-DNIC corresponds to the low-spin (S = 1/2) state with a d7 electron configuration of the iron atom and predominant localization of the unpaired electron on MO(dz2) and the square planar structure of M-DNIC. On the other side, the formation of molecular orbitals of M-DNIC including orbitals of the iron atom, thiolate and nitrosyl ligands results in a transfer of electron density from sulfur atoms to the iron atom and nitrosyl ligands. Under these conditions, the positive charge on the nitrosyl ligands diminishes appreciably, the interaction of the ligands with hydroxyl ions or with thiols slows down and the hydrolysis of nitrosyl ligands and the S-nitrosating effect of the latter are not manifested. Most probably, the S-nitrosating effect of nitrosyl ligands is a result of weak binding of thiolate ligands to the iron atom under conditions favoring destabilization of M-DNIC. PMID:22505886

Electronic and spatial structures of water-soluble dinitrosyl iron complexes with thiol-containing ligands underlying their ability to act as nitric oxide and nitrosonium ion donors.

PubMed

Vanin, Anatoly F; Burbaev, Dosymzhan Sh

2011-01-01

The ability of mononuclear dinitrosyl iron commplexes (M-DNICs) with thiolate ligands to act as NO donors and to trigger S-nitrosation of thiols can be explain only in the paradigm of the model of the [Fe(+)(NO(+))(2)] core ({Fe(NO)(2)}(7) according to the Enemark-Feltham classification). Similarly, the {(RS(-))(2)Fe(+)(NO(+))(2)}(+) structure describing the distribution of unpaired electron density in M-DNIC corresponds to the low-spin (S = 1/2) state with a d(7) electron configuration of the iron atom and predominant localization of the unpaired electron on MO(d(z2)) and the square planar structure of M-DNIC. On the other side, the formation of molecular orbitals of M-DNIC including orbitals of the iron atom, thiolate and nitrosyl ligands results in a transfer of electron density from sulfur atoms to the iron atom and nitrosyl ligands. Under these conditions, the positive charge on the nitrosyl ligands diminishes appreciably, the interaction of the ligands with hydroxyl ions or with thiols slows down and the hydrolysis of nitrosyl ligands and the S-nitrosating effect of the latter are not manifested. Most probably, the S-nitrosating effect of nitrosyl ligands is a result of weak binding of thiolate ligands to the iron atom under conditions favoring destabilization of M-DNIC.

Pyrrolo-dC Metal-Mediated Base Pairs in the Reverse Watson-Crick Double Helix: Enhanced Stability of Parallel DNA and Impact of 6-Pyridinyl Residues on Fluorescence and Silver-Ion Binding.

PubMed

Yang, Haozhe; Mei, Hui; Seela, Frank

2015-07-06

Reverse Watson-Crick DNA with parallel-strand orientation (ps DNA) has been constructed. Pyrrolo-dC (PyrdC) nucleosides with phenyl and pyridinyl residues linked to the 6 position of the pyrrolo[2,3-d]pyrimidine base have been incorporated in 12- and 25-mer oligonucleotide duplexes and utilized as silver-ion binding sites. Thermal-stability studies on the parallel DNA strands demonstrated extremely strong silver-ion binding and strongly enhanced duplex stability. Stoichiometric UV and fluorescence titration experiments verified that a single (2py) PyrdC-(2py) PyrdC pair captures two silver ions in ps DNA. A structure for the PyrdC silver-ion base pair that aligns 7-deazapurine bases head-to-tail instead of head-to-head, as suggested for canonical DNA, is proposed. The silver DNA double helix represents the first example of a ps DNA structure built up of bidentate and tridentate reverse Watson-Crick base pairs stabilized by a dinuclear silver-mediated PyrdC pair. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

Oxidation promotes insertion of the CLIC1 chloride intracellular channel into the membrane.

PubMed

Goodchild, Sophia C; Howell, Michael W; Cordina, Nicole M; Littler, Dene R; Breit, Samuel N; Curmi, Paul M G; Brown, Louise Jennifer

2009-12-01

Members of the chloride intracellular channel (CLIC) family exist primarily as soluble proteins but can also auto-insert into cellular membranes to form ion channels. While little is known about the process of CLIC membrane insertion, a unique feature of mammalian CLIC1 is its ability to undergo a dramatic structural metamorphosis between a monomeric glutathione-S-transferase homolog and an all-helical dimer upon oxidation in solution. Whether this oxidation-induced metamorphosis facilitates CLIC1 membrane insertion is unclear. In this work, we have sought to characterise the role of oxidation in the process of CLIC1 membrane insertion. We examined how redox conditions modify the ability of CLIC1 to associate with and insert into the membrane using fluorescence quenching studies and a sucrose-loaded vesicle sedimentation assay to measure membrane binding. Our results suggest that oxidation of monomeric CLIC1, in the presence of membranes, promotes insertion into the bilayer more effectively than the oxidised CLIC1 dimer.

Effect of cathodic polarization on coating doxycycline on titanium surfaces.

PubMed

Geißler, Sebastian; Tiainen, Hanna; Haugen, Håvard J

2016-06-01

Cathodic polarization has been reported to enhance the ability of titanium based implant materials to interact with biomolecules by forming titanium hydride at the outermost surface layer. Although this hydride layer has recently been suggested to allow the immobilization of the broad spectrum antibiotic doxycycline on titanium surfaces, the involvement of hydride in binding the biomolecule onto titanium remains poorly understood. To gain better understanding of the influence this immobilization process has on titanium surfaces, mirror-polished commercially pure titanium surfaces were cathodically polarized in the presence of doxycycline and the modified surfaces were thoroughly characterized using atomic force microscopy, electron microscopy, secondary ion mass spectrometry, and angle-resolved X-ray spectroscopy. We demonstrated that no hydride was created during the polarization process. Doxycycline was found to be attached to an oxide layer that was modified during the electrochemical process. A bacterial assay using bioluminescent Staphylococcus epidermidis Xen43 showed the ability of the coating to reduce bacterial colonization and planktonic bacterial growth. Copyright © 2016 Elsevier B.V. All rights reserved.

HDAC6–p97/VCP controlled polyubiquitin chain turnover

PubMed Central

Boyault, Cyril; Gilquin, Benoit; Zhang, Yu; Rybin, Vladimir; Garman, Elspeth; Meyer-Klaucke, Wolfram; Matthias, Patrick; Müller, Christoph W; Khochbin, Saadi

2006-01-01

HDAC6 is a unique cytoplasmic deacetylase capable of interacting with ubiquitin. Using a combination of biophysical, biochemical and biological approaches, we have characterized the ubiquitin-binding domain of HDAC6, named ZnF-UBP, and investigated its biological functions. These studies show that the three Zn ion-containing HDAC6 ZnF-UBP domain presents the highest known affinity for ubiquitin monomers and mediates the ability of HDAC6 to negatively control the cellular polyubiquitin chain turnover. We further show that HDAC6-interacting chaperone, p97/VCP, dissociates the HDAC6–ubiquitin complexes and counteracts the ability of HDAC6 to promote the accumulation of polyubiquitinated proteins. We propose that a finely tuned balance of HDAC6 and p97/VCP concentrations determines the fate of ubiquitinated misfolded proteins: p97/VCP would promote protein degradation and ubiquitin turnover, whereas HDAC6 would favour the accumulation of ubiquitinated protein aggregates and inclusion body formation. PMID:16810319

Factor H binds to the hypervariable region of many Streptococcus pyogenes M proteins but does not promote phagocytosis resistance or acute virulence.

PubMed

Gustafsson, Mattias C U; Lannergård, Jonas; Nilsson, O Rickard; Kristensen, Bodil M; Olsen, John E; Harris, Claire L; Ufret-Vincenty, Rafael L; Stålhammar-Carlemalm, Margaretha; Lindahl, Gunnar

2013-01-01

Many pathogens express a surface protein that binds the human complement regulator factor H (FH), as first described for Streptococcus pyogenes and the antiphagocytic M6 protein. It is commonly assumed that FH recruited to an M protein enhances virulence by protecting the bacteria against complement deposition and phagocytosis, but the role of FH-binding in S. pyogenes pathogenesis has remained unclear and controversial. Here, we studied seven purified M proteins for ability to bind FH and found that FH binds to the M5, M6 and M18 proteins but not the M1, M3, M4 and M22 proteins. Extensive immunochemical analysis indicated that FH binds solely to the hypervariable region (HVR) of an M protein, suggesting that selection has favored the ability of certain HVRs to bind FH. These FH-binding HVRs could be studied as isolated polypeptides that retain ability to bind FH, implying that an FH-binding HVR represents a distinct ligand-binding domain. The isolated HVRs specifically interacted with FH among all human serum proteins, interacted with the same region in FH and showed species specificity, but exhibited little or no antigenic cross-reactivity. Although these findings suggested that FH recruited to an M protein promotes virulence, studies in transgenic mice did not demonstrate a role for bound FH during acute infection. Moreover, phagocytosis tests indicated that ability to bind FH is neither sufficient nor necessary for S. pyogenes to resist killing in whole human blood. While these data shed new light on the HVR of M proteins, they suggest that FH-binding may affect S. pyogenes virulence by mechanisms not assessed in currently used model systems.

Stability and Sugar Recognition Ability of Ricin-Like Carbohydrate Binding Domains

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

Yao, Jianzhuang; Nellas, Ricky B; Glover, Mary M

2011-01-01

Lectins are a class of proteins known for their novel binding to saccharides. Understanding this sugar recognition process can be crucial in creating structure-based designs of proteins with various biological roles. We focus on the sugar binding of a particular lectin, ricin, which has two -trefoil carbohydrate-binding domains (CRDs) found in several plant protein toxins. The binding ability of possible sites of ricin-like CRD has been puzzling. The apo and various (multiple) ligand-bound forms of the sugar-binding domains of ricin were studied by molecular dynamics simulations. By evaluating structural stability, hydrogen bond dynamics, flexibility, and binding energy, we obtained amore » detailed picture of the sugar recognition of the ricin-like CRD. Unlike what was previously believed, we found that the binding abilities of the two known sites are not independent of each other. The binding ability of one site is positively affected by the other site. While the mean positions of different binding scenarios are not altered significantly, the flexibility of the binding pockets visibly decreases upon multiple ligand binding. This change in flexibility seems to be the origin of the binding cooperativity. All the hydrogen bonds that are strong in the monoligand state are also strong in the double-ligand complex, although the stability is much higher in the latter form due to cooperativity. These strong hydrogen bonds in a monoligand state are deemed to be the essential hydrogen bonds. Furthermore, by examining the structural correlation matrix, the two domains are structurally one entity. Galactose hydroxyl groups, OH4 and OH3, are the most critical parts in both site 1 and site 2 recognition.« less

Factor H Binds to the Hypervariable Region of Many Streptococcus pyogenes M Proteins but Does Not Promote Phagocytosis Resistance or Acute Virulence

PubMed Central

Kristensen, Bodil M.; Olsen, John E.; Harris, Claire L.; Ufret-Vincenty, Rafael L.; Stålhammar-Carlemalm, Margaretha; Lindahl, Gunnar

2013-01-01

Many pathogens express a surface protein that binds the human complement regulator factor H (FH), as first described for Streptococcus pyogenes and the antiphagocytic M6 protein. It is commonly assumed that FH recruited to an M protein enhances virulence by protecting the bacteria against complement deposition and phagocytosis, but the role of FH-binding in S. pyogenes pathogenesis has remained unclear and controversial. Here, we studied seven purified M proteins for ability to bind FH and found that FH binds to the M5, M6 and M18 proteins but not the M1, M3, M4 and M22 proteins. Extensive immunochemical analysis indicated that FH binds solely to the hypervariable region (HVR) of an M protein, suggesting that selection has favored the ability of certain HVRs to bind FH. These FH-binding HVRs could be studied as isolated polypeptides that retain ability to bind FH, implying that an FH-binding HVR represents a distinct ligand-binding domain. The isolated HVRs specifically interacted with FH among all human serum proteins, interacted with the same region in FH and showed species specificity, but exhibited little or no antigenic cross-reactivity. Although these findings suggested that FH recruited to an M protein promotes virulence, studies in transgenic mice did not demonstrate a role for bound FH during acute infection. Moreover, phagocytosis tests indicated that ability to bind FH is neither sufficient nor necessary for S. pyogenes to resist killing in whole human blood. While these data shed new light on the HVR of M proteins, they suggest that FH-binding may affect S. pyogenes virulence by mechanisms not assessed in currently used model systems. PMID:23637608

Interaction analysis of chimeric metal-binding green fluorescent protein and artificial solid-supported lipid membrane by quartz crystal microbalance and atomic force microscopy.

