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Sample records for active site dynamics

  1. Dynamically achieved active site precision in enzyme catalysis.

    PubMed

    Klinman, Judith P

    2015-02-17

    CONSPECTUS: The grand challenge in enzymology is to define and understand all of the parameters that contribute to enzymes' enormous rate accelerations. The property of hydrogen tunneling in enzyme reactions has moved the focus of research away from an exclusive focus on transition state stabilization toward the importance of the motions of the heavy atoms of the protein, a role for reduced barrier width in catalysis, and the sampling of a protein conformational landscape to achieve a family of protein substates that optimize enzyme-substrate interactions and beyond. This Account focuses on a thermophilic alcohol dehydrogenase for which the chemical step of hydride transfer is rate determining across a wide range of experimental conditions. The properties of the chemical coordinate have been probed using kinetic isotope effects, indicating a transition in behavior below 30 °C that distinguishes nonoptimal from optimal C-H activation. Further, the introduction of single site mutants has the impact of either enhancing or eliminating the temperature dependent transition in catalysis. Biophysical probes, which include time dependent hydrogen/deuterium exchange and fluorescent lifetimes and Stokes shifts, have also been pursued. These studies allow the correlation of spatially resolved transitions in protein motions with catalysis. It is now possible to define a long-range network of protein motions in ht-ADH that extends from a dimer interface to the substrate binding domain across to the cofactor binding domain, over a distance of ca. 30 Å. The ongoing challenge to obtaining spatial and temporal resolution of catalysis-linked protein motions is discussed.

  2. Active-Site Hydration and Water Diffusion in Cytochrome P450cam: A Highly Dynamic Process

    SciTech Connect

    Miao, Yinglong; Baudry, Jerome Y

    2011-01-01

    Long-timescale molecular dynamics simulations (300 ns) are performed on both the apo- (i.e., camphor-free) and camphor-bound cytochrome P450cam (CYP101). Water diffusion into and out of the protein active site is observed without biased sampling methods. During the course of the molecular dynamics simulation, an average of 6.4 water molecules is observed in the camphor-binding site of the apo form, compared to zero water molecules in the binding site of the substrate-bound form, in agreement with the number of water molecules observed in crystal structures of the same species. However, as many as 12 water molecules can be present at a given time in the camphor-binding region of the active site in the case of apo-P450cam, revealing a highly dynamic process for hydration of the protein active site, with water molecules exchanging rapidly with the bulk solvent. Water molecules are also found to exchange locations frequently inside the active site, preferentially clustering in regions surrounding the water molecules observed in the crystal structure. Potential-of-mean-force calculations identify thermodynamically favored trans-protein pathways for the diffusion of water molecules between the protein active site and the bulk solvent. Binding of camphor in the active site modifies the free-energy landscape of P450cam channels toward favoring the diffusion of water molecules out of the protein active site.

  3. Correlated structural kinetics and retarded solvent dynamics at the metalloprotease active site

    SciTech Connect

    Grossman, Moran; Born, Benjamin; Heyden, Matthias; Tworowski, Dmitry; Fields, Gregg B.; Sagi, Irit; Havenith, Martina

    2011-09-18

    Solvent dynamics can play a major role in enzyme activity, but obtaining an accurate, quantitative picture of solvent activity during catalysis is quite challenging. Here, we combine terahertz spectroscopy and X-ray absorption analyses to measure changes in the coupled water-protein motions during peptide hydrolysis by a zinc-dependent human metalloprotease. These changes were tightly correlated with rearrangements at the active site during the formation of productive enzyme-substrate intermediates and were different from those in an enzyme–inhibitor complex. Molecular dynamics simulations showed a steep gradient of fast-to-slow coupled protein-water motions around the protein, active site and substrate. Our results show that water retardation occurs before formation of the functional Michaelis complex. We propose that the observed gradient of coupled protein-water motions may assist enzyme-substrate interactions through water-polarizing mechanisms that are remotely mediated by the catalytic metal ion and the enzyme active site.

  4. Molecular dynamics explorations of active site structure in designed and evolved enzymes.

    PubMed

    Osuna, Sílvia; Jiménez-Osés, Gonzalo; Noey, Elizabeth L; Houk, K N

    2015-04-21

    This Account describes the use of molecular dynamics (MD) simulations to reveal how mutations alter the structure and organization of enzyme active sites. As proposed by Pauling about 70 years ago and elaborated by many others since then, biocatalysis is efficient when functional groups in the active site of an enzyme are in optimal positions for transition state stabilization. Changes in mechanism and covalent interactions are often critical parts of enzyme catalysis. We describe our explorations of the dynamical preorganization of active sites using MD, studying the fluctuations between active and inactive conformations normally concealed to static crystallography. MD shows how the various arrangements of active site residues influence the free energy of the transition state and relates the populations of the catalytic conformational ensemble to the enzyme activity. This Account is organized around three case studies from our laboratory. We first describe the importance of dynamics in evaluating a series of computationally designed and experimentally evolved enzymes for the Kemp elimination, a popular subject in the enzyme design field. We find that the dynamics of the active site is influenced not only by the original sequence design and subsequent mutations but also by the nature of the ligand present in the active site. In the second example, we show how microsecond MD has been used to uncover the role of remote mutations in the active site dynamics and catalysis of a transesterase, LovD. This enzyme was evolved by Tang at UCLA and Codexis, Inc., and is a useful commercial catalyst for the production of the drug simvastatin. X-ray analysis of inactive and active mutants did not reveal differences in the active sites, but relatively long time scale MD in solution showed that the active site of the wild-type enzyme preorganizes only upon binding of the acyl carrier protein (ACP) that delivers the natural acyl group to the active site. In the absence of bound ACP

  5. Dynamic formation of single-atom catalytic active sites on ceria-supported gold nanoparticles

    PubMed Central

    Wang, Yang-Gang; Mei, Donghai; Glezakou, Vassiliki-Alexandra; Li, Jun; Rousseau, Roger

    2015-01-01

    Catalysis by gold supported on reducible oxides has been extensively studied, yet issues such as the nature of the catalytic site and the role of the reducible support remain fiercely debated topics. Here we present ab initio molecular dynamics simulations of an unprecedented dynamic single-atom catalytic mechanism for the oxidation of carbon monoxide by ceria-supported gold clusters. The reported dynamic single-atom catalytic mechanism results from the ability of the gold cation to strongly couple with the redox properties of the ceria in a synergistic manner, thereby lowering the energy of redox reactions. The gold cation can break away from the gold nanoparticle to catalyse carbon monoxide oxidation, adjacent to the metal/oxide interface and subsequently reintegrate back into the nanoparticle after the reaction is completed. Our study highlights the importance of the dynamic creation of active sites under reaction conditions and their essential role in catalysis. PMID:25735407

  6. Molecular dioxygen enters the active site of 12/15-lipoxygenase via dynamic oxygen access channels.

    PubMed

    Saam, Jan; Ivanov, Igor; Walther, Matthias; Holzhütter, Hermann-Georg; Kuhn, Hartmut

    2007-08-14

    Cells contain numerous enzymes that use molecular oxygen for their reactions. Often, their active sites are buried deeply inside the protein, which raises the question whether there are specific access channels guiding oxygen to the site of catalysis. Choosing 12/15-lipoxygenase as a typical example for such oxygen-dependent enzymes, we determined the oxygen distribution within the protein and defined potential routes for oxygen access. For this purpose, we have applied an integrated strategy of structural modeling, molecular dynamics simulations, site-directed mutagenesis, and kinetic measurements. First, we computed the 3D free-energy distribution for oxygen, which led to identification of four oxygen channels in the protein. All channels connect the protein surface with a region of high oxygen affinity at the active site. This region is localized opposite to the nonheme iron providing a structural explanation for the reaction specificity of this lipoxygenase isoform. The catalytically most relevant path can be obstructed by L367F exchange, which leads to a strongly increased Michaelis constant for oxygen. The blocking mechanism is explained in detail by reordering the hydrogen-bonding network of water molecules. Our results provide strong evidence that the main route for oxygen access to the active site of the enzyme follows a channel formed by transiently interconnected cavities whereby the opening and closure are governed by side chain dynamics. PMID:17675410

  7. Dynamics and Mechanism of Efficient DNA Repair Reviewed by Active-Site Mutants

    NASA Astrophysics Data System (ADS)

    Tan, Chuang; Liu, Zheyun; Li, Jiang; Guo, Xunmin; Wang, Lijuan; Zhong, Dongping

    2010-06-01

    Photolyases repair the UV-induced pyrimidine dimers in damage DNA via a photoreaction which includes a series of light-driven electron transfers between the two-electron-reduced flavin cofactor FADH^- and the dimer. We report here our systematic studies of the repair dynamics in E. coli photolyase with mutation of several active-site residues. With femtosecond resolution, we observed the significant change in the forward electron transfer from the excited FADH^- to the dimer and the back electron transfer from the repaired thymines by mutation of E274A, R226A, R342A, N378S and N378C. We also found that the mutation of E274A accelerates the bond-breaking of the thymine dimer. The dynamics changes are consistent with the quantum yield study of these mutants. These results suggest that the active-site residues play a significant role, structurally and chemically, in the DNA repair photocycle.

  8. Wobble Pairs of the HDV Ribozyme Play Specific Roles in Stabilization of Active Site Dynamics

    PubMed Central

    Sripathi, Kamali N.; Banáš, Pavel; Reblova, Kamila; Šponer, Jiři; Otyepka, Michal

    2015-01-01

    The hepatitis delta virus (HDV) is the only known human pathogen whose genome contains a catalytic RNA motif (ribozyme). The overall architecture of the HDV ribozyme is that of a double-nested pseudoknot, with two GU pairs flanking the active site. Although extensive studies have shown that mutation of either wobble results in decreased catalytic activity, little work has focused on linking these mutations to specific structural effects on catalytic fitness. Here we use molecular dynamics simulations based on an activated structure to probe the active site dynamics as a result of wobble pair mutations. In both wild-type and mutant ribozymes, the in-line fitness of the active site (as a measure of catalytic proficiency) strongly depends on the presence of a C75(N3H3+)N1(O5′) hydrogen bond, which positions C75 as the general acid for the reaction. Our mutational analyses show that each GU wobble supports catalytically fit conformations in distinct ways; the reverse G25U20 wobble promotes high in-line fitness, high occupancy of the C75(N3H3+)G1(O5′) general-acid hydrogen bond and stabilization of the G1U37 wobble, while the G1U37 wobble acts more locally by stabilizing high in-line fitness and the C75(N3H3+)G1(O5′) hydrogen bond. We also find that stable type I A-minor and P1.1 hydrogen bonding above and below the active site, respectively, prevent local structural disorder from spreading and disrupting global conformation. Taken together, our results define specific, often redundant architectural roles for several structural motifs of the HDV ribozyme active site, expanding the known roles of these motifs within all HDV-like ribozymes and other structured RNAs. PMID:25631765

  9. Docking and molecular dynamics studies at trypanothione reductase and glutathione reductase active sites.

    PubMed

    Iribarne, Federico; Paulino, Margot; Aguilera, Sara; Murphy, Miguel; Tapia, Orlando

    2002-05-01

    A theoretical docking study on the active sites of trypanothione reductase (TR) and glutathione reductase (GR) with the corresponding natural substrates, trypanothione disulfide (T[S]2) and glutathione disulfide (GSSG), is reported. Molecular dynamics simulations were carried out in order to check the robustness of the docking results. The energetic results are in agreement with previous experimental findings and show the crossed complexes have lower stabilization energies than the natural ones. To test DOCK3.5, four nitro furanic compounds, previously designed as potentially active anti-chagasic molecules, were docked at the GR and TR active sites with the DOCK3.5 procedure. A good correlation was found between differential inhibitory activity and relative interaction energy (affinity). The results provide a validation test for the use of DOCK3.5 in connection with the design of anti-chagasic drugs.

  10. Dynamics of the active site architecture in plant-type ferredoxin-NADP(+) reductases catalytic complexes.

    PubMed

    Sánchez-Azqueta, Ana; Catalano-Dupuy, Daniela L; López-Rivero, Arleth; Tondo, María Laura; Orellano, Elena G; Ceccarelli, Eduardo A; Medina, Milagros

    2014-10-01

    Kinetic isotope effects in reactions involving hydride transfer and their temperature dependence are powerful tools to explore dynamics of enzyme catalytic sites. In plant-type ferredoxin-NADP(+) reductases the FAD cofactor exchanges a hydride with the NADP(H) coenzyme. Rates for these processes are considerably faster for the plastidic members (FNR) of the family than for those belonging to the bacterial class (FPR). Hydride transfer (HT) and deuteride transfer (DT) rates for the NADP(+) coenzyme reduction of four plant-type FNRs (two representatives of the plastidic type FNRs and the other two from the bacterial class), and their temperature dependences are here examined applying a full tunnelling model with coupled environmental fluctuations. Parameters for the two plastidic FNRs confirm a tunnelling reaction with active dynamics contributions, but isotope effects on Arrhenius factors indicate a larger contribution for donor-acceptor distance (DAD) dynamics in the Pisum sativum FNR reaction than in the Anabaena FNR reaction. On the other hand, parameters for bacterial FPRs are consistent with passive environmental reorganisation movements dominating the HT coordinate and no contribution of DAD sampling or gating fluctuations. This indicates that active sites of FPRs are more organised and rigid than those of FNRs. These differences must be due to adaptation of the active sites and catalytic mechanisms to fulfil their particular metabolic roles, establishing a compromise between protein flexibility and functional optimisation. Analysis of site-directed mutants in plastidic enzymes additionally indicates the requirement of a minimal optimal architecture in the catalytic complex to provide a favourable gating contribution. PMID:24953402

  11. Dynamic formation of single-atom catalytic active sites on ceria-supported gold nanoparticles

    SciTech Connect

    Wang, Yanggang; Mei, Donghai; Glezakou, Vassiliki Alexandra; Li, Jun; Rousseau, Roger J.

    2015-03-04

    Ab initio Molecular Dynamics simulations and static Density Functional Theory calculations have been performed to investigate the reaction mechanism of CO oxidation on Au/CeO2 catalyst. It is found that under reaction condition CO adsorption significantly labializes the surface atoms of the Au cluster and leads to the formation of isolated Au+-CO species that resides on the support in the vicinity of the Au particle. In this context, we identified a dynamic single-atom catalytic mechanism at the interfacial area for CO oxidation on Au/CeO2 catalyst, which is a lower energy pathway than that of CO oxidation at the interface with the metal particle. This results from the ability of the single atom site to strongly couple with the redox properties of the support in a synergistic manner thereby lowering the barrier for redox reactions. We find that the single Au+ ion, which only exists under reaction conditions, breaks away from the Au cluster to catalyze CO oxidation and returns to the Au cluster after the catalytic cycle is completed. Generally, our study highlights the importance of the dynamic creation of active sites under reaction conditions and their essential role in a catalytic process.

  12. Dynamic formation of single-atom catalytic active sites on ceria-supported gold nanoparticles

    DOE PAGES

    Wang, Yanggang; Mei, Donghai; Glezakou, Vassiliki Alexandra; Li, Jun; Rousseau, Roger J.

    2015-03-04

    Ab initio Molecular Dynamics simulations and static Density Functional Theory calculations have been performed to investigate the reaction mechanism of CO oxidation on Au/CeO2 catalyst. It is found that under reaction condition CO adsorption significantly labializes the surface atoms of the Au cluster and leads to the formation of isolated Au+-CO species that resides on the support in the vicinity of the Au particle. In this context, we identified a dynamic single-atom catalytic mechanism at the interfacial area for CO oxidation on Au/CeO2 catalyst, which is a lower energy pathway than that of CO oxidation at the interface with themore » metal particle. This results from the ability of the single atom site to strongly couple with the redox properties of the support in a synergistic manner thereby lowering the barrier for redox reactions. We find that the single Au+ ion, which only exists under reaction conditions, breaks away from the Au cluster to catalyze CO oxidation and returns to the Au cluster after the catalytic cycle is completed. Generally, our study highlights the importance of the dynamic creation of active sites under reaction conditions and their essential role in a catalytic process.« less

  13. Ligand-dependent dynamics of the active-site lid in bacterial dimethylarginine dimethylaminohydrolase.

    PubMed

    Rasheed, Masooma; Richter, Christine; Chisty, Liisa T; Kirkpatrick, John; Blackledge, Martin; Webb, Martin R; Driscoll, Paul C

    2014-02-18

    The dimethylarginine dimethylaminohydrolase (DDAH) enzyme family has been the subject of substantial investigation as a potential therapeutic target for the regulation of vascular tension. DDAH enzymes catalyze the conversion of asymmetric N(η),N(η)-dimethylarginine (ADMA) to l-citrulline. Here the influence of substrate and product binding on the dynamic flexibility of DDAH from Pseudomonas aeruginosa (PaDDAH) has been assessed. A combination of heteronuclear NMR spectroscopy, static and time-resolved fluorescence measurements, and atomistic molecular dynamics simulations was employed. A monodisperse monomeric variant of the wild-type enzyme binds the reaction product l-citrulline with a low millimolar dissociation constant. A second variant, engineered to be catalytically inactive by substitution of the nucleophilic Cys249 residue with serine, can still convert the substrate ADMA to products very slowly. This PaDDAH variant also binds l-citrulline, but with a low micromolar dissociation constant. NMR and molecular dynamics simulations indicate that the active site "lid", formed by residues Gly17-Asp27, exhibits a high degree of internal motion on the picosecond-to-nanosecond time scale. This suggests that the lid is open in the apo state and allows substrate access to the active site that is otherwise buried. l-Citrulline binding to both protein variants is accompanied by an ordering of the lid. Modification of PaDDAH with a coumarin fluorescence reporter allowed measurement of the kinetic mechanism of the PaDDAH reaction. A combination of NMR and kinetic data shows that the catalytic turnover of the enzyme is not limited by release of the l-citrulline product. The potential to develop the coumarin-PaDDAH adduct as an l-citrulline sensor is discussed. PMID:24484052

  14. Site-specific phosphorylation and microtubule dynamics control Pyrin inflammasome activation.

    PubMed

    Gao, Wenqing; Yang, Jieling; Liu, Wang; Wang, Yupeng; Shao, Feng

    2016-08-16

    Pyrin, encoded by the MEFV gene, is best known for its gain-of-function mutations causing familial Mediterranean fever (FMF), an autoinflammatory disease. Pyrin forms a caspase-1-activating inflammasome in response to inactivating modifications of Rho GTPases by various bacterial toxins or effectors. Pyrin-mediated innate immunity is unique in that it senses bacterial virulence rather than microbial molecules, but its mechanism of activation is unknown. Here we show that Pyrin was phosphorylated in bone marrow-derived macrophages and dendritic cells. We identified Ser-205 and Ser-241 in mouse Pyrin whose phosphorylation resulted in inhibitory binding by cellular 14-3-3 proteins. The two serines underwent dephosphorylation upon toxin stimulation or bacterial infection, triggering 14-3-3 dissociation, which correlated with Pyrin inflammasome activation. We developed antibodies specific for phosphorylated Ser-205 and Ser-241, which confirmed the stimuli-induced dephosphorylation of endogenous Pyrin. Mutational analyses indicated that both phosphorylation and signal-induced dephosphorylation of Ser-205/241 are important for Pyrin activation. Moreover, microtubule drugs, including colchicine, commonly used to treat FMF, effectively blocked activation of the Pyrin inflammasome. These drugs did not affect Pyrin dephosphorylation and 14-3-3 dissociation but inhibited Pyrin-mediated apoptosis-associated Speck-like protein containing CARD (ASC) aggregation. Our study reveals that site-specific (de)phosphorylation and microtubule dynamics critically control Pyrin inflammasome activation, illustrating a fine and complex mechanism in cytosolic immunity. PMID:27482109

  15. Coordination number of zinc ions in the phosphotriesterase active site by molecular dynamics and quantum mechanics.

    PubMed

    Koca, Jaroslav; Zhan, Chang-Guo; Rittenhouse, Robert C; Ornstein, Rick L

    2003-02-01

    We have run several molecular dynamics (MD) simulations on zinc-containing phosphotriesterase (PTE) with two bound substrates, sarin and paraoxon, and with the substrate analog diethyl 4-methylbenzylphosphonate. A standard nonbonded model was employed to treat the zinc ions with the commonly used charge of +2. In all the trajectories, we observed a tightly bound water (TBW) molecule in the active site that was coordinated to the less buried zinc ion. The phosphoryl oxygen of the substrate/inhibitor was found to be coordinated to the same zinc ion so that, considering all ligands, the less buried zinc was hexa-coordinated. The hexa-coordination of this zinc ion was not seen in the deposited X-ray pdb files for PTE. Several additional MD simulations were then performed using different charges (+1, +1.5) on the zinc ions, along with ab initio and density functional theory (DFT) calculations, to evaluate the following possibilities: the crystal diffraction data were not correctly interpreted; the hexa-coordinated zinc ion in PTE is only present in solution and not in the crystal; and the hexa-coordinated zinc ion in PTE is an artifact of the force field used. A charge of +1.5 leads to a coordination number (CN) of 5 on both zinc ions, which is consistent with the results from ab initio and DFT calculations and with the latest high resolution X-ray crystal structure. The commonly used charge of +2 produces a CN of 6 on the less buried zinc. The CN on the more buried zinc ion is 5 when the substrate/inhibitor is present in the simulation, and increases to 6 when the substrate/inhibitor is removed prior to the simulation. The results of both of the MD and quantum mechanical calculations lead to the conclusion that the zinc ions in the PTE active site are both penta-coordinated, and that the MD simulations performed with the charge of +2 overestimate the CN of the zinc ions in the PTE active site. The overall protein structures in the simulations remain unaffected by the

  16. Rapid binding of a cationic active site inhibitor to wild type and mutant mouse acetylcholinesterase: Brownian dynamics simulation including diffusion in the active site gorge.

    PubMed

    Tara, S; Elcock, A H; Kirchhoff, P D; Briggs, J M; Radic, Z; Taylor, P; McCammon, J A

    1998-12-01

    It is known that anionic surface residues play a role in the long-range electrostatic attraction between acetylcholinesterase and cationic ligands. In our current investigation, we show that anionic residues also play an important role in the behavior of the ligand within the active site gorge of acetylcholinesterase. Negatively charged residues near the gorge opening not only attract positively charged ligands from solution to the enzyme, but can also restrict the motion of the ligand once it is inside of the gorge. We use Brownian dynamics techniques to calculate the rate constant kon, for wild type and mutant acetylcholinesterase with a positively charged ligand. These calculations are performed by allowing the ligand to diffuse within the active site gorge. This is an extension of previously reported work in which a ligand was allowed to diffuse only to the enzyme surface. By setting the reaction criteria for the ligand closer to the active site, better agreement with experimental data is obtained. Although a number of residues influence the movement of the ligand within the gorge, Asp74 is shown to play a particularly important role in this function. Asp74 traps the ligand within the gorge, and in this way helps to ensure a reaction.

  17. Dynamics of an Active-Site Flap Contributes to Catalysis in a JAMM Family Metallo Deubiquitinase.

    PubMed

    Bueno, Amy N; Shrestha, Rashmi K; Ronau, Judith A; Babar, Aditya; Sheedlo, Michael J; Fuchs, Julian E; Paul, Lake N; Das, Chittaranjan

    2015-10-01

    The endosome-associated deubiquitinase (DUB) AMSH is a member of the JAMM family of zinc-dependent metallo isopeptidases with high selectivity for Lys63-linked polyubiquitin chains, which play a key role in endosomal-lysosomal sorting of activated cell surface receptors. The catalytic domain of the enzyme features a flexible flap near the active site that opens and closes during its catalytic cycle. Structural analysis of its homologues, AMSH-LP (AMSH-like protein) and the fission yeast counterpart, Sst2, suggests that a conserved Phe residue in the flap may be critical for substrate binding and/or catalysis. To gain insight into the contribution of this flap in substrate recognition and catalysis, we generated mutants of Sst2 and characterized them using a combination of enzyme kinetics, X-ray crystallography, molecular dynamics simulations, and isothermal titration calorimetry (ITC). Our analysis shows that the Phe residue in the flap contributes key interactions during the rate-limiting step but not to substrate binding, since mutants of Phe403 exhibit a defect only in kcat but not in KM. Moreover, ITC studies show Phe403 mutants have similar KD for ubiquitin compared to the wild-type enzyme. The X-ray structures of both Phe403Ala and the Phe403Trp, in both the free and ubiquitin bound form, reveal no appreciable structural change that might impair substrate or alter product binding. We observed that the side chain of the Trp residue is oriented identically with respect to the isopeptide moiety of the substrate as the Phe residue in the wild-type enzyme, so the loss of activity seen in this mutant cannot be explained by the absence of a group with the ability to provide van der Waals interactions that facilitate the hyrdolysis of the Lys63-linked diubiquitin. Molecular dynamics simulations indicate that the flap in the Trp mutant is quite flexible, allowing almost free rotation of the indole side chain. Therefore, it is possible that these different dynamic

  18. Analysis of structural changes in active site of luciferase adsorbed on nanofabricated hydrophilic Si surface by molecular-dynamics simulations

    SciTech Connect

    Nishiyama, Katsuhiko; Hoshino, Tadatsugu

    2007-05-21

    Interactions between luciferase and a nanofabricated hydrophilic Si surface were explored by molecular-dynamics simulations. The structural changes in the active-site residues, the residues affecting the luciferin binding, and the residues affecting the bioluminescence color were smaller on the nanofabricated hydrophilic Si surface than on both a hydrophobic Si surface and a hydrophilic Si surface. The nanofabrication and wet-treatment techniques are expected to prevent the decrease in activity of luciferase on the Si surface.

  19. New Insights into Active Site Conformation Dynamics of E. coli PNP Revealed by Combined H/D Exchange Approach and Molecular Dynamics Simulations

    NASA Astrophysics Data System (ADS)

    Kazazić, Saša; Bertoša, Branimir; Luić, Marija; Mikleušević, Goran; Tarnowski, Krzysztof; Dadlez, Michal; Narczyk, Marta; Bzowska, Agnieszka

    2016-01-01

    The biologically active form of purine nucleoside phosphorylase (PNP) from Escherichia coli (EC 2.4.2.1) is a homohexamer unit, assembled as a trimer of dimers. Upon binding of phosphate, neighboring monomers adopt different active site conformations, described as open and closed. To get insight into the functions of the two distinctive active site conformations, virtually inactive Arg24Ala mutant is complexed with phosphate; all active sites are found to be in the open conformation. To understand how the sites of neighboring monomers communicate with each other, we have combined H/D exchange (H/DX) experiments with molecular dynamics (MD) simulations. Both methods point to the mobility of the enzyme, associated with a few flexible regions situated at the surface and within the dimer interface. Although H/DX provides an average extent of deuterium uptake for all six hexamer active sites, it was able to indicate the dynamic mechanism of cross-talk between monomers, allostery. Using this technique, it was found that phosphate binding to the wild type (WT) causes arrest of the molecular motion in backbone fragments that are flexible in a ligand-free state. This was not the case for the Arg24Ala mutant. Upon nucleoside substrate/inhibitor binding, some release of the phosphate-induced arrest is observed for the WT, whereas the opposite effects occur for the Arg24Ala mutant. MD simulations confirmed that phosphate is bound tightly in the closed active sites of the WT; conversely, in the open conformation of the active site of the WT phosphate is bound loosely moving towards the exit of the active site. In Arg24Ala mutant binary complex Pi is bound loosely, too.

  20. Improving the activity of the subtilisin nattokinase by site-directed mutagenesis and molecular dynamics simulation.

    PubMed

    Weng, Meizhi; Deng, Xiongwei; Bao, Wei; Zhu, Li; Wu, Jieyuan; Cai, Yongjun; Jia, Yan; Zheng, Zhongliang; Zou, Guolin

    2015-09-25

    Nattokinase (NK), a bacterial serine protease from Bacillus subtilis var. natto, is a potential cardiovascular drug exhibiting strong fibrinolytic activity. To broaden its commercial and medical applications, we constructed a single-mutant (I31L) and two double-mutants (M222A/I31L and T220S/I31L) by site-directed mutagenesis. Active enzymes were expressed in Escherichia coli with periplasmic secretion and were purified to homogeneity. The kinetic parameters of enzymes were examined by spectroscopy assay and isothermal titration calorimetry (ITC), and their fibrinolytic activities were determined by fibrin plate method. The substitution of Leu(31) for Ile(31) resulted in about 2-fold enhancement of catalytic efficiency (Kcat/KM) compared with wild-type NK. The specific activities of both double-mutants (M222A/I31L and T220S/I31L) were significantly increased when compared with the single-mutants (M222A and T220S) and the oxidative stability of M222A/I31L mutant was enhanced with respect to wild-type NK. This study demonstrates the feasibility of improving activity of NK by site-directed mutagenesis and shows successful protein engineering cases to improve the activity of NK as a potent therapeutic agent.

  1. Conformational dynamics of the active site loop of S-adenosylmethionine synthetase illuminated by site-directed spin labeling.

    PubMed

    Taylor, John C; Markham, George D

    2003-07-15

    S-adenosylmethionine synthetase (ATP: L-methionine S-adenosyltransferase, methionine adenosyltransferase, a.k.a. MAT) is one of numerous enzymes that have a flexible polypeptide loop that moves to gate access to the active site in a motion that is closely coupled to catalysis. Crystallographic studies of this tetrameric enzyme have shown that the loop is closed in the absence of bound substrates. However, the loop must open to allow substrate binding and a variety of data indicate that the loop is closed during the catalytic steps. Previous kinetic studies indicate that during turnover loop motion occurs on a time scale of 10(-2)s, ca. 10-fold faster than chemical transformations and turnover. Site-directed spin labeling has been used to introduce nitroxide groups at two positions in the loop to illuminate how the motion of the loop is affected by substrate binding. The two loop mutants constructed, G105C and D107C, retain wild type levels of MAT activity; attachment of a methanethiosulfonate spin label to convert the cysteine to the "R1" residue reduced the k(cat) only for the labeled D107R1 form (7-fold). The K(m) value for methionine increased 2- to 4-fold for the cysteine mutants and 2- to 7-fold for the labeled proteins, whereas the K(m) for ATP was changed by at most 2-fold. EPR spectra for both labeled proteins are nearly identical and show the presence of two major spin label environments with rotational diffusion rates differing by approximately 10-fold; the slower rate is ca. 4-fold faster than the estimated protein rotational rate. The spectra are not altered by addition of substrates or products. At both positions the less mobile conformation constitutes ca. 65% of the total species, indicating an equilibrium that only slightly favors one form, that in which the label is more immobilized. The equilibrium constant that relates the two forms is comparable to the equilibrium constant of 1.5 for a conformational change that was previously deduced from the

  2. The Role of Distant Mutations and Allosteric Regulation on LovD Active Site Dynamics

    PubMed Central

    Jiménez-Osés, Gonzalo; Osuna, Sílvia; Gao, Xue; Sawaya, Michael R.; Gilson, Lynne; Collier, Steven J.; Huisman, Gjalt W.; Yeates, Todd O.; Tang, Yi; Houk, K. N.

    2014-01-01

    Natural enzymes have evolved to perform their cellular functions under complex selective pressures, which often require their catalytic activities to be regulated by other proteins. We contrasted a natural enzyme, LovD, which acts on a protein-bound (LovF) acyl substrate, with a laboratory-generated variant that was transformed by directed evolution to accept instead a small free acyl thioester, and no longer requires the acyl carrier protein. The resulting 29-mutant variant is 1000-fold more efficient in the synthesis of the drug simvastatin than the wild-type LovD. This is the first non-patent report of the enzyme currently used for the manufacture of simvastatin, as well as the intermediate evolved variants. Crystal structures and microsecond molecular dynamics simulations revealed the mechanism by which the laboratory-generated mutations free LovD from dependence on protein-protein interactions. Mutations dramatically altered conformational dynamics of the catalytic residues, obviating the need for allosteric modulation by the acyl carrier LovF. PMID:24727900

  3. Single Active Site Mutation Causes Serious Resistance of HIV Reverse Transcriptase to Lamivudine: Insight from Multiple Molecular Dynamics Simulations.

    PubMed

    Moonsamy, Suri; Bhakat, Soumendranath; Walker, Ross C; Soliman, Mahmoud E S

    2016-03-01

    Molecular dynamics simulations, binding free energy calculations, principle component analysis (PCA), and residue interaction network analysis were employed in order to investigate the molecular mechanism of M184I single mutation which played pivotal role in making the HIV-1 reverse transcriptase (RT) totally resistant to lamivudine. Results showed that single mutations at residue 184 of RT caused (1) distortion of the orientation of lamivudine in the active site due to the steric conflict between the oxathiolane ring of lamivudine and the side chain of beta-branched amino acids Ile at position 184 which, in turn, perturbs inhibitor binding, (2) decrease in the binding affinity by (~8 kcal/mol) when compared to the wild-type, (3) variation in the overall enzyme motion as evident from the PCA for both systems, and (4) distortion of the hydrogen bonding network and atomic interactions with the inhibitor. The comprehensive analysis presented in this report can provide useful information for understanding the drug resistance mechanism against lamivudine. The results can also provide some potential clues for further design of novel inhibitors that are less susceptible to drug resistance. PMID:26972300

  4. A Dynamic Zn Site in Helicobacter pylori HypA: A Potential Mechanism for Metal-Specific Protein Activity

    SciTech Connect

    Kennedy,D.; Herbst, R.; Iwig, J.; Chivers, P.; Maroney, M.

    2007-01-01

    HypA is an accessory protein and putative metallochaperone that is critical for supplying nickel to the active site of NiFe hydrogenases. In addition to binding Ni(II), HypA is known to contain a Zn site that has been suggested to play a structural role. X-ray absorption spectroscopy has been used to show that the Zn site changes structure upon binding nickel, from a S{sub 3}(O/N)-donor ligand environment to an S{sub 4}-donor ligand environment. This provides a potential mechanism for discriminating Ni(II) from other divalent metal ions. The Ni(II) site is shown to be a six-coordinate complex composed of O/N-donors including two histidines. As such, it resembles the nickel site in UreE, a nickel metallochaperone involved in nickel incorporation into urease.

  5. Conformational flexibility related to enzyme activity: evidence for a dynamic active-site gatekeeper function of Tyr215 in Aerococcus viridans lactate oxidase

    PubMed Central

    Stoisser, Thomas; Brunsteiner, Michael; Wilson, David K.; Nidetzky, Bernd

    2016-01-01

    L-Lactate oxidase (LOX) belongs to a large family of flavoenzymes that catalyze oxidation of α-hydroxy acids. How in these enzymes the protein structure controls reactivity presents an important but elusive problem. LOX contains a prominent tyrosine in the substrate binding pocket (Tyr215 in Aerococcus viridans LOX) that is partially responsible for securing a flexible loop which sequesters the active site. To characterize the role of Tyr215, effects of substitutions of the tyrosine (Y215F, Y215H) were analyzed kinetically, crystallographically and by molecular dynamics simulations. Enzyme variants showed slowed flavin reduction and oxidation by up to 33-fold. Pyruvate release was also decelerated and in Y215F, it was the slowest step overall. A 2.6-Å crystal structure of Y215F in complex with pyruvate shows the hydrogen bond between the phenolic hydroxyl and the keto oxygen in pyruvate is replaced with a potentially stronger hydrophobic interaction between the phenylalanine and the methyl group of pyruvate. Residues 200 through 215 or 216 appear to be disordered in two of the eight monomers in the asymmetric unit suggesting that they function as a lid controlling substrate entry and product exit from the active site. Substitutions of Tyr215 can thus lead to a kinetic bottleneck in product release. PMID:27302031

  6. Active site binding modes of inhibitors of Staphylococcus aureus mevalonate diphosphate decarboxylase from docking and molecular dynamics simulations.

    PubMed

    Addo, James K; Skaff, D Andrew; Miziorko, Henry M

    2016-01-01

    Bacterial mevalonate diphosphate decarboxylase (MDD) is an attractive therapeutic target for antibacterial drug development. In this work, we discuss a combined docking and molecular dynamics strategy toward inhibitor binding to bacterial MDD. The docking parameters utilized in this study were first validated with observations for the inhibitors 6-fluoromevalonate diphosphate (FMVAPP) and diphosphoglycolylproline (DPGP) using existing structures for the Staphylococcus epidermidis enzyme. The validated docking protocol was then used to predict structures of the inhibitors bound to Staphylococcus aureus MDD using the unliganded crystal structure of Staphylococcus aureus MDD. We also investigated a possible interactions improvement by combining this docking method with molecular dynamics simulations. Thus, the predicted docking structures were analyzed in a molecular dynamics trajectory to generate dynamic models and reinforce the predicted binding modes. FMVAPP is predicted to make more extensive contacts with S. aureus MDD, forming stable hydrogen bonds with Arg144, Arg193, Lys21, Ser107, and Tyr18, as well as making stable hydrophobic interactions with Tyr18, Trp19, and Met196. The differences in predicted binding are supported by experimentally determined Ki values of 0.23 ± 0.02 and 34 ± 8 μM, for FMVAPP and DPGP, respectively. The structural information coupled with the kinetic characterization obtained from this study should be useful in defining the requirements for inhibition as well as in guiding the selection of active compounds for inhibitor optimization.

  7. Active Site Loop Dynamics of a Class IIa Fructose 1,6-Bisphosphate Aldolase from Mycobacterium tuberculosis

    SciTech Connect

    Pegan, Scott D.; Rukseree, Kamolchanok; Capodagli, Glenn C.; Baker, Erica A.; Krasnykh, Olga; Franzblau, Scott G.; Mesecar, Andrew D.

    2013-01-08

    The class II fructose 1,6-bisphosphate aldolases (FBAs, EC 4.1.2.13) comprises one of two families of aldolases. Instead of forming a Schiff base intermediate using an ε-amino group of a lysine side chain, class II FBAs utilize Zn(II) to stabilize a proposed hydroxyenolate intermediate (HEI) in the reversible cleavage of fructose 1,6-bisphosphate, forming glyceraldehyde 3-phosphate and dihydroxyacetone phosphate (DHAP). As class II FBAs have been shown to be essential in pathogenic bacteria, focus has been placed on these enzymes as potential antibacterial targets. Although structural studies of class II FBAs from Mycobacterium tuberculosis (MtFBA), other bacteria, and protozoa have been reported, the structure of the active site loop responsible for catalyzing the protonation–deprotonation steps of the reaction for class II FBAs has not yet been observed. We therefore utilized the potent class II FBA inhibitor phosphoglycolohydroxamate (PGH) as a mimic of the HEI- and DHAP-bound form of the enzyme and determined the X-ray structure of the MtFBA–PGH complex to 1.58 Å. Remarkably, we are able to observe well-defined electron density for the previously elusive active site loop of MtFBA trapped in a catalytically competent orientation. Utilization of this structural information and site-directed mutagenesis and kinetic studies conducted on a series of residues within the active site loop revealed that E169 facilitates a water-mediated deprotonation–protonation step of the MtFBA reaction mechanism. Furthermore, solvent isotope effects on MtFBA and catalytically relevant mutants were used to probe the effect of loop flexibility on catalytic efficiency. Additionally, we also reveal the structure of MtFBA in its holoenzyme form.

  8. Active site loop dynamics of a class IIa fructose 1,6-bisphosphate aldolase from Mycobacterium tuberculosis.

    PubMed

    Pegan, Scott D; Rukseree, Kamolchanok; Capodagli, Glenn C; Baker, Erica A; Krasnykh, Olga; Franzblau, Scott G; Mesecar, Andrew D

    2013-02-01

    Class II fructose 1,6-bisphosphate aldolases (FBAs, EC 4.1.2.13) comprise one of two families of aldolases. Instead of forming a Schiff base intermediate using an ε-amino group of a lysine side chain, class II FBAs utilize Zn(II) to stabilize a proposed hydroxyenolate intermediate (HEI) in the reversible cleavage of fructose 1,6-bisphosphate, forming glyceraldehyde 3-phosphate and dihydroxyacetone phosphate (DHAP). As class II FBAs have been shown to be essential in pathogenic bacteria, focus has been placed on these enzymes as potential antibacterial targets. Although structural studies of class II FBAs from Mycobacterium tuberculosis (MtFBA), other bacteria, and protozoa have been reported, the structure of the active site loop responsible for catalyzing the protonation-deprotonation steps of the reaction for class II FBAs has not yet been observed. We therefore utilized the potent class II FBA inhibitor phosphoglycolohydroxamate (PGH) as a mimic of the HEI- and DHAP-bound form of the enzyme and determined the X-ray structure of the MtFBA-PGH complex to 1.58 Å. Remarkably, we are able to observe well-defined electron density for the previously elusive active site loop of MtFBA trapped in a catalytically competent orientation. Utilization of this structural information and site-directed mutagenesis and kinetic studies conducted on a series of residues within the active site loop revealed that E169 facilitates a water-mediated deprotonation-protonation step of the MtFBA reaction mechanism. Also, solvent isotope effects on MtFBA and catalytically relevant mutants were used to probe the effect of loop flexibility on catalytic efficiency. Additionally, we also reveal the structure of MtFBA in its holoenzyme form.

  9. Probing the importance of hydrogen bonds in the active site of the subtilisin nattokinase by site-directed mutagenesis and molecular dynamics simulation

    PubMed Central

    Zheng, Zhong-liang; Ye, Mao-qing; Zuo, Zhen-yu; Liu, Zhi-gang; Tai, Keng-chang; Zou, Guo-lin

    2006-01-01

    Hydrogen bonds occurring in the catalytic triad (Asp32, His64 and Ser221) and the oxyanion hole (Asn155) are very important to the catalysis of peptide bond hydrolysis by serine proteases. For the subtilisin NK (nattokinase), a bacterial serine protease, construction and analysis of a three-dimensional structural model suggested that several hydrogen bonds formed by four residues function to stabilize the transition state of the hydrolysis reaction. These four residues are Ser33, Asp60, Ser62 and Thr220. In order to remove the effect of these hydrogen bonds, four mutants (Ser33→Ala33, Asp60→Ala60, Ser62→Ala62, and Thr220→Ala220) were constructed by site-directed mutagenesis. The results of enzyme kinetics indicated that removal of these hydrogen bonds increases the free-energy of the transition state (ΔΔGT). We concluded that these hydrogen bonds are more important for catalysis than for binding the substrate, because removal of these bonds mainly affects the kcat but not the Km values. A substrate, SUB1 (succinyl-Ala-Ala-Pro-Phe-p-nitroanilide), was used during enzyme kinetics experiments. In the present study we have also shown the results of FEP (free-energy perturbation) calculations with regard to the binding and catalysis reactions for these mutant subtilisins. The calculated difference in FEP also suggested that these four residues are more important for catalysis than binding of the substrate, and the simulated values compared well with the experimental values from enzyme kinetics. The results of MD (molecular dynamics) simulations further demonstrated that removal of these hydrogen bonds partially releases Asp32, His64 and Asn155 so that the stability of the transition state decreases. Another substrate, SUB2 (H-D-Val-Leu-Lys-p-nitroanilide), was used for FEP calculations and MD simulations. PMID:16411898

  10. Probing the importance of hydrogen bonds in the active site of the subtilisin nattokinase by site-directed mutagenesis and molecular dynamics simulation.

    PubMed

    Zheng, Zhong-liang; Ye, Mao-qing; Zuo, Zhen-yu; Liu, Zhi-gang; Tai, Keng-chang; Zou, Guo-lin

    2006-05-01

    Hydrogen bonds occurring in the catalytic triad (Asp32, His64 and Ser221) and the oxyanion hole (Asn155) are very important to the catalysis of peptide bond hydrolysis by serine proteases. For the subtilisin NK (nattokinase), a bacterial serine protease, construction and analysis of a three-dimensional structural model suggested that several hydrogen bonds formed by four residues function to stabilize the transition state of the hydrolysis reaction. These four residues are Ser33, Asp60, Ser62 and Thr220. In order to remove the effect of these hydrogen bonds, four mutants (Ser33-->Ala33, Asp60-->Ala60, Ser62-->Ala62, and Thr220-->Ala220) were constructed by site-directed mutagenesis. The results of enzyme kinetics indicated that removal of these hydrogen bonds increases the free-energy of the transition state (DeltaDeltaG(T)). We concluded that these hydrogen bonds are more important for catalysis than for binding the substrate, because removal of these bonds mainly affects the kcat but not the K(m) values. A substrate, SUB1 (succinyl-Ala-Ala-Pro-Phe-p-nitroanilide), was used during enzyme kinetics experiments. In the present study we have also shown the results of FEP (free-energy perturbation) calculations with regard to the binding and catalysis reactions for these mutant subtilisins. The calculated difference in FEP also suggested that these four residues are more important for catalysis than binding of the substrate, and the simulated values compared well with the experimental values from enzyme kinetics. The results of MD (molecular dynamics) simulations further demonstrated that removal of these hydrogen bonds partially releases Asp32, His64 and Asn155 so that the stability of the transition state decreases. Another substrate, SUB2 (H-D-Val-Leu-Lys-p-nitroanilide), was used for FEP calculations and MD simulations.

  11. Short-time dynamics of pH-dependent conformation and substrate binding in the active site of beta-glucosidases: A computational study.

    PubMed

    Flannelly, David F; Aoki, Thalia G; Aristilde, Ludmilla

    2015-09-01

    The complete degradation of cellulose to glucose is essential to carbon turnover in terrestrial ecosystems and to engineered biofuel production. A rate-limiting step in this pathway is catalyzed by beta-glucosidase (BG) enzymes, which convert cellulobiose into two glucose molecules. The activity of these enzymes has been shown to vary with solution pH. However, it is not well understood how pH influences the enzyme conformation required for catalytic action on the substrate. A structural understanding of this pH effect is important for predicting shifts in BG activity in bioreactors and environmental matrices, in addition to informing targeted protein engineering. Here we applied molecular dynamics simulations to explore conformational and substrate binding dynamics in two well-characterized BGs of bacterial (Clostridium cellulovorans) and fungal (Trichoderma reesei) origins as a function of pH. The enzymes were simulated in an explicit solvated environment, with NaCl as electrolytes, at their prominent ionization states obtained at pH 5, 6, 7, and 7.5. Our findings indicated that pH-dependent changes in the ionization states of non-catalytic residues localized outside of the immediate active site led to pH-dependent disruption of the active site conformation. This disruption interferes with favorable H-bonding interactions with catalytic residues required to initiate catalysis on the substrate. We also identified specific non-catalytic residues that are involved in stabilizing the substrate at the optimal pH for enzyme activity. The simulations further revealed the dynamics of water-bridging interactions both outside and inside the substrate binding cleft during structural changes in the enzyme-substrate complex. These findings provide new structural insights into the pH-dependent substrate binding specificity in BGs. PMID:26160737

  12. Short-time dynamics of pH-dependent conformation and substrate binding in the active site of beta-glucosidases: A computational study.

    PubMed

    Flannelly, David F; Aoki, Thalia G; Aristilde, Ludmilla

    2015-09-01

    The complete degradation of cellulose to glucose is essential to carbon turnover in terrestrial ecosystems and to engineered biofuel production. A rate-limiting step in this pathway is catalyzed by beta-glucosidase (BG) enzymes, which convert cellulobiose into two glucose molecules. The activity of these enzymes has been shown to vary with solution pH. However, it is not well understood how pH influences the enzyme conformation required for catalytic action on the substrate. A structural understanding of this pH effect is important for predicting shifts in BG activity in bioreactors and environmental matrices, in addition to informing targeted protein engineering. Here we applied molecular dynamics simulations to explore conformational and substrate binding dynamics in two well-characterized BGs of bacterial (Clostridium cellulovorans) and fungal (Trichoderma reesei) origins as a function of pH. The enzymes were simulated in an explicit solvated environment, with NaCl as electrolytes, at their prominent ionization states obtained at pH 5, 6, 7, and 7.5. Our findings indicated that pH-dependent changes in the ionization states of non-catalytic residues localized outside of the immediate active site led to pH-dependent disruption of the active site conformation. This disruption interferes with favorable H-bonding interactions with catalytic residues required to initiate catalysis on the substrate. We also identified specific non-catalytic residues that are involved in stabilizing the substrate at the optimal pH for enzyme activity. The simulations further revealed the dynamics of water-bridging interactions both outside and inside the substrate binding cleft during structural changes in the enzyme-substrate complex. These findings provide new structural insights into the pH-dependent substrate binding specificity in BGs.

  13. SufE D74R Substitution Alters Active Site Loop Dynamics To Further Enhance SufE Interaction with the SufS Cysteine Desulfurase

    PubMed Central

    Dai, Yuyuan; Kim, Dokyong; Dong, Guangchao; Busenlehner, Laura S.; Frantom, Patrick A.; Outten, F. Wayne

    2015-01-01

    Many essential metalloproteins require iron–sulfur (Fe–S) cluster cofactors for their function. In vivo persulfide formation from L-cysteine is a key step in the biogenesis of Fe–S clusters in most organisms. In Escherichia coli, the SufS cysteine desulfurase mobilizes persulfide from L-cysteine via a PLP-dependent ping-pong reaction. SufS requires the SufE partner protein to transfer the persulfide to the SufB Fe–S cluster scaffold. Without SufE, the SufS enzyme fails to efficiently turn over and remains locked in the persulfide-bound state. Coordinated protein–protein interactions mediate sulfur transfer from SufS to SufE. Multiple studies have suggested that SufE must undergo a conformational change to extend its active site Cys loop during sulfur transfer from SufS. To test this putative model, we mutated SufE Asp74 to Arg (D74R) to increase the dynamics of the SufE Cys51 loop. Amide hydrogen/deuterium exchange mass spectrometry (HDX-MS) analysis of SufE D74R revealed an increase in solvent accessibility and dynamics in the loop containing the active site Cys51 used to accept persulfide from SufS. Our results indicate that the mutant protein has a stronger binding affinity for SufS than that of wild-type SufE. In addition, SufE D74R can still enhance SufS desulfurase activity and did not show saturation at higher SufE D74R concentrations, unlike wild-type SufE. These results show that dynamic changes may shift SufE to a sulfur-acceptor state that interacts more strongly with SufS. PMID:26171726

  14. SufE D74R Substitution Alters Active Site Loop Dynamics To Further Enhance SufE Interaction with the SufS Cysteine Desulfurase.

    PubMed

    Dai, Yuyuan; Kim, Dokyong; Dong, Guangchao; Busenlehner, Laura S; Frantom, Patrick A; Outten, F Wayne

    2015-08-11

    Many essential metalloproteins require iron-sulfur (Fe-S) cluster cofactors for their function. In vivo persulfide formation from l-cysteine is a key step in the biogenesis of Fe-S clusters in most organisms. In Escherichia coli, the SufS cysteine desulfurase mobilizes persulfide from l-cysteine via a PLP-dependent ping-pong reaction. SufS requires the SufE partner protein to transfer the persulfide to the SufB Fe-S cluster scaffold. Without SufE, the SufS enzyme fails to efficiently turn over and remains locked in the persulfide-bound state. Coordinated protein-protein interactions mediate sulfur transfer from SufS to SufE. Multiple studies have suggested that SufE must undergo a conformational change to extend its active site Cys loop during sulfur transfer from SufS. To test this putative model, we mutated SufE Asp74 to Arg (D74R) to increase the dynamics of the SufE Cys51 loop. Amide hydrogen/deuterium exchange mass spectrometry (HDX-MS) analysis of SufE D74R revealed an increase in solvent accessibility and dynamics in the loop containing the active site Cys51 used to accept persulfide from SufS. Our results indicate that the mutant protein has a stronger binding affinity for SufS than that of wild-type SufE. In addition, SufE D74R can still enhance SufS desulfurase activity and did not show saturation at higher SufE D74R concentrations, unlike wild-type SufE. These results show that dynamic changes may shift SufE to a sulfur-acceptor state that interacts more strongly with SufS.

  15. Assessing hypotheses about nesting site occupancy dynamics

    USGS Publications Warehouse

    Bled, Florent; Royle, J. Andrew; Cam, Emmanuelle

    2011-01-01

    Hypotheses about habitat selection developed in the evolutionary ecology framework assume that individuals, under some conditions, select breeding habitat based on expected fitness in different habitat. The relationship between habitat quality and fitness may be reflected by breeding success of individuals, which may in turn be used to assess habitat quality. Habitat quality may also be assessed via local density: if high-quality sites are preferentially used, high density may reflect high-quality habitat. Here we assessed whether site occupancy dynamics vary with site surrogates for habitat quality. We modeled nest site use probability in a seabird subcolony (the Black-legged Kittiwake, Rissa tridactyla) over a 20-year period. We estimated site persistence (an occupied site remains occupied from time t to t + 1) and colonization through two subprocesses: first colonization (site creation at the timescale of the study) and recolonization (a site is colonized again after being deserted). Our model explicitly incorporated site-specific and neighboring breeding success and conspecific density in the neighborhood. Our results provided evidence that reproductively "successful'' sites have a higher persistence probability than "unsuccessful'' ones. Analyses of site fidelity in marked birds and of survival probability showed that high site persistence predominantly reflects site fidelity, not immediate colonization by new owners after emigration or death of previous owners. There is a negative quadratic relationship between local density and persistence probability. First colonization probability decreases with density, whereas recolonization probability is constant. This highlights the importance of distinguishing initial colonization and recolonization to understand site occupancy. All dynamics varied positively with neighboring breeding success. We found evidence of a positive interaction between site-specific and neighboring breeding success. We addressed local

  16. An Insight into the Environmental Effects of the Pocket of the Active Site of the Enzyme. Ab initio ONIOM-Molecular Dynamics (MD) Study on Cytosine Deaminase

    SciTech Connect

    Matsubara, Toshiaki; Dupuis, Michel; Aida, Misako

    2008-02-01

    We applied the ONIOM-molecular dynamics (MD) method to cytosine deaminase to examine the environmental effects of the amino acid residues in the pocket of the active site on the substrate taking account of their thermal motion. The ab initio ONIOM-MD simulations show that the substrate uracil is strongly perturbed by the amino acid residue Ile33, which sandwiches the uracil with His62, through the steric contact due to the thermal motion. As a result, the magnitude of the thermal oscillation of the potential energy and structure of the substrate uracil significantly increases. TM and MA were partly supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan.MD was supported by the Division of Chemical Sciences, Office of Basic Energy Sciences, and by the Office of Biological and Environmental Research of the U.S. Department of Energy DOE. Battelle operates Pacific Northwest National Laboratory for DOE.