PubMed

Prachayasittikul, Virapong; Isarankura Na Ayudhya, Chartchalerm; Hilterhaus, Lutz; Hinz, Andreas; Tantimongcolwat, Tanawut; Galla, Hans-Joachim

2005-02-04

Non-specific adsorption and specific interaction between a chimeric green fluorescent protein (GFP) carrying metal-binding region and the immobilized zinc ions on artificial solid-supported lipid membranes was investigated using the quartz crystal microbalance technique and the atomic force microscopy (AFM). Supported lipid bilayer, composed of octanethiol and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1,2-dioleoyl-sn-glycero-3-[N-(5-amino-1-carboxypentyl iminodiacetic acid)succinyl] (NTA-DOGS)-Zn2+, was formed on the gold electrode of quartz resonator (5 MHz). Binding of the chimeric GFP to zinc ions resulted in a rapid decrease of resonance frequency. Reversibility of the process was demonstrated via the removal of metal ions by EDTA. Nanoscale structural orientation of the chimeric GFP on the membrane was imaged by AFM. Association constant of the specific binding to metal ions was 2- to 3-fold higher than that of the non-specific adsorption, which was caused by the fluidization effect of the metal-chelating lipid molecules as well as the steric hindrance effect. This infers a possibility for a further development of biofunctionalized membrane. However, maximization is needed in order to attain closer advancement to a membrane-based sensor device.

Biophysical costs associated with tetrodotoxin resistance in the sodium channel pore of the garter snake, Thamnophis sirtalis.

PubMed

Lee, Chong Hyun; Jones, David K; Ahern, Christopher; Sarhan, Maen F; Ruben, Peter C

2011-01-01

Tetrodotoxin (TTX) is a potent toxin that specifically binds to voltage-gated sodium channels (NaV). TTX binding physically blocks the flow of sodium ions through NaV, thereby preventing action potential generation and propagation. TTX has different binding affinities for different NaV isoforms. These differences are imparted by amino acid substitutions in positions within, or proximal to, the TTX-binding site in the channel pore. These substitutions confer TTX-resistance to a variety of species. The garter snake Thamnophis sirtalis has evolved TTX-resistance over the course of an arms race, allowing some populations of snakes to feed on tetrodotoxic newts, including Taricha granulosa. Different populations of the garter snake have different degrees of TTX-resistance, which is closely related to the number of amino acid substitutions. We tested the biophysical properties and ion selectivity of NaV of three garter snake populations from Bear Lake, Idaho; Warrenton, Oregon; and Willow Creek, California. We observed changes in gating properties of TTX-resistant (TTXr) NaV. In addition, ion selectivity of TTXr NaV was significantly different from that of TTX-sensitive NaV. These results suggest TTX-resistance comes at a cost to performance caused by changes in the biophysical properties and ion selectivity of TTXr NaV.

Discovery of the ammonium substrate site on glutamine synthetase, a third cation binding site.

PubMed Central

Liaw, S. H.; Kuo, I.; Eisenberg, D.

1995-01-01

Glutamine synthetase (GS) catalyzes the ATP-dependent condensation of ammonia and glutamate to yield glutamine, ADP, and inorganic phosphate in the presence of divalent cations. Bacterial GS is an enzyme of 12 identical subunits, arranged in two rings of 6, with the active site between each pair of subunits in a ring. In earlier work, we have reported the locations within the funnel-shaped active site of the substrates glutamate and ATP and of the two divalent cations, but the site for ammonia (or ammonium) has remained elusive. Here we report the discovery by X-ray crystallography of a binding site on GS for monovalent cations, Tl+ and Cs+, which is probably the binding site for the substrate ammonium ion. Fourier difference maps show the following. (1) Tl+ and Cs+ bind at essentially the same site, with ligands being Glu 212, Tyr 179, Asp 50', Ser 53' of the adjacent subunit, and the substrate glutamate. From its position adjacent to the substrate glutamate and the cofactor ADP, we propose that this monovalent cation site is the substrate ammonium ion binding site. This proposal is supported by enzyme kinetics. Our kinetic measurements show that Tl+, Cs+, and NH4+ are competitive inhibitors to NH2OH in the gamma-glutamyl transfer reaction. (2) GS is a trimetallic enzyme containing two divalent cation sites (n1, n2) and one monovalent cation site per subunit. These three closely spaced ions are all at the active site: the distance between n1 and n2 is 6 A, between n1 and Tl+ is 4 A, and between n2 and Tl+ is 7 A. Glu 212 and the substrate glutamate are bridging ligands for the n1 ion and Tl+. (3) The presence of a monovalent cation in this site may enhance the structural stability of GS, because of its effect of balancing the negative charges of the substrate glutamate and its ligands and because of strengthening the "side-to-side" intersubunit interaction through the cation-protein bonding. (4) The presence of the cofactor ADP increases the Tl+ binding to GS because ADP binding induces movement of Asp 50' toward this monovalent cation site, essentially forming the site. This observation supports a two-step mechanism with ordered substrate binding: ATP first binds to GS, then Glu binds and attacks ATP to form gamma-glutamyl phosphate and ADP, which complete the ammonium binding site. The third substrate, an ammonium ion, then binds to GS, and then loses a proton to form the more active species ammonia, which attacks the gamma-glutamyl phosphate to yield Gln. (5) Because the products (Glu or Gln) of the reactions catalyzed by GS are determined by the molecule (water or ammonium) attacking the intermediate gamma-glutamyl phosphate, this negatively charged ammonium binding pocket has been designed naturally for high affinity of ammonium to GS, permitting glutamine synthesis to proceed in aqueous solution. PMID:8563633

Solving the Mechanism of Na+/H+ Antiporters Using Molecular Dynamics Simulations

NASA Astrophysics Data System (ADS)

Dotson, David L.

Na+/H+ antiporters are vital membrane proteins for cell homeostasis, transporting Na+ ions in exchange for H+ across the lipid bilayer. In humans, dysfunction of these transporters are implicated in hypertension, heart failure, epilepsy, and autism, making them well-established drug targets. Although experimental structures for bacterial homologs of the human Na+/H+ have been obtained, the detailed mechanism for ion transport is still not well-understood. The most well-studied of these transporters, Escherichia coli NhaA, known to transport 2 H+ for every Na+ extruded, was recently shown to bind H+ and Na+ at the same binding site, for which the two ion species compete. Using molecular dynamics simulations, the work presented in this dissertation shows that Na+ binding disrupts a previously-unidentified salt bridge between two conserved residues, suggesting that one of these residues, Lys300, may participate directly in transport of H+. This work also demonstrates that the conformational change required for ion translocation in a homolog of NhaA, Thermus thermophilus NapA, thought by some to involve only small helical movements at the ion binding site, is a large-scale, rigid-body movement of the core domain relative to the dimerization domain. This elevator-like transport mechanism translates a bound Na+ up to 10 A across the membrane. These findings constitute a major shift in the prevailing thought on the mechanism of these transporters, and serve as an exciting launchpad for new developments toward understanding that mechanism in detail.

Positronium ions and molecules

NASA Technical Reports Server (NTRS)

Ho, Y. K.

1990-01-01

Recent theoretical studies on positronium ions and molecules are discussed. A positronium ion is a three particle system consisting of two electrons in singlet spin state, and a positron. Recent studies include calculations of its binding energy, positron annihilation rate, and investigations of its doubly excited resonant states. A positronium molecule is a four body system consisting of two positrons and two electrons in an overall singlet spin state. The recent calculations of its binding energy against the dissociation into two positronium atoms, and studies of auto-detaching states in positronium molecules are discussed. These auto-dissociating states, which are believed to be part of the Rydberg series as a result of a positron attaching to a negatively charged positronium ion, Ps-, would appear as resonances in Ps-Ps scattering.

The binding energies of one and two water molecules to the first transition-row metal positive ions

NASA Technical Reports Server (NTRS)

Rosi, Marzio; Bauschlicher, Charles W., Jr.

1989-01-01

The bonding of water to the transition metal positive ions is electrostatic in origin. The electrostatic bonding is enhanced by a variety of mechanisms: mixing in 4p character, 4s-3d hybridization, and 4s promotion into the compact 3d orbital. The importance of these effects varies between the different metal ions due to changes in the separation of the metal ion atomic states. Furthermore, the change in the metal-water repulsion when a second water is added also changes the relative importance of the different metal asymptotes. The second water binding energy varies from being 11 kcal/mol smaller than the first for Mn(+) to 3 kcal/mol larger for V(+) and Fe(+).

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

Hacking RNA: Hakai promotes tumorigenesis by switching on the RNA-binding function of PSF

PubMed Central

Figueroa, Angélica; Fujita, Yasuyuki; Gorospe, Myriam

2009-01-01

Hakai, an E3 ubiquitin ligase for the E-cadherin complex, plays a crucial role in lowering cell-cell contacts in epithelial cells, a hallmark feature of tumor progression. Recently, Hakai was also found to interact with PSF (PTB-associated splicing factor). While PSF can function as a DNA-binding protein with a tumor suppressive function, its association with Hakai promotes PSF’s RNA-binding ability and post-transcriptional influence on target mRNAs. Hakai overexpression enhanced the binding of PSF to mRNAs encoding cancer-related proteins, while knockdown of Hakai reduced the RNA-binding ability of PSF. Furthermore, the knockdown of PSF suppressed Hakai-induced cell proliferation. Thus, Hakai can affect the oncogenic phenotype both by altering E-cadherin-based intercellular adhesions and by increasing PSF’s ability to bind RNAs that promote cancer-related gene expression. PMID:19855157

Molecular dynamics simulations of apocupredoxins: insights into the formation and stabilization of copper sites under entatic control.

PubMed

Abriata, Luciano A; Vila, Alejandro J; Dal Peraro, Matteo

2014-06-01

Cupredoxins perform copper-mediated long-range electron transfer (ET) in biological systems. Their copper-binding sites have evolved to force copper ions into ET-competent systems with decreased reorganization energy, increased reduction potential, and a distinct electronic structure compared with those of non-ET-competent copper complexes. The entatic or rack-induced state hypothesis explains these special properties in terms of the strain that the protein matrix exerts on the metal ions. This idea is supported by X-ray structures of apocupredoxins displaying "closed" arrangements of the copper ligands like those observed in the holoproteins; however, it implies completely buried copper-binding atoms, conflicting with the notion that they must be exposed for copper loading. On the other hand, a recent work based on NMR showed that the copper-binding regions of apocupredoxins are flexible in solution. We have explored five cupredoxins in their "closed" apo forms through molecular dynamics simulations. We observed that prearranged ligand conformations are not stable as the X-ray data suggest, although they do form part of the dynamic landscape of the apoproteins. This translates into variable flexibility of the copper-binding regions within a rigid fold, accompanied by fluctuations of the hydrogen bonds around the copper ligands. Major conformations with solvent-exposed copper-binding atoms could allow initial binding of the copper ions. An eventual subsequent incursion to the closed state would result in binding of the remaining ligands, trapping the closed conformation thanks to the additional binding energy and the fastening of noncovalent interactions that make up the rack.