  17. The role of short-range Cys171-Cys178 disulfide bond in maintaining cutinase active site integrity: A molecular dynamics simulation

    SciTech Connect

    Matak, Mehdi Youssefi; Moghaddam, Majid Erfani

    2009-12-11

    Understanding structural determinants in enzyme active site integrity can provide a good knowledge to design efficient novel catalytic machineries. Fusarium solani pisi cutinase with classic triad Ser-His-Asp is a promising enzyme to scrutinize these structural determinants. We performed two MD simulations: one, with the native structure, and the other with the broken Cys171-Cys178 disulfide bond. This disulfide bond stabilizes a turn in active site on which catalytic Asp175 is located. Functionally important H-bonds and atomic fluctuations in catalytic pocket have been changed. We proposed that this disulfide bond within active site can be considered as an important determinant of cutinase active site structural integrity.

  18. Crustal Dynamics Project: Catalogue of site information

    NASA Technical Reports Server (NTRS)

    Noll, Carey E. (Editor)

    1988-01-01

    This document represents a catalog of site information for the Crustal Dynamics Project. It contains information on and descriptions of those sites used by the Project as observing stations for making the precise geodetic measurements necessary for studies of the Earth's crustal movements and deformation.

  19. Crustal Dynamics Project: Catalogue of site information

    NASA Technical Reports Server (NTRS)

    1985-01-01

    This document represents a catalogue of site information for the Crustal Dynamics Project. It contains information and descriptions of those sites used by the Project as observing stations for making the precise geodetic measurements useful for studies of the Earth's crustal movements and deformation.

  20. On the dynamical nature of the active center in a single-site photocatalyst visualized by 4D ultrafast electron microscopy

    PubMed Central

    Yoo, Byung-Kuk; Su, Zixue; Thomas, John Meurig; Zewail, Ahmed H.

    2016-01-01

    Understanding the dynamical nature of the catalytic active site embedded in complex systems at the atomic level is critical to developing efficient photocatalytic materials. Here, we report, using 4D ultrafast electron microscopy, the spatiotemporal behaviors of titanium and oxygen in a titanosilicate catalytic material. The observed changes in Bragg diffraction intensity with time at the specific lattice planes, and with a tilted geometry, provide the relaxation pathway: the Ti4+=O2− double bond transformation to a Ti3+−O1− single bond via the individual atomic displacements of the titanium and the apical oxygen. The dilation of the double bond is up to 0.8 Å and occurs on the femtosecond time scale. These findings suggest the direct catalytic involvement of the Ti3+−O1− local structure, the significance of nonthermal processes at the reactive site, and the efficient photo-induced electron transfer that plays a pivotal role in many photocatalytic reactions. PMID:26729878

  1. Kinetic isotope effects for concerted multiple proton transfer: a direct dynamics study of an active-site model of carbonic anhydrase II.

    PubMed

    Smedarchina, Zorka; Siebrand, Willem; Fernández-Ramos, Antonio; Cui, Qiang

    2003-01-01

    The rate constant of the reaction catalyzed by the enzyme carbonic anhydrase II, which removes carbon dioxide from body fluids, is calculated for a model of the active site. The rate-determining step is proton transfer from a zinc-bound water molecule to a histidine residue via a bridge of two or more water molecules. The structure of the active site is known from X-ray studies except for the number and location of the water molecules. Model calculations are reported for a system of 58 atoms including a four-coordinated zinc ion connected to a methylimidazole molecule by a chain of two waters, constrained to reproduce the size of the active site. The structure and vibrational force field are calculated by an approximate density functional treatment of the proton-transfer step at the Self-Consistent-Charge Density Functional Tight Binding (SCC-DFTB) level. A single transition state is found indicating concerted triple proton transfer. Direct-dynamics calculations for proton and deuteron transfer and combinations thereof, based on the Approximate Instanton Method and on Variational Transition State Theory with Tunneling Corrections, are in fair agreement and yield rates that are considerably higher and kinetic isotope effects (KIEs) that are somewhat higher than experiment. Classical rate constants obtained from Transition State Theory are smaller than the quantum values but the corresponding KIEs are five times larger. For multiple proton transfer along water bridges classical KIEs are shown to be generally larger than quantum KIEs, which invalidates the standard method to distinguish tunneling and over-barrier transfer. In the present case, a three-way comparison of classical and quantum results with the observed data is necessary to conclude that proton transfer along the bridge proceeds by tunneling. The results suggest that the two-water bridge is present in low concentrations but makes a substantial contribution to proton transport because of its high

  2. Local site effects and dynamic soil behavior

    USGS Publications Warehouse

    Afak, E.

    2001-01-01

    Amplitudes of seismic waves increase significantly as they pass through soft soil layers near the earth's surface. This phenomenon, commonly known as site amplification, is a major factor influencing the extent of damage on structures. It is crucial that site amplification is accounted for when designing structures on soft soils. The characteristics of site amplification at a given site can be estimated by analytical models, as well as field tests. Analytical models require as inputs the geometry of all soil layers from surface to bedrock, their dynamic properties (e.g. density, wave velocity, damping), and the incident bedrock motions. Field tests involve recording and analyzing the dynamic response of sites to artificial excitations, ambient forces, and actual earthquakes. The most reliable estimates of site amplification are obtained by analyzing the recorded motions of the site during strong earthquakes. This paper presents a review of the types and the generating mechanisms of site amplification, and the models and methods that are used to characterize them from earthquake records. ?? 2001 Published by Elsevier Science Ltd.

  3. Salt site performance assessment activities

    SciTech Connect

    Kircher, J.F.; Gupta, S.K.

    1983-01-01

    During this year the first selection of the tools (codes) for performance assessments of potential salt sites have been tentatively selected and documented; the emphasis has shifted from code development to applications. During this period prior to detailed characterization of a salt site, the focus is on bounding calculations, sensitivity and with the data available. The development and application of improved methods for sensitivity and uncertainty analysis is a focus for the coming years activities and the subject of a following paper in these proceedings. Although the assessments to date are preliminary and based on admittedly scant data, the results indicate that suitable salt sites can be identified and repository subsystems designed which will meet the established criteria for protecting the health and safety of the public. 36 references, 5 figures, 2 tables.

  4. Crustal dynamics project site selection criteria

    NASA Technical Reports Server (NTRS)

    Allenby, R.

    1983-01-01

    The criteria for selecting site locations and constructing observing pads and monuments for the Mobile VLB1 and the satellite laser ranging systems used in the NASA/GSFC Crustal Dynamics Project are discussed. Gross system characteristics (size, shape, weight, power requirement, foot prints, etc.) are given for the Moblas, MV-1 through 3, TLRS-1 through 4 and Series instruments.

  5. Tracking dynamic team activity

    SciTech Connect

    Tambe, M.

    1996-12-31

    AI researchers are striving to build complex multi-agent worlds with intended applications ranging from the RoboCup robotic soccer tournaments, to interactive virtual theatre, to large-scale real-world battlefield simulations. Agent tracking - monitoring other agent`s actions and inferring their higher-level goals and intentions - is a central requirement in such worlds. While previous work has mostly focused on tracking individual agents, this paper goes beyond by focusing on agent teams. Team tracking poses the challenge of tracking a team`s joint goals and plans. Dynamic, real-time environments add to the challenge, as ambiguities have to be resolved in real-time. The central hypothesis underlying the present work is that an explicit team-oriented perspective enables effective team tracking. This hypothesis is instantiated using the model tracing technology employed in tracking individual agents. Thus, to track team activities, team models are put to service. Team models are a concrete application of the joint intentions framework and enable an agent to track team activities, regardless of the agent`s being a collaborative participant or a non-participant in the team. To facilitate real-time ambiguity resolution with team models: (i) aspects of tracking are cast as constraint satisfaction problems to exploit constraint propagation techniques; and (ii) a cost minimality criterion is applied to constrain tracking search. Empirical results from two separate tasks in real-world, dynamic environments one collaborative and one competitive - are provided.

  6. Lipid biomarkers and bacterial lipase activities as indicators of organic matter and bacterial dynamics in contrasted regimes at the DYFAMED site, NW Mediterranean

    NASA Astrophysics Data System (ADS)

    Bourguet, Nicolas; Goutx, Madeleine; Ghiglione, Jean-François; Pujo-Pay, Mireille; Mével, Geneviève; Momzikoff, André; Mousseau, Laure; Guigue, Catherine; Garcia, Nicole; Raimbault, Patrick; Pete, Romain; Oriol, Louise; Lefèvre, Dominique

    2009-08-01

    This study investigated the relationships between dissolved organic matter (DOM) composition and bacterial dynamics on short time scale during spring mesotrophic (March 2003) and summer oligotrophic (June 2003) regimes, in a 0-500 m depth water column with almost no advection, at the DYFAMED site, NW Mediterranean. DOM was characterized by analyzing dissolved organic carbon (DOC), colored dissolved organic matter (CDOM) and lipid class biotracers. Bacterial dynamic was assessed through the measurement of in situ bacterial lipase activity, abundance, production and bacterial community structure. We made the assumption that by coupling the ambient concentration of hydrolysable acyl-lipids with the measurement of their in situ bacterial hydrolysis rates (i.e. the free fatty acids release rate) would provide new insights about bacterial response to change in environmental conditions. The seasonal transition from spring to summer was accompanied by a significant accumulation of excess DOC (+5 μM) (ANOVA, p<0.05, n=8) in the upper layer (0-50 m). In this layer, the free fatty acids release rate to the bacterial carbon demand (BCD) ratio increased from 0.6±0.3 in March to 1.3±1.0 in June (ANOVA, p<0.05, n=8) showing that more uncoupling between the hydrolysis of the acyl-lipids and the BCD occurred during the evolution of the season, and that free fatty acids contributed to the excess DOC. The increase of lipolysis index and CDOM absorbance (from 0.24±0.17 to 0.39±0.13 and from 0.076±0.039 to 0.144±0.068; ANOVA, p<0.05, n=8, respectively), and the higher contribution of triglycerides, wax esters and phospholipids (from <5% to 12-31%) to the lipid pool reflected the change in the DOM quality. In addition to a strong increase of bacterial lipase activity per cell (51.4±29.4-418.3±290.6 Ag C cell -1 h -1), a significant percentage of ribotypes (39%) was different between spring and summer in the deep chlorophyll maximum (DCM) layer in particular, suggesting a shift

  7. The Role of the β5-α11 Loop in the Active-Site Dynamics of Acylated Penicillin-Binding Protein A from Mycobacterium tuberculosis

    SciTech Connect

    Fedarovich, Alena; Nicholas, Robert A.; Davies, Christopher

    2013-04-22

    Penicillin-binding protein A (PBPA) is a class B penicillin-binding protein that is important for cell division in Mycobacterium tuberculosis. We have determined a second crystal structure of PBPA in apo form and compared it with an earlier structure of apoenzyme. Significant structural differences in the active site region are apparent, including increased ordering of a β-hairpin loop and a shift of the SxN active site motif such that it now occupies a position that appears catalytically competent. Using two assays, including one that uses the intrinsic fluorescence of a tryptophan residue, we have also measured the second-order acylation rate constants for the antibiotics imipenem, penicillin G, and ceftriaxone. Of these, imipenem, which has demonstrable anti-tubercular activity, shows the highest acylation efficiency. Crystal structures of PBPA in complex with the same antibiotics were also determined, and all show conformational differences in the β5–α11 loop near the active site, but these differ for each β-lactam and also for each of the two molecules in the crystallographic asymmetric unit. Overall, these data reveal the β5–α11 loop of PBPA as a flexible region that appears important for acylation and provide further evidence that penicillin-binding proteins in apo form can occupy different conformational states.

  8. Characterization of active sites in zeolite catalysts

    SciTech Connect

    Eckert, J.; Bug, A.; Nicol, J.M.

    1997-11-01

    This is the final report of a three-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). Atomic-level details of the interaction of adsorbed molecules with active sites in catalysts are urgently needed to facilitate development of more effective and/or environmentally benign catalysts. To this end the authors have carried out neutron scattering studies combined with theoretical calculations of the dynamics of small molecules inside the cavities of zeolite catalysts. The authors have developed the use of H{sub 2} as a probe of adsorption sites by observing the hindered rotations of the adsorbed H{sub 2} molecule, and they were able to show that an area near the four-rings is the most likely adsorption site for H{sub 2} in zeolite A while adsorption of H{sub 2} near cations located on six-ring sites decreases in strength as Ni {approximately} Co > Ca > Zn {approximately} Na. Vibrational and rotational motions of ethylene and cyclopropane adsorption complexes were used as a measure for zeolite-adsorbate interactions. Preliminary studies of the binding of water, ammonia, and methylamines were carried out in a number of related guest-host materials.

  9. Catalysis: Elusive active site in focus

    NASA Astrophysics Data System (ADS)

    Labinger, Jay A.

    2016-08-01

    The identification of the active site of an iron-containing catalyst raises hopes of designing practically useful catalysts for the room-temperature conversion of methane to methanol, a potential fuel for vehicles. See Letter p.317

  10. A small ribozyme with dual-site kinase activity

    PubMed Central

    Biondi, Elisa; Maxwell, Adam W.R.; Burke, Donald H.

    2012-01-01

    Phosphoryl transfer onto backbone hydroxyls is a recognized catalytic activity of nucleic acids. We find that kinase ribozyme K28 possesses an unusually complex active site that promotes (thio)phosphorylation of two residues widely separated in primary sequence. After allowing the ribozyme to radiolabel itself by phosphoryl transfer from [γ-32P]GTP, DNAzyme-mediated cleavage yielded two radiolabeled cleavage fragments, indicating phosphorylation sites within each of the two cleavage fragments. These sites were mapped by alkaline digestion and primer extension pausing. Enzymatic digestion and mutational analysis identified nucleotides important for activity and established the active structure as being a constrained pseudoknot with unusual connectivity that may juxtapose the two reactive sites. Nuclease sensitivities for nucleotides near the pseudoknot core were altered in the presence of GTPγS, indicating donor-induced folding. The 5′ target site was more strongly favored in full-length ribozyme K28 (128 nt) than in truncated RNAs (58 nt). Electrophoretic mobilities of self-thiophosphorylated products on organomercurial gels are distinct from the 5′ mono-thiophosphorylated product produced by reaction with polynucleotide kinase, potentially indicating simultaneous labeling of both sites within individual RNA strands. Our evidence supports a single, compact structure with local dynamics, rather than global rearrangement, as being responsible for dual-site phosphorylation. PMID:22618879

  11. Low dielectric response in enzyme active site

    PubMed Central

    Mertz, Edward L.; Krishtalik, Lev I.

    2000-01-01

    The kinetics of charge transfer depend crucially on the dielectric reorganization of the medium. In enzymatic reactions that involve charge transfer, atomic dielectric response of the active site and of its surroundings determines the efficiency of the protein as a catalyst. We report direct spectroscopic measurements of the reorganization energy associated with the dielectric response in the active site of α-chymotrypsin. A chromophoric inhibitor of the enzyme is used as a spectroscopic probe. We find that water strongly affects the dielectric reorganization in the active site of the enzyme in solution. The reorganization energy of the protein matrix in the vicinity of the active site is similar to that of low-polarity solvents. Surprisingly, water exhibits an anomalously high dielectric response that cannot be described in terms of the dielectric continuum theory. As a result, sequestering the active site from the aqueous environment inside low-dielectric enzyme body dramatically reduces the dielectric reorganization. This reduction is particularly important for controlling the rate of enzymatic reactions. PMID:10681440

  12. Dynamics of Transcription Factor Binding Site Evolution

    PubMed Central

    Tuğrul, Murat; Paixão, Tiago; Barton, Nicholas H.; Tkačik, Gašper

    2015-01-01

    Evolution of gene regulation is crucial for our understanding of the phenotypic differences between species, populations and individuals. Sequence-specific binding of transcription factors to the regulatory regions on the DNA is a key regulatory mechanism that determines gene expression and hence heritable phenotypic variation. We use a biophysical model for directional selection on gene expression to estimate the rates of gain and loss of transcription factor binding sites (TFBS) in finite populations under both point and insertion/deletion mutations. Our results show that these rates are typically slow for a single TFBS in an isolated DNA region, unless the selection is extremely strong. These rates decrease drastically with increasing TFBS length or increasingly specific protein-DNA interactions, making the evolution of sites longer than ∼ 10 bp unlikely on typical eukaryotic speciation timescales. Similarly, evolution converges to the stationary distribution of binding sequences very slowly, making the equilibrium assumption questionable. The availability of longer regulatory sequences in which multiple binding sites can evolve simultaneously, the presence of “pre-sites” or partially decayed old sites in the initial sequence, and biophysical cooperativity between transcription factors, can all facilitate gain of TFBS and reconcile theoretical calculations with timescales inferred from comparative genomics. PMID:26545200

  13. Water in the Active Site of Ketosteroid Isomerase

    PubMed Central

    Hanoian, Philip; Hammes-Schiffer, Sharon

    2011-01-01

    Classical molecular dynamics simulations were utilized to investigate the structural and dynamical properties of water in the active site of ketosteroid isomerase (KSI) to provide insight into the role of these water molecules in the enzyme-catalyzed reaction. This reaction is thought to proceed via a dienolate intermediate that is stabilized by hydrogen bonding with residues Tyr16 and Asp103. A comparative study was performed for the wild-type (WT) KSI and the Y16F, Y16S, and Y16F/Y32F/Y57F (FFF) mutants. These systems were studied with three different bound ligands: equilenin, which is an intermediate analog, and the intermediate states of two steroid substrates. Several distinct water occupation sites were identified in the active site of KSI for the WT and mutant systems. Three additional sites were identified in the Y16S mutant that were not occupied in WT KSI or the other mutants studied. The number of water molecules directly hydrogen bonded to the ligand oxygen was approximately two waters in the Y16S mutant, one water in the Y16F and FFF mutants, and intermittent hydrogen bonding of one water molecule in WT KSI. The molecular dynamics trajectories of the Y16F and FFF mutants reproduced the small conformational changes of residue 16 observed in the crystal structures of these two mutants. Quantum mechanical/molecular mechanical calculations of 1H NMR chemical shifts of the protons in the active site hydrogen-bonding network suggest that the presence of water in the active site does not prevent the formation of short hydrogen bonds with far-downfield chemical shifts. The molecular dynamics simulations indicate that the active site water molecules exchange much more frequently for WT KSI and the FFF mutant than for the Y16F and Y16S mutants. This difference is most likely due to the hydrogen-bonding interaction between Tyr57 and an active site water molecule that is persistent in the Y16F and Y16S mutants but absent in the FFF mutant and significantly less

  14. Energy transfer at the active sites of heme proteins

    SciTech Connect

    Dlott, D.D.; Hill, J.R.

    1995-12-31

    Experiments using a picosecond pump-probe apparatus at the Picosecond Free-electron Laser Center at Stanford University, were performed to investigate the relaxation of carbon monoxide bound to the active sites of heme proteins. The significance of these experiments is two-fold: (1) they provide detailed information about molecular dynamics occurring at the active sites of proteins; and (2) they provide insight into the nature of vibrational relaxation processes in condensed matter. Molecular engineering is used to construct various molecular systems which are studied with the FEL. We have studied native proteins, mainly myoglobin obtained from different species, mutant proteins produced by genetic engineering using recombinant DNA techniques, and a variety of model systems which mimic the structures of the active sites of native proteins, which are produced using molecular synthesis. Use of these different systems permits us to investigate how specific molecular structural changes affect dynamical processes occurring at the active sites. This research provides insight into the problems of how different species needs are fulfilled by heme proteins which have greatly different functionality, which is induced by rather small structural changes.

  15. Concurrent cooperativity and substrate inhibition in the epoxidation of carbamazepine by cytochrome P450 3A4 active site mutants inspired by molecular dynamics simulations.

    PubMed

    Müller, Christian S; Knehans, Tim; Davydov, Dmitri R; Bounds, Patricia L; von Mandach, Ursula; Halpert, James R; Caflisch, Amedeo; Koppenol, Willem H

    2015-01-27

    Cytochrome P450 3A4 (CYP3A4) is the major human P450 responsible for the metabolism of carbamazepine (CBZ). To explore the mechanisms of interactions of CYP3A4 with this anticonvulsive drug, we carried out multiple molecular dynamics (MD) simulations, starting with the complex of CYP3A4 manually docked with CBZ. On the basis of these simulations, we engineered CYP3A4 mutants I369F, I369L, A370V, and A370L, in which the productive binding orientation was expected to be stabilized, thus leading to increased turnover of CBZ to the 10,11-epoxide product. In addition, we generated CYP3A4 mutant S119A as a control construct with putative destabilization of the productive binding pose. Evaluation of the kinetics profiles of CBZ epoxidation demonstrate that CYP3A4-containing bacterial membranes (bactosomes) as well as purified CYP3A4 (wild-type and mutants I369L/F) exhibit substrate inhibition in reconstituted systems. In contrast, mutants S119A and A370V/L exhibit S-shaped profiles that are indicative of homotropic cooperativity. MD simulations with two to four CBZ molecules provide evidence that the substrate-binding pocket of CYP3A4 can accommodate more than one molecule of CBZ. Analysis of the kinetics profiles of CBZ metabolism with a model that combines the formalism of the Hill equation with an allowance for substrate inhibition demonstrates that the mechanism of interactions of CBZ with CYP3A4 involves multiple substrate-binding events (most likely three). Despite the retention of the multisite binding mechanism in the mutants, functional manifestations reveal an exquisite sensitivity to even minor structural changes in the binding pocket that are introduced by conservative substitutions such as I369F, I369L, and A370V.

  16. Oxidation of butane and butene on the (100) face of (VO) sub 2 P sub 2 O sub 7 : A dynamic view in terms of the crystallochemical model of active sites

    SciTech Connect

    Ziolkowski, J. ); Bordes, E.; Courtine, P. )

    1990-03-01

    The structure of the (100) face of (VO){sub 2}P{sub 2}O{sub 7} and its performance in the oxidation of n-butane and butenes to maleic anhydride have been analyzed in terms of the crystallochemical model of active sites (CMAS). Analysis involves the heats of adsorption of oxygen, hydrogen (as a component of OH), and water as well as the heats of their movement along the surface, which allows determination of the energetically easiest pathways of elementary steps and gives insight into the reaction dynamics. The catalyst (100) (VO){sub 2}P{sub 2}O{sub 7} is found to work in a surface-oxidized state, all cations being covered with oxygen. The active site for the direct oxidation of n-butane to maleic anhydride is found to be situated between four protruding, undersaturated oxygens (2 {times} V-O, 2 {times} P-O). The reaction is thought to be initiated by H bonding at both terminal carbons. The desorption of water and migration of surface oxygen (which produces the pairs of adjacent vacancies to be filled by O{sub 2} molecules) that constitute a substep of the concerted reoxidation seem to be rate determining. Oxidation of butenes on (100) (VO){sub 2}P{sub 2}O{sub 7} is thought to be initiated by adsorption of C{double bond}C over unsaturated oxygens. In view of the surface structure, this adsorption limits the number of active oxygens with which the hydrocarbon may interact and favors a mild and nonselective oxidation to epoxybutanes, crotonaldehyde, hydrofuran, furan, and acetaldehyde. Minor yields are expected due to difficult reoxidation and competitive adsorption. Theoretical predictions are shown to agree with experimental data.

  17. Active Site Dynamical Effects in the Hydrogen Transfer Rate-limiting Step in the Catalysis of Linoleic Acid by Soybean Lipoxygenase-1 (SLO-1): Primary and Secondary Isotope Contributions.

    PubMed

    Phatak, Prasad; Venderley, Jordan; Debrota, John; Li, Junjie; Iyengar, Srinivasan S

    2015-07-30

    Using ab initio molecular dynamics (AIMD) simulations that facilitate the treatment of rare events, we probe the active site participation in the rate-determining hydrogen transfer step in the catalytic oxidation of linoleic acid by soybean lipoxygenase-1 (SLO-1). The role of two different active site components is probed. (a) On the hydrogen atom acceptor side of the active site, the hydrogen bonding propensity between the acceptor side hydroxyl group, which is bound to the iron cofactor, and the backbone carboxyl group of isoleucine (residue number 839) is studied toward its role in promoting the hydrogen transfer event. Primary and secondary (H/D) isotope effects are also probed and a definite correlation with subtle secondary H/D isotope effects is found. With increasing average nuclear kinetic energy, the increase in transfer probability is enhanced due to the presence of the hydrogen bond between the backbone carbonyl of I839 and the acceptor oxygen. Further increase in average nuclear kinetic energy reduces the strength of this secondary hydrogen bond which leads to a deterioration in hydrogen transfer rates and finally embrances an Arrhenius-like behavior. (b) On the hydrogen atom donor side, the coupling between vibrational modes predominantly localized on the donor-side linoleic acid group and the reactive mode is probed. There appears to be a qualitative difference in the coupling between modes that belong to linoleic acid and the hydrogen transfer mode, for hydrogen and deuterium transfer. For example, the donor side secondary hydrogen atom is much more labile (by nearly a factor of 5) during deuterium transfer as compared to the case for hydrogen transfer. This appears to indicate a greater coupling between the modes belonging to the linoleic acid scaffold and the deuterium transfer mode and also provides a new rationalization for the abnormal (nonclassical) secondary isotope effect results obtained by Knapp, Rickert, and Klinman in J. Am. Chem. Soc

  18. Structural and Dynamic Features of F-recruitment Site Driven Substrate Phosphorylation by ERK2

    PubMed Central

    Piserchio, Andrea; Ramakrishan, Venkatesh; Wang, Hsin; Kaoud, Tamer S.; Arshava, Boris; Dutta, Kaushik; Dalby, Kevin N.; Ghose, Ranajeet

    2015-01-01

    The F-recruitment site (FRS) of active ERK2 binds F-site (Phe-x-Phe-Pro) sequences found downstream of the Ser/Thr phospho-acceptor on cellular substrates. Here we apply NMR methods to analyze the interaction between active ERK2 (ppERK2), and a 13-residue F-site-bearing peptide substrate derived from its cellular target, the transcription factor Elk-1. Our results provide detailed insight into previously elusive structural and dynamic features of FRS/F-site interactions and FRS-driven substrate phosphorylation. We show that substrate F-site engagement significantly quenches slow dynamics involving the ppERK2 activation-loop and the FRS. We also demonstrate that the F-site phenylalanines make critical contacts with ppERK2, in contrast to the proline whose cis-trans isomerization has no significant effect on F-site recognition by the kinase FRS. Our results support a mechanism where phosphorylation of the disordered N-terminal phospho-acceptor is facilitated by its increased productive encounters with the ppERK2 active site due to docking of the proximal F-site at the kinase FRS. PMID:26054059

  19. Active Sites Environmental Monitoring Program: Action levels

    SciTech Connect

    Ashwood, J.S.; Ashwood, T.L.

    1991-10-01

    The Active Sites Environmental Monitoring Program (ASEMP) was established at Oak Ridge National Laboratory to provide for early leak detection and to monitor performance of the active low-level waste disposal facilities in Solid Waste Storage Area (SWSA) 6 and the transuranic waste storage areas in SWSA 5 North. Early leak detection is accomplished by sampling runoff, groundwater, and perched water in burial trenches. Sample results are compared to action levels that represent background contamination by naturally occurring and fallout-derived radionuclides. 15 refs., 3 figs., 12 tabs.

  20. Site-Directed Spectroscopic Probes of Actomyosin Structural Dynamics

    PubMed Central

    Thomas, David D.; Kast, David; Korman, Vicci L.

    2010-01-01

    Spectroscopy of myosin and actin has entered a golden age. High-resolution crystal structures of isolated actin and myosin have been used to construct detailed models for the dynamic actomyosin interactions that move muscle. Improved protein mutagenesis and expression technologies have facilitated site-directed labeling with fluorescent and spin probes. Spectroscopic instrumentation has achieved impressive advances in sensitivity and resolution. Here we highlight the contributions of site-directed spectroscopic probes to understanding the structural dynamics of myosin II and its actin complexes in solution and muscle fibers. We emphasize studies that probe directly the movements of structural elements within the myosin catalytic and light-chain domains, and changes in the dynamics of both actin and myosin due to their alternating strong and weak interactions in the ATPase cycle. A moving picture emerges in which single biochemical states produce multiple structural states, and transitions between states of order and dynamic disorder power the actomyosin engine. PMID:19416073

  1. Polarizability of the active site of cytochrome c reduces the activation barrier for electron transfer

    NASA Astrophysics Data System (ADS)

    Dinpajooh, Mohammadhasan; Martin, Daniel R.; Matyushov, Dmitry V.

    2016-06-01

    Enzymes in biology’s energy chains operate with low energy input distributed through multiple electron transfer steps between protein active sites. The general challenge of biological design is how to lower the activation barrier without sacrificing a large negative reaction free energy. We show that this goal is achieved through a large polarizability of the active site. It is polarized by allowing a large number of excited states, which are populated quantum mechanically by electrostatic fluctuations of the protein and hydration water shells. This perspective is achieved by extensive mixed quantum mechanical/molecular dynamics simulations of the half reaction of reduction of cytochrome c. The barrier for electron transfer is consistently lowered by increasing the number of excited states included in the Hamiltonian of the active site diagonalized along the classical trajectory. We suggest that molecular polarizability, in addition to much studied electrostatics of permanent charges, is a key parameter to consider in order to understand how enzymes work.

  2. Polarizability of the active site of cytochrome c reduces the activation barrier for electron transfer

    PubMed Central

    Dinpajooh, Mohammadhasan; Martin, Daniel R.; Matyushov, Dmitry V.

    2016-01-01

    Enzymes in biology’s energy chains operate with low energy input distributed through multiple electron transfer steps between protein active sites. The general challenge of biological design is how to lower the activation barrier without sacrificing a large negative reaction free energy. We show that this goal is achieved through a large polarizability of the active site. It is polarized by allowing a large number of excited states, which are populated quantum mechanically by electrostatic fluctuations of the protein and hydration water shells. This perspective is achieved by extensive mixed quantum mechanical/molecular dynamics simulations of the half reaction of reduction of cytochrome c. The barrier for electron transfer is consistently lowered by increasing the number of excited states included in the Hamiltonian of the active site diagonalized along the classical trajectory. We suggest that molecular polarizability, in addition to much studied electrostatics of permanent charges, is a key parameter to consider in order to understand how enzymes work. PMID:27306204

  3. Conformational Transitions in Human AP Endonuclease 1 and Its Active Site Mutant during Abasic Site Repair†

    PubMed Central

    Kanazhevskaya, Lyubov Yu.; Koval, Vladimir V.; Zharkov, Dmitry O.; Strauss, Phyllis R.; Fedorova, Olga S.

    2010-01-01

    AP endonuclease 1 (APE 1) is a crucial enzyme of the base excision repair pathway (BER) in human cells. APE1 recognizes apurinic/apyrimidinic (AP) sites and makes a nick in the phosphodiester backbone 5′ to them. The conformational dynamics and presteady-state kinetics of wild-type APE1 and its active site mutant, Y171F-P173L-N174K, have been studied. To observe conformational transitions occurring in the APE1 molecule during the catalytic cycle, we detected intrinsic tryptophan fluorescence of the enzyme under single turnover conditions. DNA duplexes containing a natural AP site, its tetrahydrofuran analogue, or a 2′-deoxyguanosine residue in the same position were used as specific substrates or ligands. The stopped-flow experiments have revealed high flexibility of the APE1 molecule and the complexity of the catalytic process. The fluorescent traces indicate that wild-type APE1 undergoes at least four conformational transitions during the processing of abasic sites in DNA. In contrast, nonspecific interactions of APE1 with undamaged DNA can be described by a two-step kinetic scheme. Rate and equilibrium constants were extracted from the stopped-flow and fluorescence titration data for all substrates, ligands, and products. A replacement of three residues at the enzymatic active site including the replacement of tyrosine 171 with phenylalanine in the enzyme active site resulted in a 2 × 104-fold decrease in the reaction rate and reduced binding affinity. Our data indicate the important role of conformational changes in APE1 for substrate recognition and catalysis. PMID:20575528

  4. Dynamic coupling of regulated binding sites and cycling myosin heads in striated muscle.

    PubMed

    Campbell, Kenneth S

    2014-03-01

    In an activated muscle, binding sites on the thin filament and myosin heads switch frequently between different states. Because the status of the binding sites influences the status of the heads, and vice versa, the binding sites and myosin heads are dynamically coupled. The functional consequences of this coupling were investigated using MyoSim, a new computer model of muscle. MyoSim extends existing models based on Huxley-type distribution techniques by incorporating Ca(2+) activation and cooperative effects. It can also simulate arbitrary cross-bridge schemes set by the researcher. Initial calculations investigated the effects of altering the relative speeds of binding-site and cross-bridge kinetics, and of manipulating cooperative processes. Subsequent tests fitted simulated force records to experimental data recorded using permeabilized myocardial preparations. These calculations suggest that the rate of force development at maximum activation is limited by myosin cycling kinetics, whereas the rate at lower levels of activation is limited by how quickly binding sites become available. Additional tests investigated the behavior of transiently activated cells by driving simulations with experimentally recorded Ca(2+) signals. The unloaded shortening profile of a twitching myocyte could be reproduced using a model with two myosin states, cooperative activation, and strain-dependent kinetics. Collectively, these results demonstrate that dynamic coupling of binding sites and myosin heads is important for contractile function.

  5. Active site of ribulosebisphosphate carboxylase/oxygenase

    SciTech Connect

    Hartman, F.C.; Stringer, C.D.; Milanez, S.; Lee, E.H.

    1985-01-01

    Previous affinity labeling studies and comparative sequence analyses have identified two different lysines at the active site of ribulosebisphosphate carboxylase/oxygenase and have suggested their essentiality to function. The essential lysines occupy positions 166 and 329 in the Rhodospirillum rubrum enzyme and positions 175 and 334 in the spinach enzyme. Based on the pH-dependencies of inactivations of the two enzymes by trinitrobenzene sulfonate, Lys-166 (R. rubrum enzyme) exhibits a pK/sub a/ of 7.9 and Lys-334 (spinach enzyme) exhibits a pK/sub a/ of 9.0. These low pK/sub a/ values as well as the enhanced nucleophilicities of the lysyl residues argue that both are important to catalysis rather than to substrate binding. Lys-166 may correspond to the essential base that initiates catalysis and that displays a pK/sub a/ of 7.5 in the pH-curve for V/sub max//K/sub m/. Cross-linking experiments with 4,4'-diisothiocyano-2,2'-disulfonate stilbene demonstrate that the two active-site lysines are within 12 A. 50 refs., 7 figs., 1 tab.

  6. Active Sites Environmental Monitoring Program: Program plan

    SciTech Connect

    Ashwood, T.L.; Wickliff, D.S.; Morrissey, C.M.

    1992-02-01

    The Active Sites Environmental Monitoring Program (ASEMP), initiated in 1989, provides early detection and performance monitoring of transuranic (TRU) waste and active low-level waste (LLW) facilities at Oak Ridge National Laboratory (ORNL) in accordance with US Department of Energy (DOE) Order 5820.2A. Active LLW facilities in Solid Waste Storage Area (SWSA) 6 include Tumulus I and Tumulus II, the Interim Waste Management Facility (IWMF), LLW silos, high-range wells, asbestos silos, and fissile wells. The tumulus pads and IWMF are aboveground, high-strength concrete pads on which concrete vaults containing metal boxes of LLW are placed; the void space between the boxes and vaults is filled with grout. Eventually, these pads and vaults will be covered by an engineered multilayered cap. All other LLW facilities in SWSA 6 are below ground. In addition, this plan includes monitoring of the Hillcut Disposal Test Facility (HDTF) in SWSA 6, even though this facility was completed prior to the data of the DOE order. In SWSA 5 North, the TRU facilities include below-grade engineered caves, high-range wells, and unlined trenches. All samples from SWSA 6 are screened for alpha and beta activity, counted for gamma-emitting isotopes, and analyzed for tritium. In addition to these analytes, samples from SWSA 5 North are analyzed for specific transuranic elements.

  7. Conformational dynamics of bacterial and human cytoplasmic models of the ribosomal A-site.

    PubMed

    Panecka, Joanna; Šponer, Jiří; Trylska, Joanna

    2015-05-01

    The aminoacyl-tRNA binding site (A-site) is located in helix 44 of small ribosomal subunit. The mobile adenines 1492 and 1493 (Escherichia coli numbering), forming the A-site bulge, act as a functional switch that ensures mRNA decoding accuracy. Structural data on the oligonucleotide models mimicking the ribosomal A-site with sequences corresponding to bacterial and human cytoplasmic sites confirm that this RNA motif forms also without the ribosome context. We performed all-atom molecular dynamics simulations of these crystallographic A-site models to compare their conformational properties. We found that the human A-site bulge is more internally flexible than the bacterial one and has different base pairing preferences, which result in the overall different shapes of these bulges and cation density distributions. Also, in the human A-site model we observed repetitive destacking of A1492, while A1493 was more stably paired than in the bacterial variant. Based on the dynamics of the A-sites we suggest why aminoglycoside antibiotics, which target the bacterial A-site, have lower binding affinities and anti-translational activities toward the human variant. PMID:25748164

  8. Highly Dense Isolated Metal Atom Catalytic Sites: Dynamic Formation and In Situ Observations.

    PubMed

    Chen, Yaxin; Kasama, Takeshi; Huang, Zhiwei; Hu, Pingping; Chen, Jianmin; Liu, Xi; Tang, Xingfu

    2015-11-23

    Atomically dispersed noble-metal catalysts with highly dense active sites are promising materials with which to maximise metal efficiency and to enhance catalytic performance; however, their fabrication remains challenging because metal atoms are prone to sintering, especially at a high metal loading. A dynamic process of formation of isolated metal atom catalytic sites on the surface of the support, which was achieved starting from silver nanoparticles by using a thermal surface-mediated diffusion method, was observed directly by using in situ electron microscopy and in situ synchrotron X-ray diffraction. A combination of electron microscopy images with X-ray absorption spectra demonstrated that the silver atoms were anchored on five-fold oxygen-terminated cavities on the surface of the support to form highly dense isolated metal active sites, leading to excellent reactivity in catalytic oxidation at low temperature. This work provides a general strategy for designing atomically dispersed noble-metal catalysts with highly dense active sites.

  9. Metal active site elasticity linked to activation of homocysteine in methionine synthases

    SciTech Connect

    Koutmos, Markos; Pejchal, Robert; Bomer, Theresa M.; Matthews, Rowena G.; Smith, Janet L.; Ludwig, Martha L.

    2008-04-02

    Enzymes possessing catalytic zinc centers perform a variety of fundamental processes in nature, including methyl transfer to thiols. Cobalamin-independent (MetE) and cobalamin-dependent (MetH) methionine synthases are two such enzyme families. Although they perform the same net reaction, transfer of a methyl group from methyltetrahydrofolate to homocysteine (Hcy) to form methionine, they display markedly different catalytic strategies, modular organization, and active site zinc centers. Here we report crystal structures of zinc-replete MetE and MetH, both in the presence and absence of Hcy. Structural investigation of the catalytic zinc sites of these two methyltransferases reveals an unexpected inversion of zinc geometry upon binding of Hcy and displacement of an endogenous ligand in both enzymes. In both cases a significant movement of the zinc relative to the protein scaffold accompanies inversion. These structures provide new information on the activation of thiols by zinc-containing enzymes and have led us to propose a paradigm for the mechanism of action of the catalytic zinc sites in these and related methyltransferases. Specifically, zinc is mobile in the active sites of MetE and MetH, and its dynamic nature helps facilitate the active site conformational changes necessary for thiol activation and methyl transfer.

  10. Control of active sites in flocculation: Concept of equivalent active sites''

    SciTech Connect

    Behl, S.; Moudgil, B.M. . Dept. of Materials Science and Engineering)

    1993-12-01

    Flocculation and dispersion of solids are strong functions of the amount and conformation of the adsorbed polymer. Regions of dispersion and flocculation of solids with particular polymer molecules may be deduced from saturation adsorption data. The concept of equivalent active sites'' is proposed to explain flocculation and dispersion behavior irrespective of the amount or conformation of the adsorbed polymer. The concept has been further extended to study the selective flocculation process.

  11. Bologna in Context: A Horizontal Perspective on the Dynamics of Governance Sites for a Europe of Knowledge

    ERIC Educational Resources Information Center

    Gornitzka, Ase

    2010-01-01

    This article presents a horizontal perspective on the dynamics of governance sites currently active for the European of Knowledge and places the Bologna process in this wider European level context. It introduces two dynamics of change in political organisation: (a) institutional differentiation and specialisation and (b) the interaction between…

  12. Collective dynamics of active filament complexes

    NASA Astrophysics Data System (ADS)

    Nogucci, Hironobu; Ishihara, Shuji

    2016-05-01

    Networks of biofilaments are essential for the formation of cellular structures that support various biological functions. For the most part, previous studies have investigated the collective dynamics of rodlike biofilaments; however, the shapes of the actual subcellular components are often more elaborate. In this study, we considered an active object composed of two active filaments, which represents the progression from rodlike biofilaments to complex-shaped biofilaments. Specifically, we numerically assessed the collective behaviors of these active objects in two dimensions and observed several types of dynamics, depending on the density and the angle of the two filaments as shape parameters of the object. Among the observed collective dynamics, a moving density band that we named a "moving smectic" is introduced here for the first time. By analyzing the trajectories of individual objects and the interactions among them, this study demonstrated how interactions among active biofilaments with complex shapes could produce collective dynamics in a nontrivial manner.

  13. Dissecting the active site of a photoreceptor protein

    NASA Astrophysics Data System (ADS)

    Hoff, Wouter; Hara, Miwa; Ren, Jie; Moghadam, Farzaneh; Xie, Aihua; Kumauchi, Masato

    While enzymes are quite large molecules, functionally important chemical events are often limited to a small region of the protein: the active site. The physical and chemical properties of residues at such active sites are often strongly altered compared to the same groups dissolved in water. Understanding such effects is important for unraveling the mechanisms underlying protein function and for protein engineering, but has proven challenging. Here we report on our ongoing efforts on using photoactive yellow protein (PYP), a bacterial photoreceptor, as a model system for such effects. We will report on the following questions: How many residues affect active site properties? Are these residues in direct physical contact with the active site? Can functionally important residues be recognized in the crystal structure of a protein? What structural resolution is needed to understand active sites? What spectroscopic techniques are most informative? Which weak interactions dominate active site properties?

  14. Phosphorylation Site Dynamics of Early T-cell Receptor Signaling

    PubMed Central

    Rigbolt, Kristoffer T. G.; Hu, Bin; Hlavacek, William S.; Blagoev, Blagoy

    2014-01-01

    In adaptive immune responses, T-cell receptor (TCR) signaling impacts multiple cellular processes and results in T-cell differentiation, proliferation, and cytokine production. Although individual protein–protein interactions and phosphorylation events have been studied extensively, we lack a systems-level understanding of how these components cooperate to control signaling dynamics, especially during the crucial first seconds of stimulation. Here, we used quantitative proteomics to characterize reshaping of the T-cell phosphoproteome in response to TCR/CD28 co-stimulation, and found that diverse dynamic patterns emerge within seconds. We detected phosphorylation dynamics as early as 5 s and observed widespread regulation of key TCR signaling proteins by 30 s. Development of a computational model pointed to the presence of novel regulatory mechanisms controlling phosphorylation of sites with central roles in TCR signaling. The model was used to generate predictions suggesting unexpected roles for the phosphatase PTPN6 (SHP-1) and shortcut recruitment of the actin regulator WAS. Predictions were validated experimentally. This integration of proteomics and modeling illustrates a novel, generalizable framework for solidifying quantitative understanding of a signaling network and for elucidating missing links. PMID:25147952

  15. Mars Surveyor Project Landing Site Activities

    NASA Technical Reports Server (NTRS)

    Gulick, Virginia C.; Briggs, Geoffrey; Saunders, R. Stephen; Gilmore, Martha; Soderblom, Larry

    1999-01-01

    The Mars Surveyor Program --now a cooperative program led by NASA and CNES along with other international partners -- is underway. It has the primary science objective of furthering our understanding of the biological potential and possible biological history of Mars and has the complementary objective of improving our understanding of martian climate evolution and planetary history The missions will develop technology and acquire data necessary for eventual human Exploration. Launches of orbiters, landers and rovers will take place in 2001 and in 2003; in 2005 a complete system will be launched capable of returning samples to Earth by 2008. A key aspect of the program is the selection of landing sites. This abstract 1) reports on the status of the landing site selection process that begins with the 2001 lander mission and 2) outlines be opportunities for the Mars community to provide input into the landing site selection process.

  16. Mars Surveyor Project Landing Site Activities

    NASA Technical Reports Server (NTRS)

    Gulick, V. C.; Briggs, Geoffrey; Saunders, R. Stephen; Gilmore, Martha; Soderblom, Larry

    1999-01-01

    The Mars Surveyor Program -- now a cooperative program led by NASA and CNES along with other international partners -- is underway. It has the primary science objective of furthering our understanding of the biological potential and possible biological history of Mars and has the complementary objective of improving our understanding of martian climate evolution and planetary history. The missions will develop technology and acquire data necessary for eventual human exploration. Launches of orbiters, landers and rovers will take place in 2001 and in 2003; in 2005 a complete system will be launched capable of returning samples to Earth by 2008. A key aspect of the program is the selection of landing sites. This abstract 1) reports on the status of the landing site selection process that begins with the 2001 lander mission and 2) outlines the opportunities for the Mars community to provide input into the landing site selection process.

  17. Single Cell Analysis of Transcriptional Activation Dynamics

    PubMed Central

    Rafalska-Metcalf, Ilona U.; Powers, Sara Lawrence; Joo, Lucy M.; LeRoy, Gary; Janicki, Susan M.

    2010-01-01

    Background Gene activation is thought to occur through a series of temporally defined regulatory steps. However, this process has not been completely evaluated in single living mammalian cells. Methodology/Principal Findings To investigate the timing and coordination of gene activation events, we tracked the recruitment of GCN5 (histone acetyltransferase), RNA polymerase II, Brd2 and Brd4 (acetyl-lysine binding proteins), in relation to a VP16-transcriptional activator, to a transcription site that can be visualized in single living cells. All accumulated rapidly with the VP16 activator as did the transcribed RNA. RNA was also detected at significantly more transcription sites in cells expressing the VP16-activator compared to a p53-activator. After α-amanitin pre-treatment, the VP16-activator, GCN5, and Brd2 are still recruited to the transcription site but the chromatin does not decondense. Conclusions/Significance This study demonstrates that a strong activator can rapidly overcome the condensed chromatin structure of an inactive transcription site and supercede the expected requirement for regulatory events to proceed in a temporally defined order. Additionally, activator strength determines the number of cells in which transcription is induced as well as the extent of chromatin decondensation. As chromatin decondensation is significantly reduced after α-amanitin pre-treatment, despite the recruitment of transcriptional activation factors, this provides further evidence that transcription drives large-scale chromatin decondensation. PMID:20422051

  18. Defect dynamics in active nematics

    PubMed Central

    Giomi, Luca; Bowick, Mark J; Mishra, Prashant; Sknepnek, Rastko; Cristina Marchetti, M

    2014-01-01

    Topological defects are distinctive signatures of liquid crystals. They profoundly affect the viscoelastic behaviour of the fluid by constraining the orientational structure in a way that inevitably requires global changes not achievable with any set of local deformations. In active nematic liquid crystals, topological defects not only dictate the global structure of the director, but also act as local sources of motion, behaving as self-propelled particles. In this article, we present a detailed analytical and numerical study of the mechanics of topological defects in active nematic liquid crystals. PMID:25332389

  19. Activation of Inhibitors by Sortase Triggers Irreversible Modification of the Active Site*S

    PubMed Central

    Maresso, Anthony W.; Wu, Ruiying; Kern, Justin W.; Zhang, Rongguang; Janik, Dorota; Missiakas, Dominique M.; Duban, Mark-Eugene; Joachimiak, Andrzej; Schneewind, Olaf

    2011-01-01

    Sortases anchor surface proteins to the cell wall of Gram-positive pathogens through recognition of specific motif sequences. Loss of sortase leads to large reductions in virulence, which identifies sortase as a target for the development of antibacterials. By screening 135,625 small molecules for inhibition, we report here that aryl (β-amino)ethyl ketones inhibit sortase enzymes from staphylococci and bacilli. Inhibition of sortases occurs through an irreversible, covalent modification of their active site cysteine. Sortases specifically activate this class of molecules via β-elimination, generating a reactive olefin intermediate that covalently modifies the cysteine thiol. Analysis of the three-dimensional structure of Bacillus anthracis sortase B with and without inhibitor provides insights into the mechanism of inhibition and reveals binding pockets that can be exploited for drug discovery. PMID:17545669

  20. The bifunctional active site of s-adenosylmethionine synthetase. Roles of the active site aspartates.

    PubMed

    Taylor, J C; Markham, G D

    1999-11-12

    S-Adenosylmethionine (AdoMet) synthetase catalyzes the biosynthesis of AdoMet in a unique enzymatic reaction. Initially the sulfur of methionine displaces the intact tripolyphosphate chain (PPP(i)) from ATP, and subsequently PPP(i) is hydrolyzed to PP(i) and P(i) before product release. The crystal structure of Escherichia coli AdoMet synthetase shows that the active site contains four aspartate residues. Aspartate residues Asp-16* and Asp-271 individually provide the sole protein ligand to one of the two required Mg(2+) ions (* denotes a residue from a second subunit); aspartates Asp-118 and Asp-238* are proposed to interact with methionine. Each aspartate has been changed to an uncharged asparagine, and the metal binding residues were also changed to alanine, to assess the roles of charge and ligation ability on catalytic efficiency. The resultant enzyme variants all structurally resemble the wild type enzyme as indicated by circular dichroism spectra and are tetramers. However, all have k(cat) reductions of approximately 10(3)-fold in AdoMet synthesis, whereas the MgATP and methionine K(m) values change by less than 3- and 8-fold, respectively. In the partial reaction of PPP(i) hydrolysis, mutants of the Mg(2+) binding residues have >700-fold reduced catalytic efficiency (k(cat)/K(m)), whereas the D118N and D238*N mutants are impaired less than 35-fold. The catalytic efficiency for PPP(i) hydrolysis by Mg(2+) site mutants is improved by AdoMet, like the wild type enzyme. In contrast AdoMet reduces the catalytic efficiency for PPP(i) hydrolysis by the D118N and D238*N mutants, indicating that the events involved in AdoMet activation are hindered in these methionyl binding site mutants. Ca(2+) uniquely activates the D271A mutant enzyme to 15% of the level of Mg(2+), in contrast to the approximately 1% Ca(2+) activation of the wild type enzyme. This indicates that the Asp-271 side chain size is a discriminator between the activating ability of Ca(2+) and the

  1. Site-level model intercomparison of high latitude and high altitude soil thermal dynamics in tundra and barren landscapes

    NASA Astrophysics Data System (ADS)

    Ekici, A.; Chadburn, S.; Chaudhary, N.; Hajdu, L. H.; Marmy, A.; Peng, S.; Boike, J.; Burke, E.; Friend, A. D.; Hauck, C.; Krinner, G.; Langer, M.; Miller, P. A.; Beer, C.

    2015-07-01

    Modeling soil thermal dynamics at high latitudes and altitudes requires representations of physical processes such as snow insulation, soil freezing and thawing and subsurface conditions like soil water/ice content and soil texture. We have compared six different land models: JSBACH, ORCHIDEE, JULES, COUP, HYBRID8 and LPJ-GUESS, at four different sites with distinct cold region landscape types, to identify the importance of physical processes in capturing observed temperature dynamics in soils. The sites include alpine, high Arctic, wet polygonal tundra and non-permafrost Arctic, thus showing how a range of models can represent distinct soil temperature regimes. For all sites, snow insulation is of major importance for estimating topsoil conditions. However, soil physics is essential for the subsoil temperature dynamics and thus the active layer thicknesses. This analysis shows that land models need more realistic surface processes, such as detailed snow dynamics and moss cover with changing thickness and wetness, along with better representations of subsoil thermal dynamics.