Rapid kinetics of iron responsive element (IRE) RNA/iron regulatory protein 1 and IRE-RNA/eIF4F complexes respond differently to metal ions.

PubMed

Khan, Mateen A; Ma, Jia; Walden, William E; Merrick, William C; Theil, Elizabeth C; Goss, Dixie J

2014-06-01

Metal ion binding was previously shown to destabilize IRE-RNA/IRP1 equilibria and enhanced IRE-RNA/eIF4F equilibria. In order to understand the relative importance of kinetics and stability, we now report rapid rates of protein/RNA complex assembly and dissociation for two IRE-RNAs with IRP1, and quantitatively different metal ion response kinetics that coincide with the different iron responses in vivo. kon, for FRT IRE-RNA binding to IRP1 was eight times faster than ACO2 IRE-RNA. Mn(2+) decreased kon and increased koff for IRP1 binding to both FRT and ACO2 IRE-RNA, with a larger effect for FRT IRE-RNA. In order to further understand IRE-mRNA regulation in terms of kinetics and stability, eIF4F kinetics with FRT IRE-RNA were determined. kon for eIF4F binding to FRT IRE-RNA in the absence of metal ions was 5-times slower than the IRP1 binding to FRT IRE-RNA. Mn(2+) increased the association rate for eIF4F binding to FRT IRE-RNA, so that at 50 µM Mn(2+) eIF4F bound more than 3-times faster than IRP1. IRP1/IRE-RNA complex has a much shorter life-time than the eIF4F/IRE-RNA complex, which suggests that both rate of assembly and stability of the complexes are important, and that allows this regulatory system to respond rapidly to change in cellular iron. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

Stoichiometry and kinetics of mercury uptake by photosynthetic bacteria.

PubMed

Kis, Mariann; Sipka, Gábor; Maróti, Péter

2017-05-01

Mercury adsorption on the cell surface and intracellular uptake by bacteria represent the key first step in the production and accumulation of highly toxic mercury in living organisms. In this work, the biophysical characteristics of mercury bioaccumulation are studied in intact cells of photosynthetic bacteria by use of analytical (dithizone) assay and physiological photosynthetic markers (pigment content, fluorescence induction, and membrane potential) to determine the amount of mercury ions bound to the cell surface and taken up by the cell. It is shown that the Hg(II) uptake mechanism (1) has two kinetically distinguishable components, (2) includes co-opted influx through heavy metal transporters since the slow component is inhibited by Ca 2+ channel blockers, (3) shows complex pH dependence demonstrating the competition of ligand binding of Hg(II) ions with H + ions (low pH) and high tendency of complex formation of Hg(II) with hydroxyl ions (high pH), and (4) is not a passive but an energy-dependent process as evidenced by light activation and inhibition by protonophore. Photosynthetic bacteria can accumulate Hg(II) in amounts much (about 10 5 ) greater than their own masses by well-defined strong and weak binding sites with equilibrium binding constants in the range of 1 (μM) -1 and 1 (mM) -1 , respectively. The strong binding sites are attributed to sulfhydryl groups as the uptake is blocked by use of sulfhydryl modifying agents and their number is much (two orders of magnitude) smaller than the number of weak binding sites. Biofilms developed by some bacteria (e.g., Rvx. gelatinosus) increase the mercury binding capacity further by a factor of about five. Photosynthetic bacteria in the light act as a sponge of Hg(II) and can be potentially used for biomonitoring and bioremediation of mercury-contaminated aqueous cultures.

Interaction of ibogaine with human α3β4-nicotinic acetylcholine receptors in different conformational states

PubMed Central

Arias, Hugo R.; Rosenberg, Avraham; Targowska-Duda, Katarzyna M.; Feuerbach, Dominik; Yuan, Xiao Juan; Jozwiak, Krzysztof; Moaddel, Ruin; Wainer, Irving W.

2015-01-01

The interaction of ibogaine and phencyclidine (PCP) with human (h) α3β4-nicotinic acetylcholine receptors (AChRs) in different conformational states was determined by functional and structural approaches including, radioligand binding assays, Ca2+ influx detections, and thermodynamic and kinetics measurements. The results established that (a) ibogaine inhibits (±)-epibatidine-induced Ca2+ influx in hα3β4 AChRs with ~9-fold higher potency than that for PCP, (b) [3H]ibogaine binds to a single site in the hα3β4 AChR ion channel with relatively high affinity (Kd = 0.46 ± 0.06 µM), and ibogaine inhibits [3H]ibogaine binding to the desensitized hα3β4 AChR with slightly higher affinity compared to the resting AChR. This is explained by a slower dissociation rate from the desensitized ion channel compared to the resting ion channel, and (c) PCP inhibits [3H]ibogaine binding to the hα3β4 AChR, suggesting overlapping sites. The experimental results correlate with the docking simulations suggesting that ibogaine and PCP interact with a binding domain located between the serine (position 6′) and valine/phenylalanine (position 13′) rings. This interaction is mediated mainly by van der Waals contacts, which is in agreement with the observed enthalpic contribution determined by non-linear chromatography. However, the calculated entropic contribution also indicates local conformational changes. Collectively our data suggest that ibogaine and PCP bind to overlapping sites located between the serine and valine/phenylalanine rings, to finally block the AChR ion channel, and in the case of ibogaine, to probably maintain the AChR in the desensitized state for longer time. PMID:20684041

Interaction of ibogaine with human alpha3beta4-nicotinic acetylcholine receptors in different conformational states.

PubMed

Arias, Hugo R; Rosenberg, Avraham; Targowska-Duda, Katarzyna M; Feuerbach, Dominik; Yuan, Xiao Juan; Jozwiak, Krzysztof; Moaddel, Ruin; Wainer, Irving W

2010-09-01

The interaction of ibogaine and phencyclidine (PCP) with human (h) alpha3beta4-nicotinic acetylcholine receptors (AChRs) in different conformational states was determined by functional and structural approaches including, radioligand binding assays, Ca2+ influx detections, and thermodynamic and kinetics measurements. The results established that (a) ibogaine inhibits (+/-)-epibatidine-induced Ca2+ influx in h(alpha)3beta4 AChRs with approximately 9-fold higher potency than that for PCP, (b) [3H]ibogaine binds to a single site in the h(alpha)3beta4 AChR ion channel with relatively high affinity (Kd = 0.46 +/- 0.06 microM), and ibogaine inhibits [3H]ibogaine binding to the desensitized h(alpha)3beta4 AChR with slightly higher affinity compared to the resting AChR. This is explained by a slower dissociation rate from the desensitized ion channel compared to the resting ion channel, and (c) PCP inhibits [3H]ibogaine binding to the h(alpha)3beta4 AChR, suggesting overlapping sites. The experimental results correlate with the docking simulations suggesting that ibogaine and PCP interact with a binding domain located between the serine (position 6') and valine/phenylalanine (position 13') rings. This interaction is mediated mainly by van der Waals contacts, which is in agreement with the observed enthalpic contribution determined by non-linear chromatography. However, the calculated entropic contribution also indicates local conformational changes. Collectively our data suggest that ibogaine and PCP bind to overlapping sites located between the serine and valine/phenylalanine rings, to finally block the AChR ion channel, and in the case of ibogaine, to probably maintain the AChR in the desensitized state for longer time.

Structural characterization of agonist and antagonist-bound acetylcholine-binding protein from Aplysia californica.

PubMed

Hansen, Scott B; Sulzenbacher, Gerlind; Huxford, Tom; Marchot, Pascale; Bourne, Yves; Taylor, Palmer

2006-01-01

Nicotinic acetylcholine receptors (nAChRs) are well-characterized allosteric transmembrane proteins involved in the rapid gating of ions elicited by ACh. These receptors belong to the Cys-loop superfamily of ligand-gated ion channels, which also includes GABAA and GABAC, 5-HT3, and glycine receptors. The nAChRs are homo- or heteromeric pentamers of structurally related subunits that encompass an extracellular N-terminal ligand-binding domain, four transmembrane-spanning regions that form the ion channel, and an extended intracellular region between spans 3 and 4. Ligand binding triggers conformational changes that are transmitted to the transmembrane-spanning region, leading to gating and changes in membrane potential. The four transmembrane spans on each of the five subunits create a substantial region of hydrophobicity that precludes facile crystallization of this protein. However the freshwater snail, Lymnaea stagnalis, produces a soluble homopentameric protein, termed the ACh-binding protein (AChBP), which binds ACh (Smit et al., 2001). Its structure was determined recently (Brejc et al., 2001) at high resolution, revealing the structural scaffold for nAChR, and has become a functional and structural surrogate of the nAChR ligand-binding domain. We have characterized an AChBP from Aplysia californica and determined distinct ligand-binding properties when compared to those of L. stagnalis, including ligand specificity for the nAChR alpha7 subtype-specific alpha-conotoxin ImI (Hansen et al., 2004).

Synthesis, crystal structure and investigation of mononuclear copper(II) and zinc(II) complexes of a new carboxylate rich tripodal ligand and their interaction with carbohydrates in alkaline aqueous solution

PubMed Central

Stewart, Christopher D.; Pedraza, Mayra; Arman, Hadi; Fan, Hua-Jun; Schilling, Eduardo Luiz; Szpoganicz, Bruno; Musie, Ghezai T.

2016-01-01

A new carboxylate rich asymmetric tripodal ligand, N-[2-carboxybenzomethyl]-N-[carboxymethyl]-β-alanine (H3camb), and its di-copper(II), (NH4)2[1]2, and di-zinc(II), ((CH3)4 N)2[2]2, complexes have been synthesized as carbohydrate binding models in aqueous solutions. The ligand and complexes have been fully characterized using several techniques, including single crystal X-ray diffraction. The interactions of (NH4)2[1]2 and ((CH3)4 N)2[2]2 with D-glucose, D-mannose, D-xylose and xylitol in aqueous alkaline media were investigated using UV–Vis and 13C-NMR spectroscopic techniques, respectively. The molar conductance, NMR and ESI–MS studies indicate that the complexes dissociate in solution to produce the respective complex anions, 1− and 2−. Complexes 1− and 2− showed chelating ability towards the naturally abundant and biologically relevant sugars, D-glucose, D-mannose, D-xylose, and xylitol. The complex ions bind to one molar equivalent of the sugars, even in the presence of stoichiometric excess of the substrates, in solution. Experimentally obtained spectroscopic data and computational results suggest that the substrates bind to the metal center in a bidentate fashion. Apparent binding constant values, pKapp, between the complexes and the substrates were determined and a specific mode of substrate binding is proposed. The pKapp and relativistic density functional theory (DFT) calculated Gibbs free energy values indicate that D-mannose displayed the strongest interaction with the complexes. Syntheses, characterizations, detailed substrate binding studies using spectroscopic techniques, single crystal X-ray diffraction and geometry optimizations of the complex-substrates with DFT calculations are also reported. PMID:25969174

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.

Expression and purification of recombinant proteins in Escherichia coli tagged with the metal-binding protein CusF.