  2. Topology and Dynamics of Active Nematic Vesicles

    PubMed Central

    Keber, Felix C.; Loiseau, Etienne; Sanchez, Tim; DeCamp, Stephen J.; Giomi, Luca; Bowick, Mark J.; Marchetti, M. Cristina; Dogic, Zvonimir; Bausch, Andreas R.

    2015-01-01

    Engineering synthetic materials that mimic the remarkable complexity of living organisms is a fundamental challenge in science and technology. We study the spatiotemporal patterns that emerge when an active nematic film of microtubules and molecular motors is encapsulated within a shape-changing lipid vesicle. Unlike in equilibrium systems, where defects are largely static structures, in active nematics defects move spontaneously and can be described as self-propelled particles. The combination of activity, topological constraints and vesicle deformability produces a myriad of dynamical states. We highlight two dynamical modes: a tunable periodic state that oscillates between two defect configurations, and shape-changing vesicles with streaming filopodia-like protrusions. These results demonstrate how biomimetic materials can be obtained when topological constraints are used to control the non-equilibrium dynamics of active matter. PMID:25190790

  3. The active site of ribulose-bisphosphate carboxylase/oxygenase

    SciTech Connect

    Hartman, F.C.

    1991-01-01

    The active site of ribulose-bisphosphate carboxylase/oxygenase requires interacting domains of adjacent, identical subunits. Most active-site residues are located within the loop regions of an eight-stranded {beta}/{alpha}-barrel which constitutes the larger C-terminal domain; additional key residues are located within a segment of the smaller N-terminal domain which partially covers the mouth of the barrel. Site-directed mutagenesis of the gene encoding the enzyme from Rhodospirillum rubrum has been used to delineate functions of active-site residues. 6 refs., 2 figs.

  4. The dynamics of zinc sites in proteins: electronic basis for coordination sphere expansion at structural sites.

    PubMed

    Daniel, A Gerard; Farrell, Nicholas P

    2014-12-01

    The functional role assumed by zinc in proteins is closely tied to the variable dynamics around its coordination sphere arising by virtue of its flexibility in bonding. Modern experimental and computational methods allow the detection and study of previously unknown features of bonding between zinc and its ligands in protein environment. These discoveries are occurring just in time as novel biological functions of zinc, which involve rather unconventional coordination trends, are emerging. In this sense coordination sphere expansion of structural zinc sites, as observed in our previous experiments, is a novel phenomenon. Here we explore the electronic and structural requirements by simulating this phenomenon in structural zinc sites using DFT computations. For this purpose, we have chosen MPW1PW91 and a mixed basis set combination as the DFT method through benchmarking, because it accurately reproduces structural parameters of experimentally characterized zinc compounds. Using appropriate models, we show that the greater ionic character of zinc coordination would allow for coordination sphere expansion if the steric and electrostatic repulsions of the ligands are attenuated properly. Importantly, through the study of electronic and structural aspects of the models used, we arrive at a comprehensive bonding model, explaining the factors that influence coordination of zinc in proteins. The proposed model along with the existing knowledge would enhance our ability to predict zinc binding sites in proteins, which is today of growing importance given the predicted enormity of the zinc proteome.

  5. Savannah River Site prioritization of transition activities

    SciTech Connect

    Finley, R.H.

    1993-11-01

    Effective management of SRS conversion from primarily a production facility to other missions (or Decontamination and Decommissioning (D&D)) requires a systematic and consistent method of prioritizing the transition activities. This report discusses the design of a prioritizing method developed to achieve systematic and consistent methods of prioritizing these activities.

  6. DOE site performance assessment activities. Radioactive Waste Technical Support Program

    SciTech Connect

    Not Available

    1990-07-01

    Information on performance assessment capabilities and activities was collected from eight DOE sites. All eight sites either currently dispose of low-level radioactive waste (LLW) or plan to dispose of LLW in the near future. A survey questionnaire was developed and sent to key individuals involved in DOE Order 5820.2A performance assessment activities at each site. The sites surveyed included: Hanford Site (Hanford), Idaho National Engineering Laboratory (INEL), Los Alamos National Laboratory (LANL), Nevada Test Site (NTS), Oak Ridge National Laboratory (ORNL), Paducah Gaseous Diffusion Plant (Paducah), Portsmouth Gaseous Diffusion Plant (Portsmouth), and Savannah River Site (SRS). The questionnaire addressed all aspects of the performance assessment process; from waste source term to dose conversion factors. This report presents the information developed from the site questionnaire and provides a comparison of site-specific performance assessment approaches, data needs, and ongoing and planned activities. All sites are engaged in completing the radioactive waste disposal facility performance assessment required by DOE Order 5820.2A. Each site has achieved various degrees of progress and have identified a set of critical needs. Within several areas, however, the sites identified common needs and questions.

  7. Safety Oversight of Decommissioning Activities at DOE Nuclear Sites

    SciTech Connect

    Zull, Lawrence M.; Yeniscavich, William

    2008-01-15

    The Defense Nuclear Facilities Safety Board (Board) is an independent federal agency established by Congress in 1988 to provide nuclear safety oversight of activities at U.S. Department of Energy (DOE) defense nuclear facilities. The activities under the Board's jurisdiction include the design, construction, startup, operation, and decommissioning of defense nuclear facilities at DOE sites. This paper reviews the Board's safety oversight of decommissioning activities at DOE sites, identifies the safety problems observed, and discusses Board initiatives to improve the safety of decommissioning activities at DOE sites. The decommissioning of former defense nuclear facilities has reduced the risk of radioactive material contamination and exposure to the public and site workers. In general, efforts to perform decommissioning work at DOE defense nuclear sites have been successful, and contractors performing decommissioning work have a good safety record. Decommissioning activities have recently been completed at sites identified for closure, including the Rocky Flats Environmental Technology Site, the Fernald Closure Project, and the Miamisburg Closure Project (the Mound site). The Rocky Flats and Fernald sites, which produced plutonium parts and uranium materials for defense needs (respectively), have been turned into wildlife refuges. The Mound site, which performed R and D activities on nuclear materials, has been converted into an industrial and technology park called the Mound Advanced Technology Center. The DOE Office of Legacy Management is responsible for the long term stewardship of these former EM sites. The Board has reviewed many decommissioning activities, and noted that there are valuable lessons learned that can benefit both DOE and the contractor. As part of its ongoing safety oversight responsibilities, the Board and its staff will continue to review the safety of DOE and contractor decommissioning activities at DOE defense nuclear sites.

  8. Temporal dynamics of biogeochemical processes at the Norman Landfill site

    NASA Astrophysics Data System (ADS)

    Arora, Bhavna; Mohanty, Binayak P.; McGuire, Jennifer T.; Cozzarelli, Isabelle M.

    2013-10-01

    The temporal variability observed in redox sensitive species in groundwater can be attributed to coupled hydrological, geochemical, and microbial processes. These controlling processes are typically nonstationary, and distributed across various time scales. Therefore, the purpose of this study is to investigate biogeochemical data sets from a municipal landfill site to identify the dominant modes of variation and determine the physical controls that become significant at different time scales. Data on hydraulic head, specific conductance, δ2H, chloride, sulfate, nitrate, and nonvolatile dissolved organic carbon were collected between 1998 and 2000 at three wells at the Norman Landfill site in Norman, OK. Wavelet analysis on this geochemical data set indicates that variations in concentrations of reactive and conservative solutes are strongly coupled to hydrologic variability (water table elevation and precipitation) at 8 month scales, and to individual eco-hydrogeologic framework (such as seasonality of vegetation, surface-groundwater dynamics) at 16 month scales. Apart from hydrologic variations, temporal variability in sulfate concentrations can be associated with different sources (FeS cycling, recharge events) and sinks (uptake by vegetation) depending on the well location and proximity to the leachate plume. Results suggest that nitrate concentrations show multiscale behavior across temporal scales for different well locations, and dominant variability in dissolved organic carbon for a closed municipal landfill can be larger than 2 years due to its decomposition and changing content. A conceptual framework that explains the variability in chemical concentrations at different time scales as a function of hydrologic processes, site-specific interactions, and/or coupled biogeochemical effects is also presented.

  9. Temporal dynamics of biogeochemical processes at the Norman Landfill site

    USGS Publications Warehouse

    Arora, Bhavna; Mohanty, Binayak P.; McGuire, Jennifer T.; Cozzarelli, Isabelle M.

    2013-01-01

    The temporal variability observed in redox sensitive species in groundwater can be attributed to coupled hydrological, geochemical, and microbial processes. These controlling processes are typically nonstationary, and distributed across various time scales. Therefore, the purpose of this study is to investigate biogeochemical data sets from a municipal landfill site to identify the dominant modes of variation and determine the physical controls that become significant at different time scales. Data on hydraulic head, specific conductance, δ2H, chloride, sulfate, nitrate, and nonvolatile dissolved organic carbon were collected between 1998 and 2000 at three wells at the Norman Landfill site in Norman, OK. Wavelet analysis on this geochemical data set indicates that variations in concentrations of reactive and conservative solutes are strongly coupled to hydrologic variability (water table elevation and precipitation) at 8 month scales, and to individual eco-hydrogeologic framework (such as seasonality of vegetation, surface-groundwater dynamics) at 16 month scales. Apart from hydrologic variations, temporal variability in sulfate concentrations can be associated with different sources (FeS cycling, recharge events) and sinks (uptake by vegetation) depending on the well location and proximity to the leachate plume. Results suggest that nitrate concentrations show multiscale behavior across temporal scales for different well locations, and dominant variability in dissolved organic carbon for a closed municipal landfill can be larger than 2 years due to its decomposition and changing content. A conceptual framework that explains the variability in chemical concentrations at different time scales as a function of hydrologic processes, site-specific interactions, and/or coupled biogeochemical effects is also presented.

  10. Mutations of fumarase that distinguish between the active site and a nearby dicarboxylic acid binding site.

    PubMed Central

    Weaver, T.; Lees, M.; Banaszak, L.

    1997-01-01

    Two mutant forms of fumarase C from E. coli have been made using PCR and recombinant DNA. The recombinant form of the protein included a histidine arm on the C-terminal facilitating purification. Based on earlier studies, two different carboxylic acid binding sites, labeled A- and B-, were observed in crystal structures of the wild type and inhibited forms of the enzyme. A histidine at each of the sites was mutated to an asparagine. H188N at the A-site resulted in a large decrease in specific activity, while the H129N mutation at the B-site had essentially no effect. From the results, we conclude that the A-site is indeed the active site, and a dual role for H188 as a potential catalytic base is proposed. Crystal structures of the two mutant proteins produced some unexpected results. Both mutations reduced the affinity for the carboxylic acids at their respective sites. The H129N mutant should be particularly useful in future kinetic studies because it sterically blocks the B-site with the carboxyamide of asparagine assuming the position of the ligand's carboxylate. In the H188N mutation at the active site, the new asparagine side chain still interacts with an active site water that appears to have moved slightly as a result of the mutation. PMID:9098893

  11. Ionizable Side Chains at Catalytic Active Sites of Enzymes

    PubMed Central

    Jimenez-Morales, David; Liang, Jie

    2012-01-01

    Catalytic active sites of enzymes of known structure can be well defined by a modern program of computational geometry. The CASTp program was used to define and measure the volume of the catalytic active sites of 573 enzymes in the Catalytic Site Atlas database. The active sites are identified as catalytic because the amino acids they contain are known to participate in the chemical reaction catalyzed by the enzyme. Acid and base side chains are reliable markers of catalytic active sites. The catalytic active sites have 4 acid and 5 base side chains, in an average volume of 1072 Å3. The number density of acid side chains is 8.3 M (in chemical units); the number density of basic side chains is 10.6 M. The catalytic active site of these enzymes is an unusual electrostatic and steric environment in which side chains and reactants are crowded together in a mixture more like an ionic liquid than an ideal infinitely dilute solution. The electrostatics and crowding of reactants and side chains seems likely to be important for catalytic function. In three types of analogous ion channels, simulation of crowded charges accounts for the main properties of selectivity measured in a wide range of solutions and concentrations. It seems wise to use mathematics designed to study interacting complex fluids when making models of the catalytic active sites of enzymes. PMID:22484856

  12. Active Polar Two-Fluid Macroscopic Dynamics

    NASA Astrophysics Data System (ADS)

    Pleiner, Harald; Svensek, Daniel; Brand, Helmut R.

    2014-03-01

    We study the dynamics of systems with a polar dynamic preferred direction. Examples include the pattern-forming growth of bacteria (in a solvent, shoals of fish (moving in water currents), flocks of birds and migrating insects (flying in windy air). Because the preferred direction only exists dynamically, but not statically, the macroscopic variable of choice is the macroscopic velocity associated with the motion of the active units. We derive the macroscopic equations for such a system and discuss novel static, reversible and irreversible cross-couplings connected to this second velocity. We find a normal mode structure quite different compared to the static descriptions, as well as linear couplings between (active) flow and e.g. densities and concentrations due to the genuine two-fluid transport derivatives. On the other hand, we get, quite similar to the static case, a direct linear relation between the stress tensor and the structure tensor. This prominent ``active'' term is responsible for many active effects, meaning that our approach can describe those effects as well. In addition, we also deal with explicitly chiral systems, which are important for many active systems. In particular, we find an active flow-induced heat current specific for the dynamic chiral polar order.

  13. Dynamic Disturbance Processes Create Dynamic Lek Site Selection in a Prairie Grouse.

    PubMed

    Hovick, Torre J; Allred, Brady W; Elmore, R Dwayne; Fuhlendorf, Samuel D; Hamilton, Robert G; Breland, Amber

    2015-01-01

    It is well understood that landscape processes can affect habitat selection patterns, movements, and species persistence. These selection patterns may be altered or even eliminated as a result of changes in disturbance regimes and a concomitant management focus on uniform, moderate disturbance across landscapes. To assess how restored landscape heterogeneity influences habitat selection patterns, we examined 21 years (1991, 1993-2012) of Greater Prairie-Chicken (Tympanuchus cupido) lek location data in tallgrass prairie with restored fire and grazing processes. Our study took place at The Nature Conservancy's Tallgrass Prairie Preserve located at the southern extent of Flint Hills in northeastern Oklahoma. We specifically addressed stability of lek locations in the context of the fire-grazing interaction, and the environmental factors influencing lek locations. We found that lek locations were dynamic in a landscape with interacting fire and grazing. While previous conservation efforts have treated leks as stable with high site fidelity in static landscapes, a majority of lek locations in our study (i.e., 65%) moved by nearly one kilometer on an annual basis in this dynamic setting. Lek sites were in elevated areas with low tree cover and low road density. Additionally, lek site selection was influenced by an interaction of fire and patch edge, indicating that in recently burned patches, leks were located near patch edges. These results suggest that dynamic and interactive processes such as fire and grazing that restore heterogeneity to grasslands do influence habitat selection patterns in prairie grouse, a phenomenon that is likely to apply throughout the Greater Prairie-Chicken's distribution when dynamic processes are restored. As conservation moves toward restoring dynamic historic disturbance patterns, it will be important that siting and planning of anthropogenic structures (e.g., wind energy, oil and gas) and management plans not view lek locations as static

  14. Dynamic Disturbance Processes Create Dynamic Lek Site Selection in a Prairie Grouse

    PubMed Central

    Hovick, Torre J.; Allred, Brady W.; Elmore, R. Dwayne; Fuhlendorf, Samuel D.; Hamilton, Robert G.; Breland, Amber

    2015-01-01

    It is well understood that landscape processes can affect habitat selection patterns, movements, and species persistence. These selection patterns may be altered or even eliminated as a result of changes in disturbance regimes and a concomitant management focus on uniform, moderate disturbance across landscapes. To assess how restored landscape heterogeneity influences habitat selection patterns, we examined 21 years (1991, 1993–2012) of Greater Prairie-Chicken (Tympanuchus cupido) lek location data in tallgrass prairie with restored fire and grazing processes. Our study took place at The Nature Conservancy’s Tallgrass Prairie Preserve located at the southern extent of Flint Hills in northeastern Oklahoma. We specifically addressed stability of lek locations in the context of the fire-grazing interaction, and the environmental factors influencing lek locations. We found that lek locations were dynamic in a landscape with interacting fire and grazing. While previous conservation efforts have treated leks as stable with high site fidelity in static landscapes, a majority of lek locations in our study (i.e., 65%) moved by nearly one kilometer on an annual basis in this dynamic setting. Lek sites were in elevated areas with low tree cover and low road density. Additionally, lek site selection was influenced by an interaction of fire and patch edge, indicating that in recently burned patches, leks were located near patch edges. These results suggest that dynamic and interactive processes such as fire and grazing that restore heterogeneity to grasslands do influence habitat selection patterns in prairie grouse, a phenomenon that is likely to apply throughout the Greater Prairie-Chicken’s distribution when dynamic processes are restored. As conservation moves toward restoring dynamic historic disturbance patterns, it will be important that siting and planning of anthropogenic structures (e.g., wind energy, oil and gas) and management plans not view lek locations as

  15. Changes in active site histidine hydrogen bonding trigger cryptochrome activation.

    PubMed

    Ganguly, Abir; Manahan, Craig C; Top, Deniz; Yee, Estella F; Lin, Changfan; Young, Michael W; Thiel, Walter; Crane, Brian R

    2016-09-01

    Cryptochrome (CRY) is the principal light sensor of the insect circadian clock. Photoreduction of the Drosophila CRY (dCRY) flavin cofactor to the anionic semiquinone (ASQ) restructures a C-terminal tail helix (CTT) that otherwise inhibits interactions with targets that include the clock protein Timeless (TIM). All-atom molecular dynamics (MD) simulations indicate that flavin reduction destabilizes the CTT, which undergoes large-scale conformational changes (the CTT release) on short (25 ns) timescales. The CTT release correlates with the conformation and protonation state of conserved His378, which resides between the CTT and the flavin cofactor. Poisson-Boltzmann calculations indicate that flavin reduction substantially increases the His378 pKa Consistent with coupling between ASQ formation and His378 protonation, dCRY displays reduced photoreduction rates with increasing pH; however, His378Asn/Arg variants show no such pH dependence. Replica-exchange MD simulations also support CTT release mediated by changes in His378 hydrogen bonding and verify other responsive regions of the protein previously identified by proteolytic sensitivity assays. His378 dCRY variants show varying abilities to light-activate TIM and undergo self-degradation in cellular assays. Surprisingly, His378Arg/Lys variants do not degrade in light despite maintaining reactivity toward TIM, thereby implicating different conformational responses in these two functions. Thus, the dCRY photosensory mechanism involves flavin photoreduction coupled to protonation of His378, whose perturbed hydrogen-bonding pattern alters the CTT and surrounding regions. PMID:27551082

  16. Doc Dynamics In The Northwestern Mediterranean Sea (dyfamed Site)

    NASA Astrophysics Data System (ADS)

    Avril, Bernard

    Dissolved Organic Carbon (DOC) distribution and dynamics are investigated at the DYFAMED site (central Ligurian Sea, NW Mediterranean) in relation to hydrologi- cal and biological contexts, using a 4-year time-series dataset (1991-1994). The DY- FAMED site is regarded as a one-dimensional station where simple hydrological mechanisms prevail and where the ecosystem is quite well understood. An average vertical profile of DOC concentration ([DOC]) indicates that maximal concentrations and variability are concentrated in the surface layers. For depth >800 m, the annual variations are on average similar to the analytical standard deviation (~2 µM). The "composite" [DOC] distribution (average distribution over a typical year, integrating about 40 monthly profiles) for surface waters (0-200 m) is closely related to hydro- logical and phytoplanktonic forcings. It exhibits summer DOC accumulation in sur- face waters, due to spring-summer stratification and successive phytoplanktonic events such as spring and summer blooms, and winter DOC removal to deeper waters, due to intense vertical mixing. The analysis of vertical [DOC] gradient at 100-m depth as a function of the integrated DOC content in the 0-100-m layer makes it possible to objectively distinguish 3 specific periods: the winter vertical mixing period, the pe- riod of stratification and spring phytoplankton bloom, and the period of stratification re-inforcement and summer-fall phytoplankton bloom. We calculate the vertical DOC fluxes to deep waters using this large dataset. The seasonal variations of the "compos- ite" [DOC] distribution in surface waters are significantly correlated to the Apparent Oxygen Utilization (AOU) distribution, but the biogeochemical significance of such a correlation is still under examination. The global significance of our local findings is presented and the role of the oceanic DOC in the global carbon cycle is emphasized, especially with respect to several current issues, such as the

  17. Characterizing tunable dynamics in an active gel

    NASA Astrophysics Data System (ADS)

    Henkin, Gil; Decamp, Stephen; Chen, Daniel; Dogic, Zvonimir

    2014-03-01

    We experimentally investigate dynamics of an active gel of bundled microtubules that is driven to far-from-equilibrium steady states by clusters of kinesin molecular motors. Upon the addition of ATP, the coordinated action of thousands of molecular motors drives this gel to an active, percolating state that persists for hours and is only limited by the stability of constituent proteins and the availability of the chemical fuel ATP. We extensively characterize how enhanced transport in emergent macroscopic flows depends on relevant molecular parameters, including ATP, motor, and depletant concentrations, microtubule concentration and length, as well as structure of the motor clusters. Our results show that the properties and dynamics of this active isotropic gel are highly tunable, suggesting that this is an ideal system for studying the behavior of active materials.

  18. Deuterium reveals the dynamics of notch activation.

    PubMed

    Raphael, Kopan

    2011-04-13

    Notch activation requires unfolding of a juxtamembrane negative regulatory domain (NRR). Tiyanont et al. (2011) analyzed the dynamics of NRR unfolding in the presence of EGTA. As predicted from the crystal structure and deletion analyses, the lin-Notch repeats unfold first, facilitating access by ADAM proteases. Surprisingly, the heterodimerization domain remains stable.

  19. Time- and Site- Resolved Dynamics in a Circuit Topological Insulator

    NASA Astrophysics Data System (ADS)

    Jia, Ningyuan; Owens, Clai; Sommer, Ariel; Schuster, David; Simon, Jonathan

    2014-05-01

    With the discovery of the quantum Hall effect and topological insulators there has been an outpouring of ideas to harness topologically knotted band-structures in the design of state-of-the art, disorder-insensitive materials. Here we demonstrate the first simultaneous site- and time- resolved measurements of a time reversal invariant topological insulator, realized in a novel RF circuit topology. In this meta-material, we induce global topology in the band structure via local braiding in a capacitor-inductor network. We observe a gapped density of states consistent with a modified Hofstadter spectrum at a flux per plaquette of ϕ = π / 2 . In-situ probes reveal spatial localization within the bulk energy-gaps, as well as de-localized edge states. Time-resolved dynamics demonstrate a splitting of localized excitations into spin-resolved edge-modes. The RF circuit paradigm is naturally compatible widely proposed non-local coupling schemes, allowing us to implement a Mobius topological insulator inaccessible to conventional materials. Combining local braiding in an RF circuit with circuit-QED techniques, provides a direct path to topologically ordered quantum phases of matter.

  20. Site-specific dissociation dynamics of H2/D2 on Ag(111) and Co(0001) and the validity of the site-averaging model.

    PubMed

    Hu, Xixi; Jiang, Bin; Xie, Daiqian; Guo, Hua

    2015-09-21

    Dissociative chemisorption of polyatomic molecules on metal surfaces involves high-dimensional dynamics, of which quantum mechanical treatments are computationally challenging. A promising reduced-dimensional approach approximates the full-dimensional dynamics by a weighted average of fixed-site results. To examine the performance of this site-averaging model, we investigate two distinct reactions, namely, hydrogen dissociation on Co(0001) and Ag(111), using accurate first principles potential energy surfaces (PESs). The former has a very low barrier of ∼0.05 eV while the latter is highly activated with a barrier of ∼1.15 eV. These two systems allow the investigation of not only site-specific dynamical behaviors but also the validity of the site-averaging model. It is found that the reactivity is not only controlled by the barrier height but also by the topography of the PES. Moreover, the agreement between the site-averaged and full-dimensional results is much better on Ag(111), though quantitative in neither system. Further quasi-classical trajectory calculations showed that the deviations can be attributed to dynamical steering effects, which are present in both reactions at all energies.

  1. Site-specific dissociation dynamics of H2/D2 on Ag(111) and Co(0001) and the validity of the site-averaging model

    NASA Astrophysics Data System (ADS)

    Hu, Xixi; Jiang, Bin; Xie, Daiqian; Guo, Hua

    2015-09-01

    Dissociative chemisorption of polyatomic molecules on metal surfaces involves high-dimensional dynamics, of which quantum mechanical treatments are computationally challenging. A promising reduced-dimensional approach approximates the full-dimensional dynamics by a weighted average of fixed-site results. To examine the performance of this site-averaging model, we investigate two distinct reactions, namely, hydrogen dissociation on Co(0001) and Ag(111), using accurate first principles potential energy surfaces (PESs). The former has a very low barrier of ˜0.05 eV while the latter is highly activated with a barrier of ˜1.15 eV. These two systems allow the investigation of not only site-specific dynamical behaviors but also the validity of the site-averaging model. It is found that the reactivity is not only controlled by the barrier height but also by the topography of the PES. Moreover, the agreement between the site-averaged and full-dimensional results is much better on Ag(111), though quantitative in neither system. Further quasi-classical trajectory calculations showed that the deviations can be attributed to dynamical steering effects, which are present in both reactions at all energies.

  2. Ultrafast excited-state charge-transfer dynamics in laccase type I copper site.

    PubMed

    Delfino, Ines; Viola, Daniele; Cerullo, Giulio; Lepore, Maria

    2015-01-01

    Femtosecond pump-probe spectroscopy was used to investigate the excited state dynamics of the T1 copper site of laccase from Pleurotus ostreatus, by exciting its 600 nm charge transfer band with a 15-fs pulse and probing over a broad range in the visible region. The decay of the pump-induced ground-state bleaching occurs in a single step and is modulated by clearly visible oscillations. Global analysis of the two-dimensional differential transmission map shows that the excited state exponentially decays with a time constant of 375 fs, thus featuring a decay rate slower than those occurring in quite all the investigated T1 copper site proteins. The ultrashort pump pulse induces a vibrational coherence in the protein, which is mainly assigned to ground state activity, as expected in a system with fast excited state decay. Vibrational features are discussed also in comparison with the traditional resonance Raman spectrum of the enzyme. The results indicate that both excited state dynamics and vibrational modes associated with the T1 Cu laccase charge transfer have main characteristics similar to those of all the T1 copper site-containing proteins. On the other hand, the differences observed for laccase from P. ostreatus further confirm the peculiar hypothesized trigonal T1 Cu site geometry. PMID:25819432

  3. Ultrafast excited-state charge-transfer dynamics in laccase type I copper site.

    PubMed

    Delfino, Ines; Viola, Daniele; Cerullo, Giulio; Lepore, Maria

    2015-01-01

    Femtosecond pump-probe spectroscopy was used to investigate the excited state dynamics of the T1 copper site of laccase from Pleurotus ostreatus, by exciting its 600 nm charge transfer band with a 15-fs pulse and probing over a broad range in the visible region. The decay of the pump-induced ground-state bleaching occurs in a single step and is modulated by clearly visible oscillations. Global analysis of the two-dimensional differential transmission map shows that the excited state exponentially decays with a time constant of 375 fs, thus featuring a decay rate slower than those occurring in quite all the investigated T1 copper site proteins. The ultrashort pump pulse induces a vibrational coherence in the protein, which is mainly assigned to ground state activity, as expected in a system with fast excited state decay. Vibrational features are discussed also in comparison with the traditional resonance Raman spectrum of the enzyme. The results indicate that both excited state dynamics and vibrational modes associated with the T1 Cu laccase charge transfer have main characteristics similar to those of all the T1 copper site-containing proteins. On the other hand, the differences observed for laccase from P. ostreatus further confirm the peculiar hypothesized trigonal T1 Cu site geometry.

  4. The dynamics of pain: Evidence for simultaneous site-specific habituation and site-nonspecific sensitization in thermal pain

    PubMed Central

    Jepma, Marieke; Jones, Matt; Wager, Tor D.

    2014-01-01

    Repeated exposure to noxious stimuli changes their painfulness, due to multiple adaptive processes in the peripheral and central nervous system. Somewhat paradoxically, repeated stimulation can produce an increase (sensitization) or a decrease (habituation) in pain. Adaptation processes may also be body-site-specific or operate across body sites, and considering this distinction may help explain the conditions under which habituation vs. sensitization occurs. To dissociate the effects of site-specific and site-nonspecific adaptation processes, we examined reported pain in 100 participants during counterbalanced sequences of noxious thermal stimulation on multiple skin sites. Analysis of pain ratings revealed two opposing sequential effects: repeated stimulations of the same skin site produced temperature-dependent habituation, whereas repeated stimulations across different sites produced sensitization. Stimulation trials were separated by ~20 seconds and sensitization was unrelated to the distance between successively stimulated sites, suggesting that neither temporal nor spatial summation occurred. To explain these effects, we propose a dynamic model with two adaptation processes, one site-specific and one site-nonspecific. The model explains 93% of the variance in the group-mean pain ratings after controlling for current stimulation temperature, with its estimated parameters showing evidence for habituation for the site-specific process and sensitization for the site-nonspecific process. The two pain-adaptation processes revealed in this study, and the ability to disentangle them, may hold keys to understanding multiple pain-regulatory mechanisms and their disturbance in chronic-pain syndromes. Perspective This article presents novel evidence for simultaneous site-specific habituation and site-nonspecific sensitization in thermal pain, which can be disentangled (and the direction and strength of each process estimated) by a dynamic model. The dissociation of site

  5. Active Site Metal Occupancy and Cyclic Di-GMP Phosphodiesterase Activity of Thermotoga maritima HD-GYP.

    PubMed

    Miner, Kyle D; Kurtz, Donald M

    2016-02-16

    HD-GYPs make up a subclass of the metal-dependent HD phosphohydrolase superfamily and catalyze conversion of cyclic di(3',5')-guanosine monophosphate (c-di-GMP) to 5'-phosphoguanylyl-(3'→5')-guanosine (pGpG) and GMP. Until now, the only reported crystal structure of an HD-GYP that also exhibits c-di-GMP phosphodiesterase activity contains a His/carboxylate ligated triiron active site. However, other structural and phylogenetic correlations indicate that some HD-GYPs contain dimetal active sites. Here we provide evidence that an HD-GYP c-di-GMP phosphodiesterase, TM0186, from Thermotoga maritima can accommodate both di- and trimetal active sites. We show that an as-isolated iron-containing TM0186 has an oxo/carboxylato-bridged diferric site, and that the reduced (diferrous) form is necessary and sufficient to catalyze conversion of c-di-GMP to pGpG, but that conversion of pGpG to GMP requires more than two metals per active site. Similar c-di-GMP phosphodiesterase activities were obtained with divalent iron or manganese. On the basis of activity correlations with several putative metal ligand residue variants and molecular dynamics simulations, we propose that TM0186 can accommodate both di- and trimetal active sites. Our results also suggest that a Glu residue conserved in a subset of HD-GYPs is required for formation of the trimetal site and can also serve as a labile ligand to the dimetal site. Given the anaerobic growth requirement of T. maritima, we suggest that this HD-GYP can function in vivo with either divalent iron or manganese occupying di- and trimetal sites.

  6. Random bursts determine dynamics of active filaments

    PubMed Central

    Weber, Christoph A.; Suzuki, Ryo; Schaller, Volker; Aranson, Igor S.; Bausch, Andreas R.; Frey, Erwin

    2015-01-01

    Constituents of living or synthetic active matter have access to a local energy supply that serves to keep the system out of thermal equilibrium. The statistical properties of such fluctuating active systems differ from those of their equilibrium counterparts. Using the actin filament gliding assay as a model, we studied how nonthermal distributions emerge in active matter. We found that the basic mechanism involves the interplay between local and random injection of energy, acting as an analog of a thermal heat bath, and nonequilibrium energy dissipation processes associated with sudden jump-like changes in the system’s dynamic variables. We show here how such a mechanism leads to a nonthermal distribution of filament curvatures with a non-Gaussian shape. The experimental curvature statistics and filament relaxation dynamics are reproduced quantitatively by stochastic computer simulations and a simple kinetic model. PMID:26261319

  7. Random bursts determine dynamics of active filaments.

    PubMed

    Weber, Christoph A; Suzuki, Ryo; Schaller, Volker; Aranson, Igor S; Bausch, Andreas R; Frey, Erwin

    2015-08-25

    Constituents of living or synthetic active matter have access to a local energy supply that serves to keep the system out of thermal equilibrium. The statistical properties of such fluctuating active systems differ from those of their equilibrium counterparts. Using the actin filament gliding assay as a model, we studied how nonthermal distributions emerge in active matter. We found that the basic mechanism involves the interplay between local and random injection of energy, acting as an analog of a thermal heat bath, and nonequilibrium energy dissipation processes associated with sudden jump-like changes in the system's dynamic variables. We show here how such a mechanism leads to a nonthermal distribution of filament curvatures with a non-Gaussian shape. The experimental curvature statistics and filament relaxation dynamics are reproduced quantitatively by stochastic computer simulations and a simple kinetic model.

  8. Active site - a site of binding of affinity inhibitors in baker's yeast inorganic pyrophosphatase

    SciTech Connect

    Svyato, I.E.; Sklyankina, V.A.; Avaeva, S.M.

    1986-03-20

    The interaction of the enzyme-substrate complex with methyl phosphate, O-phosphoethanolamine, O-phosphopropanolamine, N-acetylphosphoserine, and phosphoglyolic acid, as well as pyrophosphatase, modified by monoesters of phosphoric acid, with pyrophosphate and tripolyphosphate, was investigated. It was shown that the enzyme containing the substrate in the active site does not react with monophosphates, but modified pyrophosphatase entirely retains the ability to bind polyanions to the regulatory site. It is concluded that the inactivation of baker's yeast inorganic pyrophosphatase by monoesters of phosphoric acid, which are affinity inhibitors of it, is the result of modification of the active site of the enzyme.

  9. A novel approach to predict active sites of enzyme molecules.

    PubMed

    Chou, Kuo-Chen; Cai, Yu-dong

    2004-04-01

    Enzymes are critical in many cellular signaling cascades. With many enzyme structures being solved, there is an increasing need to develop an automated method for identifying their active sites. However, given the atomic coordinates of an enzyme molecule, how can we predict its active site? This is a vitally important problem because the core of an enzyme molecule is its active site from the viewpoints of both pure scientific research and industrial application. In this article, a topological entity was introduced to characterize the enzymatic active site. Based on such a concept, the covariant discriminant algorithm was formulated for identifying the active site. As a paradigm, the serine hydrolase family was demonstrated. The overall success rate by jackknife test for a data set of 88 enzyme molecules was 99.92%, and that for a data set of 50 independent enzyme molecules was 99.91%. Meanwhile, it was shown through an example that the prediction algorithm can also be used to find any typographic error of a PDB file in annotating the constituent amino acids of catalytic triad and to suggest a possible correction. The very high success rates are due to the introduction of a covariance matrix in the prediction algorithm that makes allowance for taking into account the coupling effects among the key constituent atoms of active site. It is anticipated that the novel approach is quite promising and may become a useful high throughput tool in enzymology, proteomics, and structural bioinformatics. PMID:14997541

  10. Dynamics of active layer in wooded palsas of northern Quebec

    NASA Astrophysics Data System (ADS)

    Jean, Mélanie; Payette, Serge

    2014-02-01

    Palsas are organic or mineral soil mounds having a permafrost core. Palsas are widespread in the circumpolar discontinuous permafrost zone. The annual dynamics and evolution of the active layer, which is the uppermost layer over the permafrost table and subjected to the annual freeze-thaw cycle, are influenced by organic layer thickness, snow depth, vegetation type, topography and exposure. This study examines the influence of vegetation types, with an emphasis on forest cover, on active layer dynamics of palsas in the Boniface River watershed (57°45‧ N, 76°00‧ W). In this area, palsas are often colonized by black spruce trees (Picea mariana (Mill.) B.S.P.). Thaw depth and active layer thickness were monitored on 11 wooded or non-wooded mineral and organic palsas in 2009, 2010 and 2011. Snow depth, organic layer thickness, and vegetation types were assessed. The mapping of a palsa covered by various vegetation types and a large range of organic layer thickness were used to identify the factors influencing the spatial patterns of thaw depth and active layer. The active layer was thinner and the thaw rate slower in wooded palsas, whereas it was the opposite in more exposed sites such as forest openings, shrubs and bare ground. Thicker organic layers were associated with thinner active layers and slower thaw rates. Snow depth was not an important factor influencing active layer dynamics. The topography of the mapped palsa was uneven, and the environmental factors such as organic layer, snow depth, and vegetation types were heterogeneously distributed. These factors explain a part of the spatial variation of the active layer. Over the 3-year long study, the area of one studied palsa decreased by 70%. In a context of widespread permafrost decay, increasing our understanding of factors that influence the dynamics of wooded and non-wooded palsas and understanding of the role of vegetation cover will help to define the response of discontinuous permafrost landforms

  11. Growth exponents in surface models with non-active sites

    NASA Astrophysics Data System (ADS)

    Santos, M.; Figueiredo, W.; Aarão Reis, F. D. A.

    2006-11-01

    In this work, we studied the role played by the inactive sites present on the substrate of a growing surface. In our model, one particle sticks at the surface if the site where it falls is an active site. However, we allow the deposited particle to diffuse along the surface in accordance with some mechanism previously defined. Using Monte Carlo simulations, and some analytical results, we have investigated the model in (1+1) and (2+1) dimensions considering different relaxation mechanisms. We show that the consideration of non-active sites is a crucial point in the model. In fact, we have seen that the saturation regime is not observed for any value of the density of inactive sites. Besides, the growth exponent β turns to be one, at long times, whatever the mechanism of diffusion we consider in one and two dimensions.

  12. Modulation of Active Site Electronic Structure by the Protein Matrix to Control [NiFe] Hydrogenase Reactivity

    SciTech Connect

    Smith, Dayle MA; Raugei, Simone; Squier, Thomas C.

    2014-09-30

    Control of the reactivity of the nickel center of the [NiFe] hydrogenase and other metalloproteins commonly involves outer coordination sphere ligands that act to modify the geometry and physical properties of the active site metal centers. We carried out a combined set of classical molecular dynamics and quantum/classical mechanics calculations to provide quantitative estimates of how dynamic fluctuations of the active site within the protein matrix modulate the electronic structure at the catalytic center. Specifically we focused on the dynamics of the inner and outer coordination spheres of the cysteinate-bound Ni–Fe cluster in the catalytically active Ni-C state. There are correlated movements of the cysteinate ligands and the surrounding hydrogen-bonding network, which modulate the electron affinity at the active site and the proton affinity of a terminal cysteinate. On the basis of these findings, we hypothesize a coupling between protein dynamics and electron and proton transfer reactions critical to dihydrogen production.

  13. Dynamic patterns of academic forum activities

    NASA Astrophysics Data System (ADS)

    Zhao, Zhi-Dan; Gao, Ya-Chun; Cai, Shi-Min; Zhou, Tao

    2016-11-01

    A mass of traces of human activities show rich dynamic patterns. In this article, we comprehensively investigate the dynamic patterns of 50 thousands of researchers' activities in Sciencenet, the largest multi-disciplinary academic community in China. Through statistical analyses, we found that (i) there exists a power-law scaling between the frequency of visits to an academic forum and the number of corresponding visitors, with the exponent being about 1.33; (ii) the expansion process of academic forums obeys the Heaps' law, namely the number of distinct visited forums to the number of visits grows in a power-law form with exponent being about 0.54; (iii) the probability distributions of time intervals and the number of visits taken to revisit the same academic forum both follow power-laws, indicating the existence of memory effect in academic forum activities. On the basis of these empirical results, we propose a dynamic model that incorporates the exploration, preferential return with memory effect, which can well reproduce the observed scaling laws.

  14. Architecture and active site of particulate methane monooxygenase

    PubMed Central

    Culpepper, Megen A.; Rosenzweig, Amy C.

    2012-01-01

    Particulate methane monooxygenase (pMMO) is an integral membrane metalloenzyme that oxidizes methane to methanol in methanotrophic bacteria, organisms that live on methane gas as their sole carbon source. Understanding pMMO function has important implications for bioremediation applications and for the development of new, environmentally friendly catalysts for the direct conversion of methane to methanol. Crystal structures of pMMOs from three different methanotrophs reveal a trimeric architecture, consisting of three copies each of the pmoB, pmoA, and pmoC subunits. There are three distinct metal centers in each protomer of the trimer, mononuclear and dinuclear copper sites in the periplasmic regions of pmoB and a mononuclear site within the membrane that can be occupied by copper or zinc. Various models for the pMMO active site have been proposed within these structural constraints, including dicopper, tricopper, and diiron centers. Biochemical and spectroscopic data on pMMO and recombinant soluble fragments, denoted spmoB proteins, indicate that the active site involves copper and is located at the site of the dicopper center in the pmoB subunit. Initial spectroscopic evidence for O2 binding at this site has been obtained. Despite these findings, questions remain about the active site identity and nuclearity and will be the focus of future studies. PMID:22725967

  15. Methanopyrus kandleri topoisomerase V contains three distinct AP lyase active sites in addition to the topoisomerase active site.

    PubMed

    Rajan, Rakhi; Osterman, Amy; Mondragón, Alfonso

    2016-04-20

    Topoisomerase V (Topo-V) is the only topoisomerase with both topoisomerase and DNA repair activities. The topoisomerase activity is conferred by a small alpha-helical domain, whereas the AP lyase activity is found in a region formed by 12 tandem helix-hairpin-helix ((HhH)2) domains. Although it was known that Topo-V has multiple repair sites, only one had been mapped. Here, we show that Topo-V has three AP lyase sites. The atomic structure and Small Angle X-ray Scattering studies of a 97 kDa fragment spanning the topoisomerase and 10 (HhH)2 domains reveal that the (HhH)2 domains extend away from the topoisomerase domain. A combination of biochemical and structural observations allow the mapping of the second repair site to the junction of the 9th and 10th (HhH)2 domains. The second site is structurally similar to the first one and to the sites found in other AP lyases. The 3rd AP lyase site is located in the 12th (HhH)2 domain. The results show that Topo-V is an unusual protein: it is the only known protein with more than one (HhH)2 domain, the only known topoisomerase with dual activities and is also unique by having three AP lyase repair sites in the same polypeptide. PMID:26908655

  16. Methanopyrus kandleri topoisomerase V contains three distinct AP lyase active sites in addition to the topoisomerase active site

    PubMed Central

    Rajan, Rakhi; Osterman, Amy; Mondragón, Alfonso

    2016-01-01

    Topoisomerase V (Topo-V) is the only topoisomerase with both topoisomerase and DNA repair activities. The topoisomerase activity is conferred by a small alpha-helical domain, whereas the AP lyase activity is found in a region formed by 12 tandem helix-hairpin-helix ((HhH)2) domains. Although it was known that Topo-V has multiple repair sites, only one had been mapped. Here, we show that Topo-V has three AP lyase sites. The atomic structure and Small Angle X-ray Scattering studies of a 97 kDa fragment spanning the topoisomerase and 10 (HhH)2 domains reveal that the (HhH)2 domains extend away from the topoisomerase domain. A combination of biochemical and structural observations allow the mapping of the second repair site to the junction of the 9th and 10th (HhH)2 domains. The second site is structurally similar to the first one and to the sites found in other AP lyases. The 3rd AP lyase site is located in the 12th (HhH)2 domain. The results show that Topo-V is an unusual protein: it is the only known protein with more than one (HhH)2 domain, the only known topoisomerase with dual activities and is also unique by having three AP lyase repair sites in the same polypeptide. PMID:26908655

  17. The Three Mycobacterium tuberculosis Antigen 85 Isoforms Have Unique Substrates and Activities Determined by Non-active Site Regions*

    PubMed Central

    Backus, Keriann M.; Dolan, Michael A.; Barry, Conor S.; Joe, Maju; McPhie, Peter; Boshoff, Helena I. M.; Lowary, Todd L.; Davis, Benjamin G.; Barry, Clifton E.

    2014-01-01

    The three isoforms of antigen 85 (A, B, and C) are the most abundant secreted mycobacterial proteins and catalyze transesterification reactions that synthesize mycolated arabinogalactan, trehalose monomycolate (TMM), and trehalose dimycolate (TDM), important constituents of the outermost layer of the cellular envelope of Mycobacterium tuberculosis. These three enzymes are nearly identical at the active site and have therefore been postulated to exist to evade host immunity. Distal to the active site is a second putative carbohydrate-binding site of lower homology. Mutagenesis of the three isoforms at this second site affected both substrate selectivity and overall catalytic activity in vitro. Using synthetic and natural substrates, we show that these three enzymes exhibit unique selectivity; antigen 85A more efficiently mycolates TMM to form TDM, whereas C (and to a lesser extent B) has a higher rate of activity using free trehalose to form TMM. This difference in substrate selectivity extends to the hexasaccharide fragment of cell wall arabinan. Mutation of secondary site residues from the most active isoform (C) into those present in A or B partially interconverts this substrate selectivity. These experiments in combination with molecular dynamics simulations reveal that differences in the N-terminal helix α9, the adjacent Pro216–Phe228 loop, and helix α5 are the likely cause of changes in activity and substrate selectivity. These differences explain the existence of three isoforms and will allow for future work in developing inhibitors. PMID:25028517

  18. Dynamics and Emergent Structures in Active Fluids

    NASA Astrophysics Data System (ADS)

    Baskaran, Aparna

    2014-03-01

    In this talk, we consider an active fluid of colloidal sized particles, with the primary manifestation of activity being a self-replenishing velocity along one body axis of the particle. This is a minimal model for varied systems such as bacterial colonies, cytoskeletal filament motility assays vibrated granular particles and self propelled diffusophoretic colloids, depending on the nature of interaction among the particles. Using microscopic Brownian dynamics simulations, coarse-graining using the tools of non-equilibrium statistical mechanics and analysis of macroscopic hydrodynamic theories, we characterize emergent structures seen in these systems, which are determined by the symmetry of the interactions among the active units, such as propagating density waves, dense stationary bands, asters and phase separated isotropic clusters. We identify a universal mechanism, termed ``self-regulation,'' as the underlying physics that leads to these structures in diverse systems. Support from NSF through DMR-1149266 and DMR-0820492.

  19. Active Sites Environmental Monitoring Program: Mid-FY 1991 report

    SciTech Connect

    Ashwood, T.L.; Wickliff, D.S.; Morrissey, C.M.

    1991-10-01

    This report summarizes the activities of the Active Sites Environmental Monitoring Program (ASEMP) from October 1990 through March 1991. The ASEMP was established in 1989 by Solid Waste Operations and the Environmental Sciences Division to provide early detection and performance monitoring at active low-level radioactive waste (LLW) disposal sites in Solid Waste Storage Area (SWSA) 6 and transuranic (TRU) waste storage sites in SWSA 5 as required by chapters II and III of US Department of Energy Order 5820.2A. Monitoring results continue to demonstrate the no LLW is being leached from the storage vaults on the tumulus pads. Loading of vaults on Tumulus II began during this reporting period and 115 vaults had been loaded by the end of March 1991.

  20. Interface dynamics explain assembly dependency of influenza neuraminidase catalytic activity

    PubMed Central

    von Grafenstein, Susanne; Wallnoefer, Hannes G.; Kirchmair, Johannes; Fuchs, Julian E.; Huber, Roland G.; Schmidtke, Michaela; Sauerbrei, Andreas; Rollinger, Judith M.; Liedl, Klaus R.

    2015-01-01

    Influenza virus neuraminidase (iNA) is a homotetrameric surface protein of the influenza virus and an established target for antiviral drugs. In contrast to neuraminidases (NAs) of other biological systems (non-iNAs), enzymatic activity of iNA is only observed in a quaternary assembly and iNA needs the tetramerization to mediate enzymatic activity. Obviously, differences on a molecular level between iNA and non-iNAs are responsible for this intriguing observation. Comparison between protein structures and multiple sequence alignment allow the identification of differences in amino acid composition in crucial regions of the enzyme, such as next to the conserved D151 and the 150-loop. These differences in amino acid sequence and protein tetramerization are likely to alter the dynamics of the system. Therefore, we performed molecular dynamics simulations to investigate differences in the molecular flexibility of monomers, dimers, and tetramers of iNAs of subtype N1 (avian 2004, pandemic 1918 and pandemic 2009 iNA) and as comparison the non-iNA monomer from Clostridium perfringens. We show that conformational transitions of iNA are crucially influenced by its assembly state. The protein–protein interface induces a complex hydrogen-bonding network between the 110-helix and the 150-loop, which consequently stabilizes the structural arrangement of the binding site. Therefore, we claim that these altered dynamics are responsible for the dependence of iNA’s catalytic activity on the tetrameric assembly. Only the tetramerization-induced balance between stabilization and altered local flexibility in the binding site provides the appropriate arrangement of key residues for iNA’s catalytic activity. PMID:24279589

  1. Active chemisorption sites in functionalized ionic liquids for carbon capture.

    PubMed

    Cui, Guokai; Wang, Jianji; Zhang, Suojiang

    2016-07-25

    Development of novel technologies for the efficient and reversible capture of CO2 is highly desired. In the last decade, CO2 capture using ionic liquids has attracted intensive attention from both academia and industry, and has been recognized as a very promising technology. Recently, a new approach has been developed for highly efficient capture of CO2 by site-containing ionic liquids through chemical interaction. This perspective review focuses on the recent advances in the chemical absorption of CO2 using site-containing ionic liquids, such as amino-based ionic liquids, azolate ionic liquids, phenolate ionic liquids, dual-functionalized ionic liquids, pyridine-containing ionic liquids and so on. Other site-containing liquid absorbents such as amine-based solutions, switchable solvents, and functionalized ionic liquid-amine blends are also investigated. Strategies have been discussed for how to activate the existent reactive sites and develop novel reactive sites by physical and chemical methods to enhance CO2 absorption capacity and reduce absorption enthalpy. The carbon capture mechanisms of these site-containing liquid absorbents are also presented. Particular attention has been paid to the latest progress in CO2 capture in multiple-site interactions by amino-free anion-functionalized ionic liquids. In the last section, future directions and prospects for carbon capture by site-containing ionic liquids are outlined.