PubMed

Cantu-Bustos, J Enrique; Vargas-Cortez, Teresa; Morones-Ramirez, Jose Ruben; Balderas-Renteria, Isaias; Galbraith, David W; McEvoy, Megan M; Zarate, Xristo

2016-05-01

Production of recombinant proteins in Escherichia coli has been improved considerably through the use of fusion proteins, because they increase protein solubility and facilitate purification via affinity chromatography. In this article, we propose the use of CusF as a new fusion partner for expression and purification of recombinant proteins in E. coli. Using a cell-free protein expression system, based on the E. coli S30 extract, Green Fluorescent Protein (GFP) was expressed with a series of different N-terminal tags, immobilized on self-assembled protein microarrays, and its fluorescence quantified. GFP tagged with CusF showed the highest fluorescence intensity, and this was greater than the intensities from corresponding GFP constructs that contained MBP or GST tags. Analysis of protein production in vivo showed that CusF produces large amounts of soluble protein with low levels of inclusion bodies. Furthermore, fusion proteins can be exported to the cellular periplasm, if CusF contains the signal sequence. Taking advantage of its ability to bind copper ions, recombinant proteins can be purified with readily available IMAC resins charged with this metal ion, producing pure proteins after purification and tag removal. We therefore recommend the use of CusF as a viable alternative to MBP or GST as a fusion protein/affinity tag for the production of soluble recombinant proteins in E. coli. Copyright © 2016 Elsevier Inc. All rights reserved.

Assigning crystallographic electron densities with free energy calculations—The case of the fluoride channel Fluc

PubMed Central

2018-01-01

Approximately 90% of the structures in the Protein Data Bank (PDB) were obtained by X-ray crystallography or electron microscopy. Whereas the overall quality of structure is considered high, thanks to a wide range of tools for structure validation, uncertainties may arise from density maps of small molecules, such as organic ligands, ions or water, which are non-covalently bound to the biomolecules. Even with some experience and chemical intuition, the assignment of such disconnected electron densities is often far from obvious. In this study, we suggest the use of molecular dynamics (MD) simulations and free energy calculations, which are well-established computational methods, to aid in the assignment of ambiguous disconnected electron densities. Specifically, estimates of (i) relative binding affinities, for instance between an ion and water, (ii) absolute binding free energies, i.e., free energies for transferring a solute from bulk solvent to a binding site, and (iii) stability assessments during equilibrium simulations may reveal the most plausible assignments. We illustrate this strategy using the crystal structure of the fluoride specific channel (Fluc), which contains five disconnected electron densities previously interpreted as four fluoride and one sodium ion. The simulations support the assignment of the sodium ion. In contrast, calculations of relative and absolute binding free energies as well as stability assessments during free MD simulations suggest that four of the densities represent water molecules instead of fluoride. The assignment of water is compatible with the loss of these densities in the non-conductive F82I/F85I mutant of Fluc. We critically discuss the role of the ion force fields for the calculations presented here. Overall, these findings indicate that MD simulations and free energy calculations are helpful tools for modeling water and ions into crystallographic density maps. PMID:29771936

Hydrogen peroxide-mediated oxidative stress disrupts calcium binding on calmodulin: More evidence for oxidative stress in vitiligo

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

Schallreuter, K.U.; Gibbons, N.C.J.; Zothner, C.

Patients with acute vitiligo have low epidermal catalase expression/activities and accumulate 10{sup -3} M H{sub 2}O{sub 2}. One consequence of this severe oxidative stress is an altered calcium homeostasis in epidermal keratinocytes and melanocytes. Here, we show decreased epidermal calmodulin expression in acute vitiligo. Since 10{sup -3}M H{sub 2}O{sub 2} oxidises methionine and tryptophan residues in proteins, we examined calcium binding to calmodulin in the presence and absence of H{sub 2}O{sub 2} utilising {sup 45}calcium. The results showed that all four calcium atoms exchanged per molecule of calmodulin. Since oxidised calmodulin looses its ability to activate calcium ATPase, enzyme activitiesmore » were followed in full skin biopsies from lesional skin of patients with acute vitiligo (n = 6) and healthy controls (n = 6). The results yielded a 4-fold decrease of ATPase activities in the patients. Computer simulation of native and oxidised calmodulin confirmed the loss of all four calcium ions from their specific EF-hand domains. Taken together H{sub 2}O{sub 2}-mediated oxidation affects calcium binding in calmodulin leading to perturbed calcium homeostasis and perturbed L-phenylalanine-uptake in the epidermis of acute vitiligo.« less

Synthesis and biological evaluation of Santacruzamate-A based analogues.

PubMed

Randino, Rosario; Gazzerro, Patrizia; Mazitschek, Ralph; Rodriquez, Manuela

2017-12-15

Several derivatives of Santacruzamate-A, a natural product that is structurally related to SAHA, were synthesized to explore the potential of carbamates and oxalylamides as novel biasing element for targeting the catalytic site of zinc-dependent histone deacetylases (HDACs). An additional class of Santacruzamate-A derivatives was synthesized to investigate the influence of the cap group and the linker element on HDAC inhibitory activity. All compounds were evaluated in dose response for their in vitro cytotoxic activity in MTT assay in HCT116 cells. HDAC inhibitory activity was evaluated in vitro by western blot analysis for histone hyperacetylation assay and biochemically for representative human HDACs isoforms. Two novel compounds were identified to exhibit potent time dependent anti proliferative activity. However, unlike hydroxamic acid analogues, the tested Santacruzamate-A derivatives showed no noticeable HDAC inhibitory activity. The ethylcarbamate moiety as unusual zinc-binding group displayed no ability to coordinate the zinc ion and thus, presumably, was not able to reproduce known inhibitor-substrate zinc-binding group interactions with the HDAC catalytic site. This study confirmed that the accommodation of the zinc-binding group is deeply critical of the positioning of the linker and the projection of the cap group toward the different surface pockets of the enzyme. Copyright © 2017 Elsevier Ltd. All rights reserved.

Computational Design of Metal Ion Sequestering Agents

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

Hay, Benjamin P.; Rapko, Brian M.

Organic ligands that exhibit a high degree of metal ion recognition are essential precursors for developing separation processes and sensors for metal ions. Since the beginning of the nuclear era, much research has focused on discovering ligands that target specific radionuclides. Members of the Group 1A and 2A cations (e.g., Cs, Sr, Ra) and the f-block metals (actinides and lanthanides) are of primary concern to DOE. Although there has been some success in identifying ligand architectures that exhibit a degree of metal ion recognition, the ability to control binding affinity and selectivity remains a significant challenge. The traditional approach formore » discovering such ligands has involved lengthy programs of organic synthesis and testing that, in the absence of reliable methods for screening compounds before synthesis, have resulted in much wasted research effort. This project seeks to enhance and strengthen the traditional approach through computer-aided design of new and improved host molecules. Accurate electronic structure calculations are coupled with experimental data to provide fundamental information about ligand structure and the nature of metal-donor group interactions (design criteria). This fundamental information then is used in a molecular mechanics model (MM) that helps us rapidly screen proposed ligand architectures and select the best members from a set of potential candidates. By using combinatorial methods, molecule building software has been developed that generates large numbers of candidate architectures for a given set of donor groups. The specific goals of this project are: • further understand the structural and energetic aspects of individual donor group- metal ion interactions and incorporate this information within the MM framework • further develop and evaluate approaches for correlating ligand structure with reactivity toward metal ions, in other words, screening capability • use molecule structure building software to generate large numbers of candidate ligand architectures for given sets of donor groups • screen candidates and identify ligand architectures that will exhibit enhanced metal ion recognition. These new capabilities are being applied to ligand systems identified under other DOEsponsored projects where studies have suggested that modifying existing architectures will lead to dramatic enhancements in metal ion binding affinity and selectivity. With this in mind, we are collaborating with Professors R. T. Paine (University of New Mexico), K. N. Raymond (University of California, Berkeley), and J. E. Hutchison (University of Oregon), and Dr. B. A. Moyer (Oak Ridge National Laboratory) to obtain experimental validation of the predicted new ligand structures. Successful completion of this study will yield molecular-level insight into the role that ligand architecture plays in controlling metal ion complexation and will provide a computational approach to ligand design.« less

Structural insights into the T6SS effector protein Tse3 and the Tse3-Tsi3 complex from Pseudomonas aeruginosa reveal a calcium-dependent membrane-binding mechanism.

PubMed

Lu, Defen; Shang, Guijun; Zhang, Heqiao; Yu, Qian; Cong, Xiaoyan; Yuan, Jupeng; He, Fengjuan; Zhu, Chunyuan; Zhao, Yanyu; Yin, Kun; Chen, Yuanyuan; Hu, Junqiang; Zhang, Xiaodan; Yuan, Zenglin; Xu, Sujuan; Hu, Wei; Cang, Huaixing; Gu, Lichuan

2014-06-01

The opportunistic pathogen Pseudomonas aeruginosa uses the type VI secretion system (T6SS) to deliver the muramidase Tse3 into the periplasm of rival bacteria to degrade their peptidoglycan (PG). Concomitantly, P. aeruginosa uses the periplasm-localized immunity protein Tsi3 to prevent potential self-intoxication caused by Tse3, and thus gains an edge over rival bacteria in fierce niche competition. Here, we report the crystal structures of Tse3 and the Tse3-Tsi3 complex. Tse3 contains an annexin repeat-like fold at the N-terminus and a G-type lysozyme fold at the C-terminus. One loop in the N-terminal domain (Loop 12) and one helix (α9) from the C-terminal domain together anchor Tse3 and the Tse3-Tsi3 complex to membrane in a calcium-dependent manner in vitro, and this membrane-binding ability is essential for Tse3's activity. In the C-terminal domain, a Y-shaped groove present on the surface likely serves as the PG binding site. Two calcium-binding motifs are also observed in the groove and these are necessary for Tse3 activity. In the Tse3-Tsi3 structure, three loops of Tsi3 insert into the substrate-binding groove of Tse3, and three calcium ions present at the interface of the complex are indispensable for the formation of the Tse3-Tsi3 complex. © 2014 John Wiley & Sons Ltd.

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

Soil-modified carbon paste electrode: a useful tool in environmental assessment of heavy metal ion binding interactions.

PubMed

Svegl, I G; Ogorevc, B

2000-08-01

Carbon paste electrodes (CPEs) modified with different soils in their native form were prepared to create a soil-like solid phase suitable for application in studies of heavy metal ion uptake and binding interactions. The preparation of CPEs modified with five different soils was examined and their heavy metal ion uptake behavior investigated using a model Cu(II) aqueous solution. Metal ions were accumulated under open circuit conditions and were determined after a medium exchange using differential pulse anodic stripping voltammetry, applying preelectrolysis at -0.7 V. The soil-modified CPE accumulation behavior, including the linearity of the current response versus Cu(II) concentration, the influence of the pH on the solution, and the uptake kinetics, was thoroughly investigated. The correlation between the soil-modified CPE uptake capability and the standard soil parameters, such as ion exchange capacity, soil pH, organic matter and clay content, were evaluated for all five examined soils. The influence of selected endogenous cations (K(I), Ca(II), Fe(III)) on the transfer of Cu(II) ions from a solution to the simulated soil solid phase was examined and is discussed. Preliminary examinations of the soil-modified CPE uptake behavior with some exogenous heavy metal ions of strong environmental interest (Pb(II), Hg(II), Cd(II) and Ag(I)) are also presented. This work demonstrates some attractive possibilities for the application of a soil-modified CPE in studying soil-heavy metal ion binding interactions, with a further potential use as a new environmental sensor appropriate for fist on-site testing of polluted soils.

Influence of natural organic matter source on copper speciation as demonstrated by Cu binding to fish gills, by ion selective electrode, and by DGT gel sampler

USGS Publications Warehouse

Luider, C.D.; Crusius, John; Playle, R.C.; Curtis, P.J.