  2. Active chemisorption sites in functionalized ionic liquids for carbon capture.

    PubMed

    Cui, Guokai; Wang, Jianji; Zhang, Suojiang

    2016-07-25

    Development of novel technologies for the efficient and reversible capture of CO2 is highly desired. In the last decade, CO2 capture using ionic liquids has attracted intensive attention from both academia and industry, and has been recognized as a very promising technology. Recently, a new approach has been developed for highly efficient capture of CO2 by site-containing ionic liquids through chemical interaction. This perspective review focuses on the recent advances in the chemical absorption of CO2 using site-containing ionic liquids, such as amino-based ionic liquids, azolate ionic liquids, phenolate ionic liquids, dual-functionalized ionic liquids, pyridine-containing ionic liquids and so on. Other site-containing liquid absorbents such as amine-based solutions, switchable solvents, and functionalized ionic liquid-amine blends are also investigated. Strategies have been discussed for how to activate the existent reactive sites and develop novel reactive sites by physical and chemical methods to enhance CO2 absorption capacity and reduce absorption enthalpy. The carbon capture mechanisms of these site-containing liquid absorbents are also presented. Particular attention has been paid to the latest progress in CO2 capture in multiple-site interactions by amino-free anion-functionalized ionic liquids. In the last section, future directions and prospects for carbon capture by site-containing ionic liquids are outlined. PMID:27243042

  3. Studies on the active site of pig plasma amine oxidase.

    PubMed Central

    Collison, D; Knowles, P F; Mabbs, F E; Rius, F X; Singh, I; Dooley, D M; Cote, C E; McGuirl, M

    1989-01-01

    Amine oxidase from pig plasma (PPAO) has two bound Cu2+ ions and at least one pyrroloquinoline quinone (PQQ) moiety as cofactors. It is shown that recovery of activity by copper-depleted PPAO is linear with respect to added Cu2+ ions. Recovery of e.s.r. and optical spectral characteristics of active-site copper parallel the recovery of catalytic activity. These results are consistent with both Cu2+ ions contributing to catalysis. Further e.s.r. studies indicate that the two copper sites in PPAO, unlike those in amine oxidases from other sources, are chemically distinct. These comparative studies establish that non-identity of the Cu2+ ions in PPAO is not a requirement for amine oxidase activity. It is shown through the use of a new assay procedure that there are two molecules of PQQ bound per molecule of protein in PPAO; only the more reactive of these PQQ moieties is required for activity. PMID:2559715

  4. CO Oxidation on Au/TiO2: Condition-Dependent Active Sites and Mechanistic Pathways.

    PubMed

    Wang, Yang-Gang; Cantu, David C; Lee, Mal-Soon; Li, Jun; Glezakou, Vassiliki-Alexandra; Rousseau, Roger

    2016-08-24

    We present results of ab initio electronic structure and molecular dynamics simulations (AIMD), as well as a microkinetic model of CO oxidation catalyzed by TiO2 supported Au nanocatalysts. A coverage-dependent microkinetic analysis, based on energetics obtained with density functional methods, shows that the dominant kinetic pathway, activated oxygen species, and catalytic active sites are all strongly depended on both temperature and oxygen partial pressure. Under oxidizing conditions and T < 400 K, the prevalent pathway involves a dynamic single atom catalytic mechanism. This reaction is catalyzed by a transient Au-CO species that migrates from the Au-cluster onto a surface oxygen adatom. It subsequently reacts with the TiO2 support via a Mars van Krevelen mechanism to form CO2 and finally the Au atom reintegrates back into the gold cluster to complete the catalytic cycle. At 300 ≤ T ≤ 600 K, oxygen-bound single Oad-Au(+)-CO sites and the perimeter Au-sites of the nanoparticle work in tandem to optimally catalyze the reaction. Above 600 K, a variety of alternate pathways associated with both single-atom and the perimeter sites of the Au nanoparticle are found to be active. Under low oxygen pressures, Oad-Au(+)-CO species can be a source of catalyst deactivation and the dominant pathway involves only Au-perimeter sites. A detailed comparison of the current model and the existing literature resolves many apparent inconsistencies in the mechanistic interpretations. PMID:27480512

  5. Enhanced Enzyme Kinetic Stability by Increasing Rigidity within the Active Site*

    PubMed Central

    Xie, Yuan; An, Jiao; Yang, Guangyu; Wu, Geng; Zhang, Yong; Cui, Li; Feng, Yan

    2014-01-01

    Enzyme stability is an important issue for protein engineers. Understanding how rigidity in the active site affects protein kinetic stability will provide new insight into enzyme stabilization. In this study, we demonstrated enhanced kinetic stability of Candida antarctica lipase B (CalB) by mutating the structurally flexible residues within the active site. Six residues within 10 Å of the catalytic Ser105 residue with a high B factor were selected for iterative saturation mutagenesis. After screening 2200 colonies, we obtained the D223G/L278M mutant, which exhibited a 13-fold increase in half-life at 48 °C and a 12 °C higher T5015, the temperature at which enzyme activity is reduced to 50% after a 15-min heat treatment. Further characterization showed that global unfolding resistance against both thermal and chemical denaturation also improved. Analysis of the crystal structures of wild-type CalB and the D223G/L278M mutant revealed that the latter formed an extra main chain hydrogen bond network with seven structurally coupled residues within the flexible α10 helix that are primarily involved in forming the active site. Further investigation of the relative B factor profile and molecular dynamics simulation confirmed that the enhanced rigidity decreased fluctuation of the active site residues at high temperature. These results indicate that enhancing the rigidity of the flexible segment within the active site may provide an efficient method for improving enzyme kinetic stability. PMID:24448805

  6. Comparative Sensitivity Analysis of Muscle Activation Dynamics.

    PubMed

    Rockenfeller, Robert; Günther, Michael; Schmitt, Syn; Götz, Thomas

    2015-01-01

    We mathematically compared two models of mammalian striated muscle activation dynamics proposed by Hatze and Zajac. Both models are representative for a broad variety of biomechanical models formulated as ordinary differential equations (ODEs). These models incorporate parameters that directly represent known physiological properties. Other parameters have been introduced to reproduce empirical observations. We used sensitivity analysis to investigate the influence of model parameters on the ODE solutions. In addition, we expanded an existing approach to treating initial conditions as parameters and to calculating second-order sensitivities. Furthermore, we used a global sensitivity analysis approach to include finite ranges of parameter values. Hence, a theoretician striving for model reduction could use the method for identifying particularly low sensitivities to detect superfluous parameters. An experimenter could use it for identifying particularly high sensitivities to improve parameter estimation. Hatze's nonlinear model incorporates some parameters to which activation dynamics is clearly more sensitive than to any parameter in Zajac's linear model. Other than Zajac's model, Hatze's model can, however, reproduce measured shifts in optimal muscle length with varied muscle activity. Accordingly we extracted a specific parameter set for Hatze's model that combines best with a particular muscle force-length relation. PMID:26417379

  7. Computer simulation of the active site of human serum cholinesterase

    SciTech Connect

    Kefang Jiao; Song Li; Zhengzheng Lu

    1996-12-31

    The first 3D-structure of acetylchelinesterase from Torpedo California electric organ (T.AChE) was published by JL. Sussman in 1991. We have simulated 3D-structure of human serum cholinesterase (H.BuChE) and the active site of H.BuChE. It is discovered by experiment that the residue of H.BuChE is still active site after a part of H.BuChE is cut. For example, the part of 21KD + 20KD is active site of H.BuChE. The 20KD as it is. Studies on these peptides by Hemelogy indicate that two active peptides have same negative electrostatic potential maps diagram. These negative electrostatic areas attached by acetyl choline with positive electrostatic potency. We predict that 147...236 peptide of AChE could be active site because it was as 20KD as with negative electrostatic potential maps. We look forward to proving from other ones.

  8. Outside-binding site mutations modify the active site's shapes in neuraminidase from influenza A H1N1.

    PubMed

    Tolentino-Lopez, Luis; Segura-Cabrera, Aldo; Reyes-Loyola, Paola; Zimic, Mirko; Quiliano, Miguel; Briz, Veronica; Muñoz-Fernández, Angeles; Rodríguez-Pérez, Mario; Ilizaliturri-Flores, Ian; Correa-Basurto, Jose

    2013-01-01

    The recent occurrence of 2009 influenza A (H1N1) pandemic as well as others has raised concern of a far more dangerous outcome should this virus becomes resistant to current drug therapies. The number of clinical cases that are resistant to oseltamivir (Tamiflu®) is larger than the limited number of neuraminidase (NA) mutations (H275Y, N295S, and I223R) that have been identified at the active site and that are associated to oseltamivir resistance. In this study, we have performed a comparative analysis between a set of NAs that have the most representative mutations located outside the active site. The recently crystallized NA-oseltamivir complex (PDB ID: 3NSS) was used as a wild-type structure. After selecting the target NA sequences, their three-dimensional (3D) structure was built using 3NSS as a template by homology modeling. The 3D NA models were refined by molecular dynamics (MD) simulations. The refined models were used to perform a docking study, using oseltamivir as a ligand. Furthermore, the docking results were refined by free-energy analysis using the MM-PBSA method. The analysis of the MD simulation results showed that the NA models reached convergence during the first 10 ns. Visual inspection and structural measures showed that the mutated NA active sites show structural variations. The docking and MM-PBSA results from the complexes showed different binding modes and free energy values. These results suggest that distant mutations located outside the active site of NA affect its structure and could be considered to be a new source of resistance to oseltamivir, which agrees with reports in the clinical literature.

  9. Electromagnetic imaging of dynamic brain activity

    SciTech Connect

    Mosher, J.; Leahy, R. . Dept. of Electrical Engineering); Lewis, P.; Lewine, J.; George, J. ); Singh, M. . Dept. of Radiology)

    1991-01-01

    Neural activity in the brain produces weak dynamic electromagnetic fields that can be measured by an array of sensors. Using a spatio-temporal modeling framework, we have developed a new approach to localization of multiple neural sources. This approach is based on the MUSIC algorithm originally developed for estimating the direction of arrival of signals impinging on a sensor array. We present applications of this technique to magnetic field measurements of a phantom and of a human evoked somatosensory response. The results of the somatosensory localization are mapped onto the brain anatomy obtained from magnetic resonance images.

  10. Electromagnetic imaging of dynamic brain activity

    SciTech Connect

    Mosher, J.; Leahy, R.; Lewis, P.; Lewine, J.; George, J.; Singh, M.

    1991-12-31

    Neural activity in the brain produces weak dynamic electromagnetic fields that can be measured by an array of sensors. Using a spatio-temporal modeling framework, we have developed a new approach to localization of multiple neural sources. This approach is based on the MUSIC algorithm originally developed for estimating the direction of arrival of signals impinging on a sensor array. We present applications of this technique to magnetic field measurements of a phantom and of a human evoked somatosensory response. The results of the somatosensory localization are mapped onto the brain anatomy obtained from magnetic resonance images.

  11. Computation of Rate Constants for Diffusion of Small Ligands to and from Buried Protein Active Sites.

    PubMed

    Wang, P-H; De Sancho, D; Best, R B; Blumberger, J

    2016-01-01

    The diffusion of ligands to actives sites of proteins is essential to enzyme catalysis and many cellular signaling processes. In this contribution we review our recently developed methodology for calculation of rate constants for diffusion and binding of small molecules to buried protein active sites. The diffusive dynamics of the ligand obtained from molecular dynamics simulation is coarse grained and described by a Markov state model. Diffusion and binding rate constants are then obtained either from the reactive flux formalism or by fitting the time-dependent population of the Markov state model to a phenomenological rate law. The method is illustrated by applications to diffusion of substrate and inhibitors in [NiFe] hydrogenase, CO-dehydrogenase, and myoglobin. We also discuss a recently developed sensitivity analysis that allows one to identify hot spots in proteins, where mutations are expected to have the strongest effects on ligand diffusion rates.

  12. Computation of Rate Constants for Diffusion of Small Ligands to and from Buried Protein Active Sites.

    PubMed

    Wang, P-H; De Sancho, D; Best, R B; Blumberger, J

    2016-01-01

    The diffusion of ligands to actives sites of proteins is essential to enzyme catalysis and many cellular signaling processes. In this contribution we review our recently developed methodology for calculation of rate constants for diffusion and binding of small molecules to buried protein active sites. The diffusive dynamics of the ligand obtained from molecular dynamics simulation is coarse grained and described by a Markov state model. Diffusion and binding rate constants are then obtained either from the reactive flux formalism or by fitting the time-dependent population of the Markov state model to a phenomenological rate law. The method is illustrated by applications to diffusion of substrate and inhibitors in [NiFe] hydrogenase, CO-dehydrogenase, and myoglobin. We also discuss a recently developed sensitivity analysis that allows one to identify hot spots in proteins, where mutations are expected to have the strongest effects on ligand diffusion rates. PMID:27497172

  13. Resonant active sites in catalytic ammonia synthesis: A structural model

    NASA Astrophysics Data System (ADS)

    Cholach, Alexander R.; Bryliakova, Anna A.; Matveev, Andrey V.; Bulgakov, Nikolai N.

    2016-03-01

    Adsorption sites Mn consisted of n adjacent atoms M, each bound to the adsorbed species, are considered within a realistic model. The sum of bonds Σ lost by atoms in a site in comparison with the bulk atoms was used for evaluation of the local surface imperfection, while the reaction enthalpy at that site was used as a measure of activity. The comparative study of Mn sites (n = 1-5) at basal planes of Pt, Rh, Ir, Fe, Re and Ru with respect to heat of N2 dissociative adsorption QN and heat of Nad + Had → NHad reaction QNH was performed using semi-empirical calculations. Linear QN(Σ) increase and QNH(Σ) decrease allowed to specify the resonant Σ for each surface in catalytic ammonia synthesis at equilibrium Nad coverage. Optimal Σ are realizable for Ru2, Re2 and Ir4 only, whereas other centers meet steric inhibition or unreal crystal structure. Relative activity of the most active sites in proportion 5.0 × 10- 5: 4.5 × 10- 3: 1: 2.5: 3.0: 1080: 2270 for a sequence of Pt4, Rh4, Fe4(fcc), Ir4, Fe2-5(bcc), Ru2, Re2, respectively, is in agreement with relevant experimental data. Similar approach can be applied to other adsorption or catalytic processes exhibiting structure sensitivity.

  14. Multi-site Phosphorylation Regulates Bim Stability and Apoptotic Activity

    PubMed Central

    Hübner, Anette; Barrett, Tamera; Flavell, Richard A.; Davis, Roger J.

    2008-01-01

    The pro-apoptotic BH3-only protein Bim is established to be an important mediator of signaling pathways that induce cell death. Multi-site phosphorylation of Bim by several members of the MAP kinase group is implicated as a regulatory mechanism that controls the apoptotic activity of Bim. To test the role of Bim phosphorylation in vivo, we constructed mice with a series of mutant alleles that express phosphorylation-defective Bim proteins. We show that mutation of the phosphorylation site Thr-112 causes decreased binding of Bim to the anti-apoptotic protein Bcl2 and can increase cell survival. In contrast, mutation of the phosphorylation sites Ser-55, Ser-65, and Ser-73 can cause increased apoptosis because of reduced proteasomal degradation of Bim. Together, these data indicate that phosphorylation can regulate Bim by multiple mechanisms and that the phosphorylation of Bim on different sites can contribute to the sensitivity of cellular apoptotic responses. PMID:18498746

  15. Analysis of Hydrogen Tunneling in an Enzyme Active Site using von Neumann Measurements

    PubMed Central

    Sumner, Isaiah; Iyengar, Srinivasan S.

    2010-01-01

    We build on our earlier quantum wavepacket study of hydrogen transfer in the biological enzyme, soybean lipoxygenase-1, by using von Neumann quantum measurement theory to gain qualitative insights into the transfer event. We treat the enzyme active site as a measurement device which acts on the tunneling hydrogen nucleus via the potential it exerts at each configuration. A series of changing active site geometries during the tunneling process effects a sequential projection of the initial, reactant state onto the final, product state. We study this process using several different kinds of von Neumann measurements and show how a discrete sequence of such measurements not only progressively increases the projection of the hydrogen nuclear wavepacket onto the product side but also favors proton over deuteron transfer. Several qualitative features of the hydrogen tunneling problem found in wavepacket dynamics studies are also recovered here. These include the shift in the “transition state” towards the reactant as a result of nuclear quantization, greater participation of excited states in the case of deuterium, and presence of critical points along the reaction coordinate that facilitate hydrogen and deuterium transfer and coincide with surface crossings. To further “tailor” the dynamics, we construct a perturbation to the sequence of measurements, that is a perturbation to the dynamical sequence of active site geometry evolution, which leads us to insight on the existence of sensitive regions of the reaction profile where subtle changes to the dynamics of the active site can have an effect on the hydrogen and deuterium transfer process. PMID:22933858

  16. Dynamics of Active Sensing and perceptual selection.

    PubMed

    Schroeder, Charles E; Wilson, Donald A; Radman, Thomas; Scharfman, Helen; Lakatos, Peter

    2010-04-01

    Sensory processing is often regarded as a passive process in which biological receptors like photoreceptors and mechanoreceptors transduce physical energy into a neural code. Recent findings, however, suggest that: first, most sensory processing is active, and largely determined by motor/attentional sampling routines; second, owing to rhythmicity in the motor routine, as well as to its entrainment of ambient rhythms in sensory regions, sensory inflow tends to be rhythmic; third, attentional manipulation of rhythms in sensory pathways is instrumental to perceptual selection. These observations outline the essentials of an Active Sensing paradigm, and argue for increased emphasis on the study of sensory processes as specific to the dynamic motor/attentional context in which inputs are acquired. PMID:20307966

  17. Dynamics of Active Sensing and Perceptual Selection

    PubMed Central

    Schroeder, Charles E; Wilson, Donald A.; Radman, Thomas; Scharfman, Helen; Lakatos, Peter

    2010-01-01

    Sensory processing is often regarded as a passive process in which a biological sensors like photo- and mechanoreceptors transducer physical energy into a neural code. Recent findings, however, suggest that: 1) most sensory processing is active, and largely determined by motor/attentional sampling routines, 2) due to rhythmicity in the motor routine, as well as to its entrainment of ambient rhythms in sensory regions, sensory inflow tends to be rhythmic, and 3) attentional manipulation of rhythms in sensory pathways is instrumental to perceptual selection. These observations outline the essentials of an Active Sensing paradigm, and argue for increased emphasis on the study of sensory processes as specific to the dynamic motor/attentional context in which inputs are acquired. PMID:20307966

  18. Activation of phenylalanine hydroxylase by phenylalanine does not require binding in the active site.

    PubMed

    Roberts, Kenneth M; Khan, Crystal A; Hinck, Cynthia S; Fitzpatrick, Paul F

    2014-12-16

    Phenylalanine hydroxylase (PheH), a liver enzyme that catalyzes the hydroxylation of excess phenylalanine in the diet to tyrosine, is activated by phenylalanine. The lack of activity at low levels of phenylalanine has been attributed to the N-terminus of the protein's regulatory domain acting as an inhibitory peptide by blocking substrate access to the active site. The location of the site at which phenylalanine binds to activate the enzyme is unknown, and both the active site in the catalytic domain and a separate site in the N-terminal regulatory domain have been proposed. Binding of catecholamines to the active-site iron was used to probe the accessibility of the active site. Removal of the regulatory domain increases the rate constants for association of several catecholamines with the wild-type enzyme by ∼2-fold. Binding of phenylalanine in the active site is effectively abolished by mutating the active-site residue Arg270 to lysine. The k(cat)/K(phe) value is down 10⁴ for the mutant enzyme, and the K(m) value for phenylalanine for the mutant enzyme is >0.5 M. Incubation of the R270K enzyme with phenylalanine also results in a 2-fold increase in the rate constants for catecholamine binding. The change in the tryptophan fluorescence emission spectrum seen in the wild-type enzyme upon activation by phenylalanine is also seen with the R270K mutant enzyme in the presence of phenylalanine. Both results establish that activation of PheH by phenylalanine does not require binding of the amino acid in the active site. This is consistent with a separate allosteric site, likely in the regulatory domain.

  19. Chemical Modification of Papain and Subtilisin: An Active Site Comparison

    ERIC Educational Resources Information Center

    St-Vincent, Mireille; Dickman, Michael

    2004-01-01

    An experiment using methyle methanethiosulfonate (MMTS) and phenylmethylsulfonyl flouride (PMSF) to specifically modify the cysteine and serine residues in the active sites of papain and subtilism respectively is demonstrated. The covalent modification of these enzymes and subsequent rescue of papain shows the beginning biochemist that proteins…

  20. Dynamics of active cellular response under stress

    NASA Astrophysics Data System (ADS)

    de, Rumi; Zemel, Assaf; Safran, Samuel

    2008-03-01

    Forces exerted by and on adherent cells are important for many physiological processes such as wound healing and tissue formation. In addition, recent experiments have shown that stem cell differentiation is controlled, at least in part, by the elasticity of the surrounding matrix. Using a simple theoretical model that includes the forces due to both the mechanosensitive nature of cells and the elastic response of the matrix, we predict the dynamics of orientation of cells. The model predicts many features observed in measurements of cellular forces and orientation including the increase with time of the forces generated by cells in the absence of applied stress and the consequent decrease of the force in the presence of quasi-static stresses. We also explain the puzzling observation of parallel alignment of cells for static and quasi-static stresses and of nearly perpendicular alignment for dynamically varying stresses. In addition, we predict the response of the cellular orientation to a sinusoidally varying applied stress as a function of frequency. The dependence of the cell orientation angle on the Poisson ratio of the surrounding material can be used to distinguish systems in which cell activity is controlled by stress from those where cell activity is controlled by strain. Reference: Nature Physics, vol. 3, pp 655 (2007).

  1. Pacemaker activity in a sensory ending with multiple encoding sites: the cat muscle spindle primary ending.

    PubMed Central

    Banks, R W; Hulliger, M; Scheepstra, K A; Otten, E

    1997-01-01

    1. A combined physiological, histological and computer modelling study was carried out on muscle spindles of the cat tenuissimus muscle to examine whether there was any correlation between the functional interaction of putative encoding sites, operated separately by static and dynamic fusimotor neurones, and the topological structure of the preterminal branches of the primary sensory ending. 2. Spindles, whose I a responses to stretch and separate and combined static and dynamic fusimotor stimulation were recorded in physiological experiments, were located in situ. Subsequently the ramifications of the sensory ending were reconstructed histologically, and the topology of the branch tree was used in computer simulations of I a responses to examine the effect of the electronic separation of encoding sites on the static-dynamic interaction pattern. 3. Interactions between separate static and dynamic inputs, manifest in responses to combineed stimulation, were quantified by a coefficient of interaction (Ci) which, by definition, was 1 for strictly linear summation of separate inputs and zero for maximum occlusion between inputs. 4. For the majority of spindles static-dynamic interactions were characterized by pronounced occlusion (C1 < 0.35). In these spindles putative encoding sites (the peripheral heminodes of the branches supplying the intrafusal fibres activated by individual fusimotor efferents) were separated by a minimum conduction path of between three and ten myelinated segments (2-9 nodes of Ranvier). In contrast, significant summation (C1, approximately 0.7) was found in only one spindle. In this case putative encoding sites were separated by a single node. 5. Occlusion was not due to encoder saturation and it could not be accounted for by any other known physiological mechanisms (intrafusal fatigue or unloading). It is therefore attributed to competitive pacemaker interaction between encoding sites which are largely selectively operated by static and

  2. N-methyl-D-aspartate recognition site ligands modulate activity at the coupled glycine recognition site.

    PubMed

    Hood, W F; Compton, R P; Monahan, J B

    1990-03-01

    In synaptic plasma membranes from rat forebrain, the potencies of glycine recognition site agonists and antagonists for modulating [3H]1-[1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP) binding and for displacing strychnine-insensitive [3H]glycine binding are altered in the presence of N-methyl-D-aspartate (NMDA) recognition site ligands. The NMDA competitive antagonist, cis-4-phosphonomethyl-2-piperidine carboxylate (CGS 19755), reduces [3H]glycine binding, and the reduction can be fully reversed by the NMDA recognition site agonist, L-glutamate. Scatchard analysis of [3H]glycine binding shows that in the presence of CGS 19755 there is no change in Bmax (8.81 vs. 8.79 pmol/mg of protein), but rather a decrease in the affinity of glycine (KD of 0.202 microM vs. 0.129 microM). Similar decreases in affinity are observed for the glycine site agonists, D-serine and 1-aminocyclopropane-1-carboxylate, in the presence of CGS 19755. In contrast, the affinity of glycine antagonists, 1-hydroxy-3-amino-2-pyrrolidone and 1-aminocyclobutane-1-carboxylate, at this [3H]glycine recognition site increases in the presence of CGS 19755. The functional consequence of this change in affinity was addressed using the modulation of [3H]TCP binding. In the presence of L-glutamate, the potency of glycine agonists for the stimulation of [3H]TCP binding increases, whereas the potency of glycine antagonists decreases. These data are consistent with NMDA recognition site ligands, through their interactions at the NMDA recognition site, modulating activity at the associated glycine recognition site.

  3. Parameterization of an Active Thermal Erosion Site, Caribou Creek, Alaska

    NASA Astrophysics Data System (ADS)

    Busey, R.; Bolton, W. R.; Cherry, J. E.; Hinzman, L. D.

    2012-12-01

    Thermokarst features are thought to be an important mechanism for landscape change in permafrost-dominated cold regions, but few such features have been incorporated into full featured landscape models. The root of this shortcoming is that historic observations are not detailed enough to parameterize a model, and the models typically do not include the relevant processes for thermal erosion. A new, dynamic thermokarst feature has been identified at the Caribou-Poker Creek Research Watershed (CPCRW) in the boreal forest of Interior Alaska. Located adjacent to a traditional use trail, this feature terminates directly in Caribou Creek. Erosion within the feature is driven predominantly by fluvial interflow. CPCRW is a Long-Term Ecological Research site underlain by varying degrees of relatively warm, discontinuous permafrost. This poster will describe the suite of measurements that have been undertaken to parameterize the ERODE model for this site, including thorough surveys, time lapse- and aerial photography, and 3-D structure from motion algorithms.

  4. Shifts of Gamma Phase across Primary Visual Cortical Sites Reflect Dynamic Stimulus-Modulated Information Transfer

    PubMed Central

    Besserve, Michel; Lowe, Scott C.; Logothetis, Nikos K.; Schölkopf, Bernhard; Panzeri, Stefano

    2015-01-01

    Distributed neural processing likely entails the capability of networks to reconfigure dynamically the directionality and strength of their functional connections. Yet, the neural mechanisms that may allow such dynamic routing of the information flow are not yet fully understood. We investigated the role of gamma band (50–80 Hz) oscillations in transient modulations of communication among neural populations by using measures of direction-specific causal information transfer. We found that the local phase of gamma-band rhythmic activity exerted a stimulus-modulated and spatially-asymmetric directed effect on the firing rate of spatially separated populations within the primary visual cortex. The relationships between gamma phases at different sites (phase shifts) could be described as a stimulus-modulated gamma-band wave propagating along the spatial directions with the largest information transfer. We observed transient stimulus-related changes in the spatial configuration of phases (compatible with changes in direction of gamma wave propagation) accompanied by a relative increase of the amount of information flowing along the instantaneous direction of the gamma wave. These effects were specific to the gamma-band and suggest that the time-varying relationships between gamma phases at different locations mark, and possibly causally mediate, the dynamic reconfiguration of functional connections. PMID:26394205

  5. Discovery of an Aurora kinase inhibitor through site-specific dynamic combinatorial chemistry.

    PubMed

    Cancilla, Mark T; He, Molly M; Viswanathan, Nina; Simmons, Robert L; Taylor, Meggin; Fung, Amy D; Cao, Kathy; Erlanson, Daniel A

    2008-07-15

    We demonstrate a fragment-based lead discovery method that combines site-directed ligand discovery with dynamic combinatorial chemistry. Our technique targets dynamic combinatorial screening to a specified region of a protein by using reversible disulfide chemistry. We have used this technology to rapidly identify inhibitors of the drug target Aurora A that span the purine-binding site and the adaptive pocket of the kinase. The binding mode of a noncovalent inhibitor has been further characterized through crystallography.

  6. Discovery of An Aurora Kinase Inhibitor Through Site-Specific Dynamic Combinatorial Chemistry

    SciTech Connect

    Cancilla, M.T.; He, M.M.; Viswanathan, N.; Simmons, R.L.; Taylor, M.; Fung, A.D.; Cao, K.; Erlanson, D.A.

    2009-05-12

    We demonstrate a fragment-based lead discovery method that combines site-directed ligand discovery with dynamic combinatorial chemistry. Our technique targets dynamic combinatorial screening to a specified region of a protein by using reversible disulfide chemistry. We have used this technology to rapidly identify inhibitors of the drug target Aurora A that span the purine-binding site and the adaptive pocket of the kinase. The binding mode of a noncovalent inhibitor has been further characterized through crystallography.

  7. Control of active sites in selective flocculation: I -- Mathematical model

    SciTech Connect

    Behl, S.; Moudgil, B.M.; Prakash, T.S. . Dept. of Materials Science and Engineering)

    1993-12-01

    Heteroflocculation has been determined to be another major reason for loss in selectivity for flocculation process. In a mathematical model developed earlier, conditions for controlling heteroflocculation were discussed. Blocking active sites to control selective adsorption of a flocculant oil a desirable solid surface is discussed. It has been demonstrated that the lower molecular weight fraction of a flocculant which is incapable of flocculating the particles is an efficient site blocking agent. The major application of selective flocculation has been in mineral processing but many potential uses exist in biological and other colloidal systems. These include purification of ceramic powders, separating hazardous solids from chemical waste, and removal of deleterious components from paper pulp.