2004-01-01

Rainbow trout (Oncorhynchus mykiss, 2 g) were exposed to 0−5 μM total copper in ion-poor water for 3 h in the presence or absence of 10 mg C/L of qualitatively different natural organic matter (NOM) derived from water spanning a large gradient in hydrologic residence time. Accumulation of Cu by trout gills was compared to Cu speciation determined by ion selective electrode (ISE) and by diffusive gradients in thin films (DGT) gel sampler technology. The presence of NOM decreased Cu uptake by trout gills as well as Cu concentrations determined by ISE and DGT. Furthermore, the source of NOM influenced Cu binding by trout gills with high-color, allochthonous NOM decreasing Cu accumulation by the gills more than low-color autochthonous NOM. The pattern of Cu binding to the NOM measured by Cu ISE and by Cu accumulation by DGT samplers was similar to the fish gill results. A simple Cu−gill binding model required an NOM Cu-binding factor (F) that depended on NOM quality to account for observed Cu accumulation by trout gills; values of F varied by a factor of 2. Thus, NOM metal-binding quality, as well as NOM quantity, are both important when assessing the bioavailability of metals such as Cu to aquatic organisms.

Two-Component System RgfA/C Activates the fbsB Gene Encoding Major Fibrinogen-Binding Protein in Highly Virulent CC17 Clone Group B Streptococcus

PubMed Central

Safadi, Rim Al; Mereghetti, Laurent; Salloum, Mazen; Lartigue, Marie-Frédérique; Virlogeux-Payant, Isabelle; Quentin, Roland; Rosenau, Agnès

2011-01-01

Group B streptococcus (GBS) strains with the highest ability to bind to human fibrinogen belong to the highly invasive clonal complex (CC) 17. To investigate the fibrinogen-binding mechanisms of CC17 strains, we determined the prevalence of fibrinogen-binding genes (fbsA and fbsB), and fbs regulator genes (rogB encoding an fbsA activator, rovS encoding an fbsA repressor and rgf encoding a two-component system [TCS] whose role on fbs genes was not determined yet) in a collection of 134 strains representing the major CCs of the species. We showed that specific gene combinations were related to particular CCs; only CC17 strains contained the fbsA, fbsB, and rgf genes combination. Non polar rgfAC deletion mutants of three CC17 serotype III strains were constructed. They showed a 3.2- to 5.1-fold increase of fbsA transcripts, a 4.8- to 6.7-fold decrease of fbsB transcripts, and a 52% to 68% decreased fibrinogen-binding ability, demonstrating that the RgfA/RgfC TCS inhibits the fbsA gene and activates the fbsB gene. The relative contribution of the two fbs genes in fibrinogen-binding ability was determined by constructing isogenic fbsA, fbsB, deletion mutants of the three CC17 strains. The ability to bind to fibrinogen was reduced by 49% to 57% in ΔfbsA mutants, and by 78% to 80% in ΔfbsB mutants, suggesting that FbsB protein plays a greater role in the fibrinogen-binding ability of CC17 strains. Moreover, the relative transcription level of fbsB gene was 9.2- to 12.7-fold higher than that of fbsA gene for the three wild type strains. Fibrinogen-binding ability could be restored by plasmid-mediated expression of rgfAC, fbsA, and fbsB genes in the corresponding deletion mutants. Thus, our results demonstrate that a specific combination of fbs genes and fbs regulator genes account for the high fibrinogen-binding ability of CC17 strains that may participate to their enhanced invasiveness for neonates as compared to strains of other CCs. PMID:21326613

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.

Anions mediate ligand binding in Adineta vaga glutamate receptor ion channels.

PubMed

Lomash, Suvendu; Chittori, Sagar; Brown, Patrick; Mayer, Mark L

2013-03-05

AvGluR1, a glutamate receptor ion channel from the primitive eukaryote Adineta vaga, is activated by alanine, cysteine, methionine, and phenylalanine, which produce lectin-sensitive desensitizing responses like those to glutamate, aspartate, and serine. AvGluR1 LBD crystal structures reveal an unusual scheme for binding dissimilar ligands that may be utilized by distantly related odorant/chemosensory receptors. Arginine residues in domain 2 coordinate the γ-carboxyl group of glutamate, whereas in the alanine, methionine, and serine complexes a chloride ion acts as a surrogate ligand, replacing the γ-carboxyl group. Removal of Cl(-) lowers affinity for these ligands but not for glutamate or aspartate nor for phenylalanine, which occludes the anion binding site and binds with low affinity. AvGluR1 LBD crystal structures and sedimentation analysis also provide insights into the evolutionary link between prokaryotic and eukaryotic iGluRs and reveal features unique to both classes, emphasizing the need for additional structure-based studies on iGluR-ligand interactions. Copyright © 2013 Elsevier Ltd. All rights reserved.

Antagonism of ligand-gated ion channel receptors: two domains of the glycine receptor alpha subunit form the strychnine-binding site.

PubMed Central

Vandenberg, R J; French, C R; Barry, P H; Shine, J; Schofield, P R

1992-01-01

The inhibitory glycine receptor (GlyR) is a member of the ligand-gated ion channel receptor superfamily. Glycine activation of the receptor is antagonized by the convulsant alkaloid strychnine. Using in vitro mutagenesis and functional analysis of the cDNA encoding the alpha 1 subunit of the human GlyR, we have identified several amino acid residues that form the strychnine-binding site. These residues were identified by transient expression of mutated cDNAs in mammalian (293) cells and examination of resultant [3H]strychnine binding, glycine displacement of [3H]strychnine, and electrophysiological responses to the application of glycine and strychnine. This mutational analysis revealed that residues from two separate domains within the alpha 1 subunit form the binding site for the antagonist strychnine. The first domain includes the amino acid residues Gly-160 and Tyr-161, and the second domain includes the residues Lys-200 and Tyr-202. These results, combined with analyses of other ligand-gated ion channel receptors, suggest a conserved tertiary structure and a common mechanism for antagonism in this receptor superfamily. PMID:1311851

An external sodium ion binding site controls allosteric gating in TRPV1 channels

PubMed Central

Jara-Oseguera, Andres; Bae, Chanhyung; Swartz, Kenton J

2016-01-01

TRPV1 channels in sensory neurons are integrators of painful stimuli and heat, yet how they integrate diverse stimuli and sense temperature remains elusive. Here, we show that external sodium ions stabilize the TRPV1 channel in a closed state, such that removing the external ion leads to channel activation. In studying the underlying mechanism, we find that the temperature sensors in TRPV1 activate in two steps to favor opening, and that the binding of sodium to an extracellular site exerts allosteric control over temperature-sensor activation and opening of the pore. The binding of a tarantula toxin to the external pore also exerts control over temperature-sensor activation, whereas binding of vanilloids influences temperature-sensitivity by largely affecting the open/closed equilibrium. Our results reveal a fundamental role of the external pore in the allosteric control of TRPV1 channel gating and provide essential constraints for understanding how these channels can be tuned by diverse stimuli. DOI: http://dx.doi.org/10.7554/eLife.13356.001 PMID:26882503

Streptomyces coelicolor SCO4226 Is a Nickel Binding Protein

PubMed Central

Jin, Hua; Zhang, Rong-Guang; Virolle, Marie-Joelle; Chen, Yuxing; Zhou, Cong-Zhao

2014-01-01

The open reading frame SCO4226 of Streptomyces coelicolor A3(2) encodes an 82-residue hypothetical protein. Biochemical assays revealed that each SCO4226 dimer binds four nickel ions. To decipher the molecular function, we solved the crystal structures of SCO4226 in both apo- and nickel-bound (Ni-SCO4226) forms at 1.30 and 2.04 Å resolution, respectively. Each subunit of SCO4226 dimer adopts a canonical ferredoxin-like fold with five β-strands flanked by two α-helices. In the structure of Ni-SCO4226, four nickel ions are coordinated at the surface of the dimer. Further biochemical assays suggested that the binding of Ni2+ triggers the self-aggregation of SCO4226 in vitro. In addition, RT-qPCR assays demonstrated that the expression of SCO4226 gene in S. coelicolor is specifically up-regulated by the addition of Ni2+, but not other divalent ions such as Cu2+, Mn2+ or Co2+. All these results suggested that SCO4226 acts as a nickel binding protein, probably required for nickel sequestration and/or detoxification. PMID:25285530

Raman microspectroscopic study of effects of Na(I) and Mg(II) ions on low pH induced DNA structural changes.

PubMed

Muntean, C M; Segers-Nolten, G M J

2003-01-01

In this work a confocal Raman microspectrometer is used to investigate the influence of Na(+) and Mg(2+) ions on the DNA structural changes induced by low pH. Measurements are carried out on calf thymus DNA at neutral pH (7) and pH 3 in the presence of low and high concentrations of Na(+) and Mg(2+) ions, respectively. It is found that low concentrations of Na(+) ions do not protect DNA against binding of H(+). High concentrations of monovalent ions can prevent protonation of the DNA double helix. Our Raman spectra show that low concentrations of Mg(2+) ions partly protect DNA against protonation of cytosine (line at 1262 cm(-1)) but do not protect adenine and guanine N(7) against binding of H(+) (characteristic lines at 1304 and 1488 cm(-1), respectively). High concentrations of Mg(2+) can prevent protonation of cytosine and protonation of adenine (disruption of AT pairs). By analyzing the line at 1488 cm(-1), which obtains most of its intensity from a guanine vibration, high magnesium salt protect the N(7) of guanine against protonation. A high salt concentration can prevent protonation of guanine, cytosine, and adenine in DNA. Higher salt concentrations cause less DNA protonation than lower salt concentrations. Magnesium ions are found to be more effective in protecting DNA against binding of H(+) as compared with calcium ions presented in a previous study. Divalent metal cations (Mg(2+), Ca(2+)) are more effective in protecting DNA against protonation than monovalent ions (Na(+)). Copyright 2003 Wiley Periodicals, Inc. Biopolymers (Biospectroscopy) 72: 000-000, 2003

Coarse-graining, Electrostatics and pH effects in phospholipid systems

NASA Astrophysics Data System (ADS)

Travesset, Alex; Vangaveti, Sweta

2010-03-01

We introduce a minimal free energy describing the interaction of charged groups and counterions including both classical electrostatic and specific interactions. The predictions of the model are compared against the standard model for describing ions next to charged interfaces, consisting of Poisson-Boltzmann theory with additional constants describing ion binding, which are specific to the counterion and the interfacial charge (``chemical binding''). It is shown that the ``chemical'' model can be appropriately described by an underlying ``physical'' model over several decades in concentration, but the extracted binding constants are not uniquely defined, as they differ depending on the particular observable quantity being studied. It is also shown that electrostatic correlations for divalent (or higher valence) ions enhance the surface charge by increasing deprotonation, an effect not properly accounted within chemical models. The model is applied to the charged phospholipids phosphatidylserine, Phosphatidc acid and Phosphoinositides and implications for different biological processes are discussed.

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

Covalent modifications of the amyloid beta peptide by hydroxynonenal: Effects on metal ion binding by monomers and insights into the fibril topology.

PubMed

Grasso, G; Komatsu, H; Axelsen, P H

2017-09-01

Amyloid β peptides (Aβ) and metal ions are associated with oxidative stress in Alzheimer's disease (AD). Oxidative stress, acting on ω-6 polyunsaturated fatty acyl chains, produces diverse products, including 4-hydroxy-2-nonenal (HNE), which can covalently modify the Aβ that helped to produce it. To examine possible feedback mechanisms involving Aβ, metal ions and HNE production, the effects of HNE modification and fibril formation on metal ion binding was investigated. Results indicate that copper(II) generally inhibits the modification of His side chains in Aβ by HNE, but that once modified, copper(II) still binds to Aβ with high affinity. Fibril formation protects only one of the three His residues in Aβ from HNE modification, and this protection is consistent with proposed models of fibril structure. These results provide insight into a network of biochemical reactions that may be operating as a consequence of oxidative stress in AD, or as part of the pathogenic process. Copyright © 2016. Published by Elsevier Inc.

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

Copper Coordination in the Full-Length, Recombinant Prion Protein†

PubMed Central

Burns, Colin S.; Aronoff-Spencer, Eliah; Legname, Giuseppe; Prusiner, Stanley B.; Antholine, William E.; Gerfen, Gary J.; Peisach, Jack; Millhauser, Glenn L.

2010-01-01

The prion protein (PrP) binds divalent copper at physiologically relevant conditions and is believed to participate in copper regulation or act as a copper-dependent enzyme. Ongoing studies aim at determining the molecular features of the copper binding sites. The emerging consensus is that most copper binds in the octarepeat domain, which is composed of four or more copies of the fundamental sequence PHGGGWGQ. Previous work from our laboratory using PrP-derived peptides, in conjunction with EPR and X-ray crystallography, demonstrated that the HGGGW segment constitutes the fundamental binding unit in the octarepeat domain [Burns et al. (2002) Biochemistry 41, 3991–4001; Aronoff-Spencer et al. (2000) Biochemistry 39, 13760–13771]. Copper coordination arises from the His imidazole and sequential deprotonated glycine amides. In this present work, recombinant, full-length Syrian hamster PrP is investigated using EPR methodologies. Four copper ions are taken up in the octarepeat domain, which supports previous findings. However, quantification studies reveal a fifth binding site in the flexible region between the octarepeats and the PrP globular C-terminal domain. A series of PrP peptide constructs show that this site involves His96 in the PrP(92–96) segment GGGTH. Further examination by X-band EPR, S-band EPR, and electron spin–echo envelope spectroscopy, demonstrates coordination by the His96 imidazole and the glycine preceding the threonine. The copper affinity for this type of binding site is highly pH dependent, and EPR studies here show that recombinant PrP loses its affinity for copper below pH 6.0. These studies seem to provide a complete profile of the copper binding sites in PrP and support the hypothesis that PrP function is related to its ability to bind copper in a pH-dependent fashion. PMID:12779334

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

Determination of geniposide in adjuvant arthritis rat plasma by ultra-high performance liquid chromatography tandem mass spectrometry method and its application to oral bioavailability and plasma protein binding ability studies.

PubMed

Chen, Jian; Wu, Hong; Xu, Guo-Bing; Dai, Miao-Miao; Hu, Shun-Li; Sun, Liang-Liang; Wang, Wei; Wang, Rong; Li, Shu-Pin; Li, Guo-Qiang

2015-04-10

A specific, sensitive and high throughput ultra-high performance liquid chromatography-electrospray ionization tandem mass spectrometric method (UHPLC-ESI-MS/MS) was established and validated to assay geniposide (GE), a promising anti-inflammatory drug, in adjuvant arthritis rat plasma: application to pharmacokinetic and oral bioavailability studies and plasma protein binding ability. Plasma samples were processed by de-proteinised with ice-cold methanol and separated on an ACQUITY UPLC™ HSS C18 column (100 mm × 2.1mm i.d., 1.8 μm particle size) at a gradient flow rate of 0.2 mL/min using acetonitrile-0.1% formic acid in water as mobile phase, and the total run time was 9 min. Mass detection was performed in selected reaction monitoring (SRM) mode with negative electro-spray ionization includes the addition of paeoniflorin (Pae) as an internal standard (IS). The mass transition ion-pair was followed as m/z 387.4 → 122.4 for GE and m/z 479.4 → 449.0 for IS. The calibration curves were linear over the concentration range of 2-50,000 ng/mL with lower limit of quantification of 2 ng/mL. The intra-day and inter-day precisions (RSD, %) of the assay were less than 8.4%, and the accuracy was within ± 6.4% in terms of relative error (RE). Extraction recovery, matrix effect and stability were satisfactory in adjuvant arthritis rat plasma. The UHPLC-ESI-MS/MS method was successfully applied to a pharmacokinetic study of GE after oral administration of depurated GE at 33, 66, 132 mg/kg and intravenous injection at 33, 66, 132 mg/kg in adjuvant arthritis (AA) rats. In addition, it was found that GE has rapid absorption and elimination, low absolute bioavailability, high plasma protein binding ability in AA rats after oral administration within the tested dosage range. It suggested that GE showed slow distribution into the intra- and extracellular space, and the binding rate was not proportionally dependent on plasma concentration of GE when the concentration of GE was below 5.0 μg/mL. Copyright © 2015 Elsevier B.V. All rights reserved.

Investigation of collision-induced dissociation products and structures of gas-phase [ M·GlyGlyHis-H]+ ( M = Fe, Ni, Cu, and Zn) complexes.

PubMed

Gannamani, Bharathi; Shin, Joong-Won

2017-02-01

Collision-induced dissociation is carried out for electrosprayed [Fe·GlyGlyHis-H] + , [Ni·GlyGlyHis-H] + , [Cu·GlyGlyHis-H] + , and [Zn·GlyGlyHis-H] + complexes. [Fe·GlyGlyHis-H] + , [Ni·GlyGlyHis-H] + , and [Zn·GlyGlyHis-H] + yield metal-bound peptide sequence ions and dehydrated ions as primary products, whereas [Cu·GlyGlyHis-H] + generates a more extensive series of metal-bound sequence ions and a product arising from the unusual loss of a formaldehyde moiety; dehydration is significantly suppressed for this complex. Density functional theory calculations show that the copper ion-deprotonated peptide binding energy is substantially higher than those in other complexes, suggesting that there is a correlation between ion-ligand binding energy and their fragmentation behavior.

Cation specific binding with protein surface charges

PubMed Central

Hess, Berk; van der Vegt, Nico F. A.

2009-01-01

Biological organization depends on a sensitive balance of noncovalent interactions, in particular also those involving interactions between ions. Ion-pairing is qualitatively described by the law of “matching water affinities.” This law predicts that cations and anions (with equal valence) form stable contact ion pairs if their sizes match. We show that this simple physical model fails to describe the interaction of cations with (molecular) anions of weak carboxylic acids, which are present on the surfaces of many intra- and extracellular proteins. We performed molecular simulations with quantitatively accurate models and observed that the order K+ < Na+ < Li+ of increasing binding affinity with carboxylate ions is caused by a stronger preference for forming weak solvent-shared ion pairs. The relative insignificance of contact pair interactions with protein surfaces indicates that thermodynamic stability and interactions between proteins in alkali salt solutions is governed by interactions mediated through hydration water molecules. PMID:19666545

Determinants of cation transport selectivity: Equilibrium binding and transport kinetics

PubMed Central

2015-01-01

The crystal structures of channels and transporters reveal the chemical nature of ion-binding sites and, thereby, constrain mechanistic models for their transport processes. However, these structures, in and of themselves, do not reveal equilibrium selectivity or transport preferences, which can be discerned only from various functional assays. In this Review, I explore the relationship between cation transport protein structures, equilibrium binding measurements, and ion transport selectivity. The primary focus is on K+-selective channels and nonselective cation channels because they have been extensively studied both functionally and structurally, but the principles discussed are relevant to other transport proteins and molecules. PMID:26078056

Effects of a Variety of Food Extracts and Juices on the Specific Binding Ability of Norovirus GII.4 P Particles

PubMed Central

LI, DAN; BAERT, LEEN; XIA, MING; ZHONG, WEIMING; JIANG, XI; UYTTENDAELE, MIEKE

2014-01-01

The effects of 13 food extracts and juices, including shellfish, fruits, and vegetables, on the binding ability of human norovirus (NoV) were examined, using P particles of human NoV GII.4 as a research surrogate. The enhancements (positive values) or reductions (negative values) of NoV P particle detection (changes in optical density at 450 nm) in the presence of different food extracts and juices as compared with P particles diluted in phosphate-buffered saline were tested by saliva-binding, enzyme-linked immunosorbent assay in triplicate. In the presence of different food extracts and juices at different concentrations, an increase or decrease of the receptor-binding ability of the NoV P particles was observed. Due to a higher specific binding and thus a higher accumulation of the viral particles, oysters may be contaminated with human NoV more often than other shellfish species (mussel, hard clams, and razor clams). Cranberry and pomegranate juices were shown to reduce the specific binding ability of human NoV P particles. No such binding inhibition effects were observed for the other tested extracts of fresh produce (strawberry, blackberry, blueberry, cherry tomato, spinach, romaine lettuce) or, notably, for raspberry, which has been associated with human NoV outbreaks. PMID:22980024

Effects of a variety of food extracts and juices on the specific binding ability of norovirus GII.4 P particles.

PubMed

Li, Dan; Baert, Leen; Xia, Ming; Zhong, Weiming; Jiang, Xi; Uyttendaele, Mieke

2012-07-01

The effects of 13 food extracts and juices, including shellfish, fruits, and vegetables, on the binding ability of human norovirus (NoV) were examined, using P particles of human NoV GII.4 as a research surrogate. The enhancements (positive values) or reductions (negative values) of NoV P particle detection (changes in optical density at 450 nm) in the presence of different food extracts and juices as compared with P particles diluted in phosphate-buffered saline were tested by saliva-binding, enzyme-linked immunosorbent assay in triplicate. In the presence of different food extracts and juices at different concentrations, an increase or decrease of the receptor-binding ability of the NoV P particles was observed. Due to a higher specific binding and thus a higher accumulation of the viral particles, oysters may be contaminated with human NoV more often than other shellfish species (mussel, hard clams, and razor clams). Cranberry and pomegranate juices were shown to reduce the specific binding ability of human NoV P particles. No such binding inhibition effects were observed for the other tested extracts of fresh produce (strawberry, blackberry, blueberry, cherry tomato, spinach, romaine lettuce) or, notably, for raspberry, which has been associated with human NoV outbreaks.

Roles of Copper-Binding Proteins in Breast Cancer.

PubMed

Blockhuys, Stéphanie; Wittung-Stafshede, Pernilla

2017-04-20

Copper ions are needed in several steps of cancer progression. However, the underlying mechanisms, and involved copper-binding proteins, are mainly elusive. Since most copper ions in the body (in and outside cells) are protein-bound, it is important to investigate what copper-binding proteins participate and, for these, how they are loaded with copper by copper transport proteins. Mechanistic information for how some copper-binding proteins, such as extracellular lysyl oxidase (LOX), play roles in cancer have been elucidated but there is still much to learn from a biophysical molecular viewpoint. Here we provide a summary of copper-binding proteins and discuss ones reported to have roles in cancer. We specifically focus on how copper-binding proteins such as mediator of cell motility 1 (MEMO1), LOX, LOX-like proteins, and secreted protein acidic and rich in cysteine (SPARC) modulate breast cancer from molecular and clinical aspects. Because of the importance of copper for invasion/migration processes, which are key components of cancer metastasis, further insights into the actions of copper-binding proteins may provide new targets to combat cancer.

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

Reactive ion etched substrates and methods of making and using

DOEpatents

Rucker, Victor C [San Francisco, CA; Shediac, Rene [Oakland, CA; Simmons, Blake A [San Francisco, CA; Havenstrite, Karen L [New York, NY

2007-08-07

Disclosed herein are substrates comprising reactive ion etched surfaces and specific binding agents immobilized thereon. The substrates may be used in methods and devices for assaying or isolating analytes in a sample. Also disclosed are methods of making the reactive ion etched surfaces.

Fluorimetric detection of Sn(2+) ion in aqueous medium using Salicylaldehyde based nanoparticles and application to natural samples analysis.

PubMed

Patil, Kishor S; Mahajan, Prasad G; Patil, Shivajirao R

2017-01-05

The fluorescent 2-[(E)-(2-phenylhydrazinylidene)methyl]phenol nanoparticles (PHPNPs) were prepared by a simple reprecipitation method. The prepared PHPNPs examined by Dynamic Light Scattering show narrower particle size distribution having an average particle size of 93.3nm. The Scanning Electron Microphotograph shows distinct spherical shaped morphology of nanoparticles. The blue shift in UV-absorption and fluorescence spectra of PHPNPs with respect to corresponding spectra of PHP in acetone solution indicates H- aggregates and Aggregation Induced Enhanced Emission (AIEE) for nanoparticles. The nanoparticles show selective tendency towards the recognition of Sn(2+) ions by enhancing the fluorescence intensity preference to Cu(2+), Fe(3+), Fe(2+), Ni(2+), NH4(+), Ca(2+), Pb(2+), Hg(2+) and Zn(2+) ions, which actually seem to quench the fluorescence of nanoparticles. The studies on Langmuir adsorption plot, fluorescence lifetime of PHPNPs, DLS-Zeta sizer, UV-visible and fluorescence titration with and without Sn(2+) helped to propose a suitable mechanism of fluorescence enhancement of nanoparticles by Sn(2+) and their binding ability during complexation. The fluorescence enhancement effect of PHPNPs induced by Sn(2+) is further used to develop an analytical method for detection of Sn(2+) from aqueous medium in environmental samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Fluorimetric detection of Sn2 + ion in aqueous medium using Salicylaldehyde based nanoparticles and application to natural samples analysis

NASA Astrophysics Data System (ADS)

Patil, Kishor S.; Mahajan, Prasad G.; Patil, Shivajirao R.

2017-01-01

The fluorescent 2-[(E)-(2-phenylhydrazinylidene)methyl]phenol nanoparticles (PHPNPs) were prepared by a simple reprecipitation method. The prepared PHPNPs examined by Dynamic Light Scattering show narrower particle size distribution having an average particle size of 93.3 nm. The Scanning Electron Microphotograph shows distinct spherical shaped morphology of nanoparticles. The blue shift in UV-absorption and fluorescence spectra of PHPNPs with respect to corresponding spectra of PHP in acetone solution indicates H- aggregates and Aggregation Induced Enhanced Emission (AIEE) for nanoparticles. The nanoparticles show selective tendency towards the recognition of Sn2 + ions by enhancing the fluorescence intensity preference to Cu2 +, Fe3 +, Fe2 +, Ni2 +, NH4+, Ca2 +, Pb2 +, Hg2 + and Zn2 + ions, which actually seem to quench the fluorescence of nanoparticles. The studies on Langmuir adsorption plot, fluorescence lifetime of PHPNPs, DLS-Zeta sizer, UV-visible and fluorescence titration with and without Sn2 + helped to propose a suitable mechanism of fluorescence enhancement of nanoparticles by Sn2 + and their binding ability during complexation. The fluorescence enhancement effect of PHPNPs induced by Sn2 + is further used to develop an analytical method for detection of Sn2 + from aqueous medium in environmental samples.

Effect of Mg(2+), Ca(2+), Sr(2+) and Ba(2+) metal ions on the antifungal activity of ZnO nanoparticles tested against Candida albicans.

PubMed

Haja Hameed, Abdulrahman Syedahamed; Karthikeyan, Chandrasekaran; Senthil Kumar, Venugopal; Kumaresan, Subramanian; Sasikumar, Seemaisamy

2015-01-01

The antifungal ability of pure and alkaline metal ion (Mg(2+), Ca(2+), Sr(2+) and Ba(2+)) doped ZnO nanoparticles (NPs) prepared by the co-precipitation method was tested against the pathogenic yeast, Candida albicans (C. albicans), and the results showed that the Mg-doped ZnO NPs possessed greater effect than the other alkaline metal ion doped ZnO NPs. The impact of the concentration of Mg doped ZnO sample on the growth of C. albicans was also studied. The Minimal Fungicidal Concentration (MFC) of the Mg doped ZnO NPs was found to be 2000 μg/ml for which the growth of C. albicans was completely inhibited. The ZnO:Mg sample (1.5mg/ml) with various concentrations of histidine reduced the fungicidal effect of the nanoparticles against C. albicans, which was deliberately explained by the role of ROS. The ZnO:Mg sample added with 5mM of histidine scavenged the ample amount of generated ROS effectively. The binding of the NPs with fungi was observed by their FESEM images and their electrostatic attraction is confirmed by the zeta potential measurement. Copyright © 2015. Published by Elsevier B.V.

Sodium ion effect on silk fibroin conformation characterized by solid-state NMR and generalized 2D NMR NMR correlation

NASA Astrophysics Data System (ADS)

Ruan, Qing-Xia; Zhou, Ping

2008-07-01

In the present work, we investigated Na + ion effect on the silk fibroin (SF) conformation. Samples are Na +-involved regenerated silk fibroin films. 13C CP-MAS NMR demonstrates that as added [Na +] increases, partial silk fibroin conformation transit from helix-form to β-form at certain Na + ion concentration which is much higher than that in Bombyx mori silkworm gland. The generalized two-dimensional NMR-NMR correlation analysis reveals that silk fibroin undergoes several intermediate states during its conformation transition process as [Na +] increase. The appearance order of the intermediates is followed as: helix and/or random coil → helix-like → β-sheet-like → β-sheet, which is the same as that produced by pH decrease from 6.8 to 4.8 in the resultant regenerated silk fibroin films. The binding sites of Na + to silk fibroin might involve the carbonyl oxygen atom of certain amino acids sequence which could promote the formation of β-sheet conformation. Since the Na +sbnd O bond is weak, the ability of Na + inducing the secondary structure transition is weaker than those of Ca 2+, Cu 2+ and even K +. It is maybe a reason why the sodium content is much lower than potassium in the silkworm gland.

Surface plasmon resonance sensing detection of mercury and lead ions based on conducting polymer composite.

PubMed

Abdi, Mahnaz M; Abdullah, Luqman Chuah; Sadrolhosseini, Amir R; Mat Yunus, Wan Mahmood; Moksin, Mohd Maarof; Tahir, Paridah Md

2011-01-01

A new sensing area for a sensor based on surface plasmon resonance (SPR) was fabricated to detect trace amounts of mercury and lead ions. The gold surface used for SPR measurements were modified with polypyrrole-chitosan (PPy-CHI) conducting polymer composite. The polymer layer was deposited on the gold surface by electrodeposition. This optical sensor was used for monitoring toxic metal ions with and without sensitivity enhancement by chitosan in water samples. The higher amounts of resonance angle unit (ΔRU) were obtained for PPy-CHI film due to a specific binding of chitosan with Pb(2+) and Hg(2+) ions. The Pb(2+) ion bind to the polymer films most strongly, and the sensor was more sensitive to Pb(2+) compared to Hg(2+). The concentrations of ions in the parts per million range produced the changes in the SPR angle minimum in the region of 0.03 to 0.07. Data analysis was done by Matlab software using Fresnel formula for multilayer system.

Principles and properties of ion flow in P2X receptors

PubMed Central

Samways, Damien S. K.; Li, Zhiyuan; Egan, Terrance M.

2014-01-01

P2X receptors are a family of trimeric ion channels that are gated by extracellular adenosine 5′-triphosphate (ATP). These receptors have long been a subject of intense research interest by virtue of their vital role in mediating the rapid and direct effects of extracellular ATP on membrane potential and cytosolic Ca2+ concentration, which in turn underpin the ability of ATP to regulate a diverse range of clinically significant physiological functions, including those associated with the cardiovascular, sensory, and immune systems. An important aspect of an ion channel's function is, of course, the means by which it transports ions across the biological membrane. A concerted effort by investigators over the last two decades has culminated in significant advances in our understanding of how P2X receptors conduct the inward flux of Na+ and Ca2+ in response to binding by ATP. However, this work has relied heavily on results from current recordings of P2X receptors altered by site-directed mutagenesis. In the absence of a 3-dimensional channel structure, this prior work provided only a vague and indirect appreciation of the relationship between structure, ion selectivity and flux. The recent publication of the crystal structures for both the closed and open channel conformations of the zebrafish P2X4 receptor has thus proved a significant boon, and has provided an important opportunity to overview the amassed functional data in the context of a working 3-dimensional model of a P2X receptor. In this paper, we will attempt to reconcile the existing functional data regarding ion permeation through P2X receptors with the available crystal structure data, highlighting areas of concordance and discordance as appropriate. PMID:24550775

A comprehensive search for calcium binding sites critical for TMEM16A calcium-activated chloride channel activity

PubMed Central

Tien, Jason; Peters, Christian J; Wong, Xiu Ming; Cheng, Tong; Jan, Yuh Nung; Jan, Lily Yeh; Yang, Huanghe

2014-01-01

TMEM16A forms calcium-activated chloride channels (CaCCs) that regulate physiological processes such as the secretions of airway epithelia and exocrine glands, the contraction of smooth muscles, and the excitability of neurons. Notwithstanding intense interest in the mechanism behind TMEM16A-CaCC calcium-dependent gating, comprehensive surveys to identify and characterize potential calcium sensors of this channel are still lacking. By aligning distantly related calcium-activated ion channels in the TMEM16 family and conducting systematic mutagenesis of all conserved acidic residues thought to be exposed to the cytoplasm, we identify four acidic amino acids as putative calcium-binding residues. Alterations of the charge, polarity, and size of amino acid side chains at these sites alter the ability of different divalent cations to activate the channel. Furthermore, TMEM16A mutant channels containing double cysteine substitutions at these residues are sensitive to the redox potential of the internal solution, providing evidence for their physical proximity and solvent accessibility. DOI: http://dx.doi.org/10.7554/eLife.02772.001 PMID:24980701

Natural hidden antibodies reacting with DNA or cardiolipin bind to thymocytes and evoke their death.

PubMed

Zamulaeva, I A; Lekakh, I V; Kiseleva, V I; Gabai, V L; Saenko, A S; Shevchenko, A S; Poverenny, A M

1997-08-18

Both free and hidden natural antibodies to DNA or cardiolipin were obtained from immunoglobulins of a normal donor. The free antibodies reacting with DNA or cardiolipin were isolated by means of affinity chromatography. Antibodies occurring in an hidden state were disengaged from the depleted immunoglobulins by ion-exchange chromatography and were then affinity-isolated on DNA or cardiolipin sorbents. We used flow cytometry to study the ability of free and hidden antibodies to bind to rat thymocytes. Simultaneously, plasma membrane integrity was tested by propidium iodide (PI) exclusion. The hidden antibodies reacted with 65.2 +/- 10.9% of the thymocytes and caused a fast plasma membrane disruption. Cells (28.7 +/- 7.1%) were stained with PI after incubation with the hidden antibodies for 1 h. The free antibodies bound to a very small fraction of the thymocytes and did not evoke death as compared to control without antibodies. The possible reason for the observed effects is difference in reactivity of the free and hidden antibodies to phospholipids. While free antibodies reacted preferentially with phosphotidylcholine, hidden antibodies reacted with cardiolipin and phosphotidylserine.

Ca2+ sensitizers: An emerging class of agents for counterbalancing weakness in skeletal muscle diseases?

PubMed

Ochala, Julien

2010-02-01

Ca(2+) ions are key regulators of skeletal muscle contraction. By binding to contractile proteins, they initiate a cascade of molecular events leading to cross-bridge formation and ultimately, muscle shortening and force production. The ability of contractile proteins to respond to Ca(2+) attachment, also known as Ca(2+) sensitivity, is often compromised in acquired and congenital skeletal muscle disorders. It constitutes, undoubtedly, a major physiological cause of weakness for patients. In this review, we discuss recent studies giving strong molecular and cellular evidence that pharmacological modulators of some of the contractile proteins, also termed Ca(2+) sensitizers, are efficient agents to improve Ca(2+) sensitivity and function in diseased skeletal muscle cells. In fact, they compensate for the impaired contractile proteins response to Ca(2+) binding. Currently, such Ca(2+) sensitizing compounds are successfully used for reducing problems in cardiac disorders. Therefore, in the future, under certain conditions, these agents may represent an emerging class of agents to enhance the quality of life of patients suffering from skeletal muscle weakness. Copyright 2009 Elsevier B.V. All rights reserved.

Bio-inspired network optimization in soft materials — Insights from the plant cell wall

NASA Astrophysics Data System (ADS)

Vincent, R. R.; Cucheval, A.; Hemar, Y.; Williams, M. A. K.

2009-01-01

The dynamic-mechanical responses of ionotropic gels made from the biopolymer pectin have recently been investigated by microrheological experiments and found to exhibit behaviour indicative of semi-flexible polymer networks. In this work we investigate the gelling behaviour of pectin systems in which an enzyme (pectinmethylesterase, PME) is used to liberate ion-binding sites on initially inert polymers, while in the presence of ions. This is in contrast to the previous work, where it was the release of ions (rather than ion-binding groups) that was controlled and the polymers had pre-existing cross-linkable moieties. In stark contrast to the semi-flexible network paradigm of biological gels and the previous work on pectin, the gels studied herein exhibit the properties of chemically cross-linked networks of flexible polymers.

The effect of Cu 2+ or Fe 3+ on the noncovalent binding of rutin with bovine serum albumin by spectroscopic analysis

NASA Astrophysics Data System (ADS)

Li, Daojin; Zhu, Mei; Xu, Chen; Chen, Jianjun; Ji, Baoming

2011-01-01

The interaction of rutin to bovine serum albumin (BSA) in aqueous solution was investigated by fluorescence spectra and ultraviolet-visible (UV-vis) spectra at pH 7.40. There are also some metal ions present in blood plasma, thus the research about the effect of metal ions on the interaction of drugs with proteins is crucial. Therefore, we have studied the effect of Cu 2+ or Fe 3+ on the interaction between rutin and BSA by using spectroscopic technique at pH 7.40, for the first time. The results of fluorescence titration indicated that rutin could quench the intrinsic fluorescence of BSA in a static quenching way. The binding sites number ( n), the binding constant ( K) and the spatial-distance ( r) of rutin with BSA without or with Cu 2+ or Fe 3+ at 310 K were calculated. The result showed that the presence of Cu 2+ or Fe 3+ increased the binding constant and changed the binding distance between the acceptor and the donor, which possibly results from the formation of metal ions-BSA complex. The effect of rutin on the conformation of BSA was also analyzed using UV under experimental conditions. Furthermore, the fluorescence displacement experiments indicated that rutin is situated within subdomain IIA of BSA.

MtrA of the sodium ion pumping methyltransferase binds cobalamin in a unique mode

PubMed Central

Wagner, Tristan; Ermler, Ulrich; Shima, Seigo

2016-01-01

In the three domains of life, vitamin B12 (cobalamin) is primarily used in methyltransferase and isomerase reactions. The methyltransferase complex MtrA–H of methanogenic archaea has a key function in energy conservation by catalysing the methyl transfer from methyl-tetrahydromethanopterin to coenzyme M and its coupling with sodium-ion translocation. The cobalamin-binding subunit MtrA is not homologous to any known B12-binding proteins and is proposed as the motor of the sodium-ion pump. Here, we present crystal structures of the soluble domain of the membrane-associated MtrA from Methanocaldococcus jannaschii and the cytoplasmic MtrA homologue/cobalamin complex from Methanothermus fervidus. The MtrA fold corresponds to the Rossmann-type α/β fold, which is also found in many cobalamin-containing proteins. Surprisingly, the cobalamin-binding site of MtrA differed greatly from all the other cobalamin-binding sites. Nevertheless, the hydrogen-bond linkage at the lower axial-ligand site of cobalt was equivalently constructed to that found in other methyltransferases and mutases. A distinct polypeptide segment fixed through the hydrogen-bond linkage in the relaxed Co(III) state might be involved in propagating the energy released upon corrinoid demethylation to the sodium-translocation site by a conformational change. PMID:27324530

Cations Stiffen Actin Filaments by Adhering a Key Structural Element to Adjacent Subunits

PubMed Central

2016-01-01

Ions regulate the assembly and mechanical properties of actin filaments. Recent work using structural bioinformatics and site-specific mutagenesis favors the existence of two discrete and specific divalent cation binding sites on actin filaments, positioned in the long axis between actin subunits. Cation binding at one site drives polymerization, while the other modulates filament stiffness and plays a role in filament severing by the regulatory protein, cofilin. Existing structural methods have not been able to resolve filament-associated cations, and so in this work we turn to molecular dynamics simulations to suggest a candidate binding pocket geometry for each site and to elucidate the mechanism by which occupancy of the “stiffness site” affects filament mechanical properties. Incorporating a magnesium ion in the “polymerization site” does not seem to require any large-scale change to an actin subunit’s conformation. Binding of a magnesium ion in the “stiffness site” adheres the actin DNase-binding loop (D-loop) to its long-axis neighbor, which increases the filament torsional stiffness and bending persistence length. Our analysis shows that bound D-loops occupy a smaller region of accessible conformational space. Cation occupancy buries key conserved residues of the D-loop, restricting accessibility to regulatory proteins and enzymes that target these amino acids. PMID:27146246

Free-energy relationships in ion channels activated by voltage and ligand

PubMed Central

Chowdhury, Sandipan

2013-01-01

Many ion channels are modulated by multiple stimuli, which allow them to integrate a variety of cellular signals and precisely respond to physiological needs. Understanding how these different signaling pathways interact has been a challenge in part because of the complexity of underlying models. In this study, we analyzed the energetic relationships in polymodal ion channels using linkage principles. We first show that in proteins dually modulated by voltage and ligand, the net free-energy change can be obtained by measuring the charge-voltage (Q-V) relationship in zero ligand condition and the ligand binding curve at highly depolarizing membrane voltages. Next, we show that the voltage-dependent changes in ligand occupancy of the protein can be directly obtained by measuring the Q-V curves at multiple ligand concentrations. When a single reference ligand binding curve is available, this relationship allows us to reconstruct ligand binding curves at different voltages. More significantly, we establish that the shift of the Q-V curve between zero and saturating ligand concentration is a direct estimate of the interaction energy between the ligand- and voltage-dependent pathway. These free-energy relationships were tested by numerical simulations of a detailed gating model of the BK channel. Furthermore, as a proof of principle, we estimate the interaction energy between the ligand binding and voltage-dependent pathways for HCN2 channels whose ligand binding curves at various voltages are available. These emerging principles will be useful for high-throughput mutagenesis studies aimed at identifying interaction pathways between various regulatory domains in a polymodal ion channel. PMID:23250866

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.

DNA Binding and Phosphorylation Regulate the Core Structure of the NF-κB p50 Transcription Factor

NASA Astrophysics Data System (ADS)

Vonderach, Matthias; Byrne, Dominic P.; Barran, Perdita E.; Eyers, Patrick A.; Eyers, Claire E.

2018-06-01

The NF-κB transcription factors are known to be extensively phosphorylated, with dynamic site-specific modification regulating their ability to dimerize and interact with DNA. p50, the proteolytic product of p105 (NF-κB1), forms homodimers that bind DNA but lack intrinsic transactivation function, functioning as repressors of transcription from κB promoters. Here, we examine the roles of specific phosphorylation events catalysed by either protein kinase A (PKAc) or Chk1, in regulating the functions of p50 homodimers. LC-MS/MS analysis of proteolysed p50 following in vitro phosphorylation allows us to define Ser328 and Ser337 as PKAc- and Chk1-mediated modifications, and pinpoint an additional four Chk1 phosphosites: Ser65, Thr152, Ser242 and Ser248. Native mass spectrometry (MS) reveals Chk1- and PKAc-regulated disruption of p50 homodimer formation through Ser337. Additionally, we characterise the Chk1-mediated phosphosite, Ser242, as a regulator of DNA binding, with a S242D p50 phosphomimetic exhibiting a > 10-fold reduction in DNA binding affinity. Conformational dynamics of phosphomimetic p50 variants, including S242D, are further explored using ion-mobility MS (IM-MS). Finally, comparative theoretical modelling with experimentally observed p50 conformers, in the absence and presence of DNA, reveals that the p50 homodimer undergoes conformational contraction during electrospray ionisation that is stabilised by complex formation with κB DNA.

[Effect of Cu2+ and Zn2+ ions in Ca-ATPase activity isolated from Pachymerus nucleorum (Fabricius) (Coleoptera: Chrysomelidae, Bruchinae) larvae].

PubMed

Dias, Decivaldo S; Coelho, Milton V

2007-01-01

ATPases, an important target of insecticides, are enzymes that hydrolyze ATP and use the energy released in that process to accomplish some type of cellular work. Pachymerus nucleorum (Fabricius) larvae possess an ATPase, that presents high Ca-ATPase activity, but no Mg-ATPase activity. In the present study, the effect of zinc and copper ions in the activity Ca-ATPase of that enzyme was tested. More than 90% of the Ca-ATPase activity was inhibited in 0.5 mM of copper ions or 0.25 mM of zinc ions. In the presence of EDTA, but not in the absence, the inhibition by zinc was reverted with the increase of calcium concentration. The inhibition by copper ions was not reverted in the presence or absence of EDTA. The Ca-ATPase was not inhibited by treatment of the ATPase fraction with copper, suggesting that the copper ion does not bind directly to the enzyme. The results suggest that zinc and copper ions form a complex with ATP and bind to the enzyme inhibiting its Ca-ATPase activity.

Crystal structure of the ATP-gated P2X[subscript 4] ion channel in the closed state

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

Kawate, Toshimitsu; Michel, Jennifer Carlisle; Birdsong, William T.

2009-08-13

P2X receptors are cation-selective ion channels gated by extracellular ATP, and are implicated in diverse physiological processes, from synaptic transmission to inflammation to the sensing of taste and pain. Because P2X receptors are not related to other ion channel proteins of known structure, there is at present no molecular foundation for mechanisms of ligand-gating, allosteric modulation and ion permeation. Here we present crystal structures of the zebrafish P2X{sub 4} receptor in its closed, resting state. The chalice-shaped, trimeric receptor is knit together by subunit-subunit contacts implicated in ion channel gating and receptor assembly. Extracellular domains, rich in {beta}-strands, have largemore » acidic patches that may attract cations, through fenestrations, to vestibules near the ion channel. In the transmembrane pore, the 'gate' is defined by an {approx}8 {angstrom} slab of protein. We define the location of three non-canonical, intersubunit ATP-binding sites, and suggest that ATP binding promotes subunit rearrangement and ion channel opening.« less

Ab initio Study of Transition metal binding to the Prion Protein

NASA Astrophysics Data System (ADS)

Cox, Daniel L.; Singh, Rajiv R. P.; Pan, Jianping

2004-03-01

Fundamental understanding of the prion protein (PrP) is of critical public health importance in view of mad cow and chronic wasting diseases. In recent years, it has been shown that the normal form (PrP^c) binds copper^1), and the structure of the copper binding domain has been elaborated. Hypotheses about toxicity associated with binding of other metals (notably manganese) have been put forward, Accordingly, using the ab initio SIESTA density functional theory code^2), we calculated the binding energy E_B(M) of M-(PrP) complexes relative to initially uncomplexed M ions, with M=Cu,Ni,Zn,Mn and (PrP)^* the minimal binding domain. The binding energy trend is E_B(Ni)>E_B(Cu)>E_B(Zn)>E_B(Mn), consistent with recent experiments apart from the surprising stability of Ni. We will also present preliminary results for binding of initially complexed M ions. *-Supported by U.S. DOE, Office of Basic Energy Sciences, Division of Materials Research 1) G.S. Jackson et al., Proc. Nat. Acad. Sci. (USA) 98, 8531 (2001). 2) P. Ordejón, et al., Phys. Rev. B53, R10441 (1996); J.M. Soler et al., J. Phys. Cond. Matt. 14, 2745 (2002).