  8. The site of activation of factor X by cancer procoagulant.

    PubMed

    Gordon, S G; Mourad, A M

    1991-12-01

    Cancer procoagulant (CP) is a cysteine proteinase found in a variety of malignant cells and tissues and in human amnion-chorion tissue. It initiates coagulation by activating factor X. However, the amino acid sequence of the substrate protein that determines the cleavage site of cysteine proteinases is different from that of the serine proteinases that normally activate factor X, such as factor IXa, VIIa and Russell's Viper Venom (RVV). Therefore, it was of interest to determine the site of cleavage of human factor X by CP. Purified CP was incubated with purified factor X and the reaction mixture was electrophoresed on a 10% Tris-tricine SDS-PAGE gel. The proteins were electroeluted on to a polyvinylidene difluoride (PVDF) membrane, and stained with Coomassie blue. The heavy chain of activated factor X was cut out of the PVDF membrane and sequenced with an Applied Biosystems 477A with on-line HPLC. The primary cleavage sequence was Asp-Ala-Ala-Asp-Leu-Asp-Pro-; two other secondary sequences Ser-Ile-Thr-Trp-Lys-Pro- and Glu-Asn-Pro-Phe-Asp-Leu were found. The penultimate amino acid on the carbonyl side of the hydrolysed amide bond plays a critical role for the recognition of the cleavage site of cysteine proteinases. These data indicate that the penultimate amino acid for the primary cleavage site of factor X by CP is proline-20 and for the secondary sites, proline-13 and proline-28. This is in contrast to arginine-52 that determines the specificity of the cleavage by normal serine proteinase activation.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Reduction of Urease Activity by Interaction with the Flap Covering the Active Site

    PubMed Central

    Macomber, Lee; Minkara, Mona S.; Hausinger, Robert P.; Merz, Kenneth M.

    2015-01-01

    With the increasing appreciation for the human microbiome coupled with the global rise of antibiotic resistant organisms, it is imperative that new methods be developed to specifically target pathogens. To that end, a novel computational approach was devised to identify compounds that reduce the activity of urease, a medically important enzyme of Helicobacter pylori, Proteus mirabilis, and many other microorganisms. Urease contains a flexible loop that covers its active site; Glide was used to identify small molecules predicted to lock this loop in an open conformation. These compounds were screened against the model urease from Klebsiella aerogenes and the natural products epigallocatechin and quercetin were shown to inhibit at low and high micromolar concentrations, respectively. These molecules exhibit a strong time-dependent inactivation of urease that was not due to their oxygen sensitivity. Rather, these compounds appear to inactivate urease by reacting with a specific Cys residue located on the flexible loop. Substitution of this cysteine by alanine in the C319A variant increased the urease resistance to both epigallocatechin and quercetin, as predicted by the computational studies. Protein dynamics are integral to the function of many enzymes; thus, identification of compounds that lock an enzyme into a single conformation presents a useful approach to define potential inhibitors. PMID:25594724

  10. New activity pattern in human interactive dynamics

    NASA Astrophysics Data System (ADS)

    Formentin, Marco; Lovison, Alberto; Maritan, Amos; Zanzotto, Giovanni

    2015-09-01

    We investigate the response function of human agents as demonstrated by written correspondence, uncovering a new pattern for how the reactive dynamics of individuals is distributed across the set of each agent’s contacts. In long-term empirical data on email, we find that the set of response times considered separately for the messages to each different correspondent of a given writer, generate a family of heavy-tailed distributions, which have largely the same features for all agents, and whose characteristic times grow exponentially with the rank of each correspondent. We furthermore show that this new behavioral pattern emerges robustly by considering weighted moving averages of the priority-conditioned response-time probabilities generated by a basic prioritization model. Our findings clarify how the range of priorities in the inputs from one’s environment underpin and shape the dynamics of agents embedded in a net of reactive relations. These newly revealed activity patterns might be universal, being present in other general interactive environments, and constrain future models of communication and interaction networks, affecting their architecture and evolution.

  11. Dynamical Network of HIV-1 Protease Mutants Reveals the Mechanism of Drug Resistance and Unhindered Activity.

    PubMed

    Appadurai, Rajeswari; Senapati, Sanjib

    2016-03-15

    HIV-1 protease variants resist drugs by active and non-active-site mutations. The active-site mutations, which are the primary or first set of mutations, hamper the stability of the enzyme and resist the drugs minimally. As a result, secondary mutations that not only increase protein stability for unhindered catalytic activity but also resist drugs very effectively arise. While the mechanism of drug resistance of the active-site mutations is through modulating the active-site pocket volume, the mechanism of drug resistance of the non-active-site mutations is unclear. Moreover, how these allosteric mutations, which are 8-21 Å distant, communicate to the active site for drug efflux is completely unexplored. Results from molecular dynamics simulations suggest that the primary mechanism of drug resistance of the secondary mutations involves opening of the flexible protease flaps. Results from both residue- and community-based network analyses reveal that this precise action of protease is accomplished by the presence of robust communication paths between the mutational sites and the functionally relevant regions: active site and flaps. While the communication is more direct in the wild type, it traverses across multiple intermediate residues in mutants, leading to weak signaling and unregulated motions of flaps. The global integrity of the protease network is, however, maintained through the neighboring residues, which exhibit high degrees of conservation, consistent with clinical data and mutagenesis studies. PMID:26892689

  12. Active-Site-Accessible, Porphyrinic Metal;#8722;Organic Framework Materials

    SciTech Connect

    Farha, Omar K.; Shultz, Abraham M.; Sarjeant, Amy A.; Nguyen, SonBinh T.; Hupp, Joseph T.

    2012-02-06

    On account of their structural similarity to cofactors found in many metallo-enzymes, metalloporphyrins are obvious potential building blocks for catalytically active, metal-organic framework (MOF) materials. While numerous porphyrin-based MOFs have already been described, versions featuring highly accessible active sites and permanent microporosity are remarkably scarce. Indeed, of the more than 70 previously reported porphyrinic MOFs, only one has been shown to be both permanently microporous and contain internally accessible active sites for chemical catalysis. Attempts to generalize the design approach used in this single successful case have failed. Reported here, however, is the synthesis of an extended family of MOFs that directly incorporate a variety of metalloporphyrins (specifically Al{sup 3+}, Zn{sup 2+}, Pd{sup 2+}, Mn{sup 3+}, and Fe{sup 3+} complexes). These robust porphyrinic materials (RPMs) feature large channels and readily accessible active sites. As an illustrative example, one of the manganese-containing RPMs is shown to be catalytically competent for the oxidation of alkenes and alkanes.

  13. Functional constituents of the active site of human neutrophil collagenase.

    PubMed

    Mookhtiar, K A; Wang, F; Van Wart, H E

    1986-05-01

    A series of chemical modification reactions has been carried out to identify functional constituents of the active site of human neutrophil collagenase. The enzyme is reversibly inhibited by the transition metal chelating agent 1,10-phenanthroline, and inhibition is fully reversed by zinc. Removal of weakly bound metal ions by gel filtration inactivates collagenase, and activity is fully restored on immediate readdition of calcium. The enzyme is unaffected by reagents that modify serine, cysteine, and arginine residues. However, reaction with the carboxyl reagents cyclohexylmorpholinocarbodiimide and Woodward's Reagent K lowers the activity of the enzyme substantially. Acetylimidazole inactivates the enzyme, but activity is completely restored on addition of hydroxylamine. The enzyme is also inactivated by tetranitromethane, indicating that it contains an essential tyrosine residue. Acylation of collagenase with diethyl pyrocarbonate, diketene, acetic anhydride, or trinitrobenzenesulfonate inactivates the enzyme, and activity is not restored on addition of hydroxylamine, indicating the presence of an essential lysine residue.

  14. Molecular dynamics simulation and site-directed mutagenesis of alcohol acyltransferase: a proposed mechanism of catalysis.

    PubMed

    Morales-Quintana, Luis; Nuñez-Tobar, María Ximena; Moya-León, María Alejandra; Herrera, Raúl

    2013-10-28

    Aroma in Vasconcellea pubescens fruit is determined by esters, which are the products of catalysis by alcohol acyltransferase (VpAAT1). VpAAT1 protein structure displayed the conserved HxxxD motif facing the solvent channel in the center of the structure. To gain insight into the role of these catalytic residues, kinetic and site-directed mutagenesis studies were carried out in VpAAT1 protein. Based on dead-end inhibition studies, the kinetic could be described in terms of a ternary complex mechanism with the H166 residue as the catalytic base. Kinetic results showed the lowest Km value for hexanoyl-CoA. Additionally, the most favorable predicted substrate orientation was observed for hexanoyl-CoA, showing a coincidence between kinetic studies and molecular docking analysis. Substitutions H166A, D170A, D170N, and D170E were evaluated in silico. The solvent channel in all mutant structures was lost, showing large differences with the native structure. Molecular docking and molecular dynamics simulations were able to describe unfavored energies for the interaction of the mutant proteins with different alcohols and acyl-CoAs. Additionally, in vitro site-directed mutagenesis of H166 and D170 in VpAAT1 induced a loss of activity, confirming the functional role of both residues for the activity, H166 being directly involved in catalysis.

  15. Erythrosin B Phosphorescence Monitors Molecular Mobility and Dynamic Site Heterogeneity in Amorphous Sucrose

    PubMed Central

    Pravinata, Linda C.; You, Yumin; Ludescher, Richard D.

    2005-01-01

    Molecular mobility modulates the chemical and physical stability of amorphous biomaterials. This study used steady-state and time-resolved phosphorescence of erythrosin B to monitor mobility in thin films of amorphous solid sucrose as a function of temperature. The phosphorescence intensity (lifetime), emission energy, and red-edge excitation effect were all sensitive to localized molecular mobility on the microsecond timescale in the glass and to more global modes of mobility activated at the glass transition. Blue shifts in the emission spectrum with time after excitation and systematic variations in the phosphorescence lifetime with wavelength indicated that emission originates from multiple sites ranging from short lifetime species with red-shifted emission spectrum to long lifetime species with blue-shifted emission spectrum; the activation energy for nonradiative decay of the triplet state was considerably larger for the blue-emitting species in both the glass and the melt. This study illustrates that phosphorescence from erythrosin B is sensitive both to local dipolar relaxations in the glass as well as more global relaxations in the sucrose melt and provides evidence of the value of phosphorescence as a probe of dynamic site heterogeneity as well as overall molecular mobility in amorphous biomaterials. PMID:15695637

  16. The Dynamic Dielectric at a Brain Functional Site and an EM Wave Approach to Functional Brain Imaging

    PubMed Central

    Li, X. P.; Xia, Q.; Qu, D.; Wu, T. C.; Yang, D. G.; Hao, W. D.; Jiang, X.; Li, X. M.

    2014-01-01

    Functional brain imaging has tremendous applications. The existing methods for functional brain imaging include functional Magnetic Resonant Imaging (fMRI), scalp electroencephalography (EEG), implanted EEG, magnetoencephalography (MEG) and Positron Emission Tomography (PET), which have been widely and successfully applied to various brain imaging studies. To develop a new method for functional brain imaging, here we show that the dielectric at a brain functional site has a dynamic nature, varying with local neuronal activation as the permittivity of the dielectric varies with the ion concentration of the extracellular fluid surrounding neurons in activation. Therefore, the neuronal activation can be sensed by a radiofrequency (RF) electromagnetic (EM) wave propagating through the site as the phase change of the EM wave varies with the permittivity. Such a dynamic nature of the dielectric at a brain functional site provides the basis for an RF EM wave approach to detecting and imaging neuronal activation at brain functional sites, leading to an RF EM wave approach to functional brain imaging. PMID:25367217

  17. The dynamic dielectric at a brain functional site and an EM wave approach to functional brain imaging.

    PubMed

    Li, X P; Xia, Q; Qu, D; Wu, T C; Yang, D G; Hao, W D; Jiang, X; Li, X M

    2014-11-04

    Functional brain imaging has tremendous applications. The existing methods for functional brain imaging include functional Magnetic Resonant Imaging (fMRI), scalp electroencephalography (EEG), implanted EEG, magnetoencephalography (MEG) and Positron Emission Tomography (PET), which have been widely and successfully applied to various brain imaging studies. To develop a new method for functional brain imaging, here we show that the dielectric at a brain functional site has a dynamic nature, varying with local neuronal activation as the permittivity of the dielectric varies with the ion concentration of the extracellular fluid surrounding neurons in activation. Therefore, the neuronal activation can be sensed by a radiofrequency (RF) electromagnetic (EM) wave propagating through the site as the phase change of the EM wave varies with the permittivity. Such a dynamic nature of the dielectric at a brain functional site provides the basis for an RF EM wave approach to detecting and imaging neuronal activation at brain functional sites, leading to an RF EM wave approach to functional brain imaging.

  18. Nest predation increases with parental activity: Separating nest site and parental activity effects

    USGS Publications Warehouse

    Martin, T.E.; Scott, J.; Menge, C.

    2000-01-01

    Alexander Skutch hypothesized that increased parental activity can increase the risk of nest predation. We tested this hypothesis using ten open-nesting bird species in Arizona, USA. Parental activity was greater during the nestling than incubation stage because parents visited the nest frequently to feed their young during the nestling stage. However, nest predation did not generally increase with parental activity between nesting stages across the ten study species. Previous investigators have found similar results. We tested whether nest site effects might yield higher predation during incubation because the most obvious sites are depredated most rapidly. We conducted experiments using nest sites from the previous year to remove parental activity. Our results showed that nest sites have highly repeatable effects on nest predation risk; poor nest sites incurred rapid predation and caused predation rates to be greater during the incubation than nestling stage. This pattern also was exhibited in a bird species with similar (i.e. controlled) parental activity between nesting stages. Once nest site effects are taken into account, nest predation shows a strong proximate increase with parental activity during the nestling stage within and across species. Parental activity and nest sites exert antagonistic influences on current estimates of nest predation between nesting stages and both must be considered in order to understand current patterns of nest predation, which is an important source of natural selection.

  19. Proposed satellite laser ranging and very long baseline interferometry sites for crustal dynamics investigations

    NASA Technical Reports Server (NTRS)

    Lowman, P. D.; Allenby, R. J.; Frey, H. V.

    1979-01-01

    Recommendations are presented for a global network of 125 sites for geodetic measurements by satellite laser ranging and very long baseline interferometry. The sites were proposed on the basis of existing facilities and scientific value for investigation of crustal dynamics as related to earthquake hazards. Tectonic problems are discussed for North America peripheral regions and for the world. The sites are presented in tables and maps, with bibliographic references.

  20. Dynamical Observations of Local Bio-molecular Sites Using Nanocrystals

    SciTech Connect

    Sasaki, Yuji C.; Okumura, Yasuaki; Ohishi, Noboru

    2004-05-12

    Recently, we succeeded time-resolved x-ray observations of picometer-scale slow Brownian motions of individual protein molecules in aqueous solutions. In this work, we investigated the relationship between individual protein molecules and the labeled nanocrystals. In order to control the number of the bonding sites on the labeled gold nanocrystals, we utilized both the periodical structure in Actin filaments and using the mercury compound. As a result, we observed free Brownian Motions of the C-terminal in F-actin, when single gold nanocrystal is linked to single G-actin.

  1. Threshold occupancy and specific cation binding modes in the hammerhead ribozyme active site are required for active conformation

    PubMed Central

    Lee, Tai-Sung; Giambaşu, George M.; Sosa, Carlos P.; Martick, Monika; Scott, William G.; York, Darrin M.

    2009-01-01

    The relationship between formation of active in-line attack conformations and monovalent (Na+) and divalent (Mg2+) metal ion binding in the hammerhead ribozyme has been explored with molecular dynamics simulations. To stabilize repulsions between negatively charged groups, different requirements of threshold occupancy of metal ions were observed in the reactant and activated precursor states both in the presence or absence of a Mg2+ in the active site. Specific bridging coordination patterns of the ions are correlated with the formation of active in-line attack conformations and can be accommodated in both cases. Furthermore, simulation results suggest that the hammerhead ribozyme folds to form an electronegative recruiting pocket that attracts high local concentrations of positive charge. The present simulations help to reconcile experiments that probe the metal ion sensitivity of hammerhead ribozyme catalysis and support the supposition that Mg2+, in addition to stabilizing active conformations, plays a specific chemical role in catalysis. PMID:19265710

  2. Active sites in char gasification: Final technical report

    SciTech Connect

    Wojtowicz, M.; Lilly, W.D.; Perkins, M.T.; Hradil, G.; Calo, J.M.; Suuberg, E.M.

    1987-09-01

    Among the key variables in the design of gasifiers and combustors is the reactivity of the chars which must be gasified or combusted. Significant loss of unburned char is unacceptable in virtually any process; the provision of sufficient residence time for complete conversion is essential. A very wide range of reactivities are observed, depending upon the nature of the char in a process. The current work focuses on furthering the understanding of gasification reactivities of chars. It has been well established that the reactivity of char to gasification generally depends upon three principal factors: (1) the concentration of ''active sites'' in the char; (2) mass transfer within the char; and (3) the type and concentration of catalytic impurities in the char. The present study primarily addresses the first factor. The subject of this research is the origin, nature, and fate of active sites in chars derived from parent hydrocarbons with coal-like structure. The nature and number of the active sites and their reactivity towards oxygen are examined in ''model'' chars derived from phenol-formaldehyde type resins. How the active sites are lost by the process of thermal annealing during heat treatment of chars are studied, and actual rate for the annealing process is derived. Since intrinsic char reactivities are of primary interest in the present study, a fair amount of attention was given to the model char synthesis and handling so that the effect of catalytic impurities and oxygen-containing functional groups in the chemical structure of the material were minimized, if not completely eliminated. The project would not be considered complete without comparing characteristic features of synthetic chars with kinetic behavior exhibited by natural chars, including coal chars.

  3. Brownian aggregation rate of colloid particles with several active sites

    SciTech Connect

    Nekrasov, Vyacheslav M.; Yurkin, Maxim A.; Chernyshev, Andrei V.; Polshchitsin, Alexey A.; Yakovleva, Galina E.; Maltsev, Valeri P.

    2014-08-14

    We theoretically analyze the aggregation kinetics of colloid particles with several active sites. Such particles (so-called “patchy particles”) are well known as chemically anisotropic reactants, but the corresponding rate constant of their aggregation has not yet been established in a convenient analytical form. Using kinematic approximation for the diffusion problem, we derived an analytical formula for the diffusion-controlled reaction rate constant between two colloid particles (or clusters) with several small active sites under the following assumptions: the relative translational motion is Brownian diffusion, and the isotropic stochastic reorientation of each particle is Markovian and arbitrarily correlated. This formula was shown to produce accurate results in comparison with more sophisticated approaches. Also, to account for the case of a low number of active sites per particle we used Monte Carlo stochastic algorithm based on Gillespie method. Simulations showed that such discrete model is required when this number is less than 10. Finally, we applied the developed approach to the simulation of immunoagglutination, assuming that the formed clusters have fractal structure.

  4. Active Sites Environmental Monitoring Program: FY 1991 report

    SciTech Connect

    Ashwood, T.L.; Hicks, D.S.; Morrissey, C.M.

    1992-11-01

    This report summarizes the activities of the Active Sites Environmental Monitoring Program (ASEMP) from April 1991 through September 1991. The ASEMP was established in 1989 by Solid Waste Operations (SWO) and the Environmental Sciences Division, both of Oak Ridge National Laboratory, to provide early detection and performance monitoring at active low-level (radioactive) waste (LLW) disposal sites in Solid Waste Storage Area (SWSA) 6 and transuranic (TRU) waste storage sites in SWSA 5 as required by chapters II and III of US Department of Energy Order 5820.2A. A new set of action levels was developed on the basis of a statistical analysis of background contamination. These new action levels have been used to evaluate results in this report. Results of ASEMP monitoring continue to demonstrate that no LLW (except [sup 3]H) is being leached from the storage vaults on the tumulus pads. Loading of vaults on Tumulus II, which began in early FY 1991, was >90% complete at the end of September 1991. Results of sampling of groundwater and surface waters is presented.

  5. Inhibition and active-site modelling of prolidase.

    PubMed

    King, G F; Crossley, M J; Kuchel, P W

    1989-03-15

    Consideration of the active-site model of prolidase led us to examine azetidine, pyrrolidine and piperidine substrate analogs as potential in vivo inhibitors of the enzyme. One of these, N-benzyloxycarbonyl-L-proline, was shown to be a potent competitive inhibitor of porcine kidney prolidase (Ki = 90 microM); its rapid protein-mediated permeation of human and sheep erythrocytes suggests that it may be effective in vivo. The higher homolog, N-benzyloxycarbonyl-L-pipecolic acid, was also a potent inhibitor of the enzyme while the antihypertensive drugs, captopril and enalaprilat, were shown to have mild and no inhibitory effects, respectively. Analysis of inhibitor action and consideration of X-ray crystallographic data of relevant Mn2+ complexes allowed the active-site model of prolidase to be further refined; a new model is presented in which the substrate acts as a bidentate ligand towards the active-site manganous ion. Various aspects of the new model help to explain why Mn2+ has been 'chosen' by the enzyme in preference to other biologically available metal ions. PMID:2924773

  6. Burial trench dynamic compaction demonstration at a humid site

    SciTech Connect

    Spalding, B.P.

    1985-01-01

    This task has the objective of determining the degree of consolidation which can be achieved by dynamic compaction of a closed burial trench within a cohesive soil formation. A seven-year-old burial trench in Solid Waste Storage Area (SWSA) 6 of Oak Ridge National Laboratory (ORNL) was selected for this demonstration. This 251 m/sup 3/ trench contained about 80 Ci of mixed radionuclides, mostly /sup 90/Sr, in 25 m/sup 3/ of waste consisting of contaminated equipment, dry solids, and demolition debris. Prior to compaction, a total trench void space of 79 m/sup 3/ was measured by pumping the trench full of water with corrections for seepage. Additional pre-compaction characterization included trench cap bulk density (1.68 kg/L), trench cap permeability (3 x 10/sup -7/ m/s), and subsurface waste/backfill hydraulic conductivity (>0.01 m/s). Compaction was achieved by repeatedly dropping a 4-ton steel-reinforced concrete cylinder from heights of 4 to 8 m using the whipline of a 70-ton crane. The average trench ground surface was depressed 0.79 m, with some sections over 2 m, yielding a surveyed volumetric depression which totaled to 64% of the measured trench void space. Trench cap (0 to 60 cm) bulk density and permeability were not affected by compaction indicating that the consolidation was largely subsurface. Neither surface nor airborne radioactive contamination were observed during repeated monitoring during the demonstration. Dynamic compaction was shown to be an excellent and inexpensive (i.e., about $20/m/sup 2/) method to collapse trench void space, thereby hastening subsidence and stabilizing the land surface. 15 refs., 10 figs., 3 tabs.

  7. Dynamics of the Thermal State of Active Layer at the Alaska North Slope and Northern Yakutia

    NASA Astrophysics Data System (ADS)

    Kholodov, A. L.; Romanovsky, V. E.; Marchenko, S.; Shiklomanov, N. I.; Fedorov-Davydov, D.

    2010-12-01

    Dynamics of the active layer is one of the most important indexes, reflecting permafrost response to the modern climate changes. Monitoring of active layer thickness dynamics is the main goal of CALM (Circumpolar Active Layer Monitoring) project. But, from different points of view, it is very important to know not only maximal depth of seasonal thawing but also dynamics of thermal field of active layer and duration of its staying in the unfrozen state. Current research was aimed on the analyzing data of temperature measurements have been done during the more then 10 years at the North Slope of Brooks Range (Alaska) and 2 years at the selected sites at the Northern Yakutia (Russia) and its comparison with the 17 to 10 years records of active layer thickness dynamics at the corresponding sites (http://www.udel.edu/Geography/calm/data/north.html). The area of investigation characterized by the typical tundra landscape and different kinds of micro topography. Reported observation sites located at the latitudinal range from 68.5 to 70.3N in Alaska and 70.5 to 71.75N in the Northern Yakutia. Observation have been done using the 1 meter long MRC probe with 11 sensors (every 10 cm) and single Campbell SCI A107 sensors in Alaska and 2-channel HOBO U23 data loggers with TMC-HD thermistors in the Northern Yakutia. Analyses of CALM data show what most observation sites in Alaska (except located near the Brooks Range and at the Arctic Ocean coast) do not subjected to the significant sustainable changes of active layer thickness over the last 10 years. At the same time active layer thickness at the Yakutian sites was increasing. Temperature observations show decreasing of the mean annual temperature at the average depth of active layer bottom at the Alaskan sites. But, because of general trend to increasing of period of thawing it does not lead to the decreasing of active layer thickness. Recent equipment deployment at the Tiksi and Allaikha sites (Northern Yakutia) does not

  8. The ArcSDE GIS Dynamic Population Model Tool for Savannah River Site Emergency Response

    SciTech Connect

    MCLANE, TRACY; JONES, DWIGHT

    2005-10-03

    The Savannah River Site (SRS) is a 310-square-mile Department of Energy site located near Aiken, South Carolina. With a workforce of over 10,000 employees and subcontractors, SRS emergency personnel must be able to respond to an emergency event in a timely and effective manner, in order to ensure the safety and security of the Site. Geographic Information Systems (GIS) provides the technology needed to give managers and emergency personnel the information they need to make quick and effective decisions. In the event of a site evacuation, knowing the number of on-site personnel to evacuate from a given area is an essential piece of information for emergency staff. SRS has developed a GIS Dynamic Population Model Tool to quickly communicate real-time information that summarizes employee populations by facility area and building and then generates dynamic maps that illustrate output statistics.

  9. Druggability analysis and classification of protein tyrosine phosphatase active sites

    PubMed Central

    Ghattas, Mohammad A; Raslan, Noor; Sadeq, Asil; Al Sorkhy, Mohammad; Atatreh, Noor

    2016-01-01

    Protein tyrosine phosphatases (PTP) play important roles in the pathogenesis of many diseases. The fact that no PTP inhibitors have reached the market so far has raised many questions about their druggability. In this study, the active sites of 17 PTPs were characterized and assessed for its ability to bind drug-like molecules. Consequently, PTPs were classified according to their druggability scores into four main categories. Only four members showed intermediate to very druggable pocket; interestingly, the rest of them exhibited poor druggability. Particularly focusing on PTP1B, we also demonstrated the influence of several factors on the druggability of PTP active site. For instance, the open conformation showed better druggability than the closed conformation, while the tight-bound water molecules appeared to have minimal effect on the PTP1B druggability. Finally, the allosteric site of PTP1B was found to exhibit superior druggability compared to the catalytic pocket. This analysis can prove useful in the discovery of new PTP inhibitors by assisting researchers in predicting hit rates from high throughput or virtual screening and saving unnecessary cost, time, and efforts via prioritizing PTP targets according to their predicted druggability. PMID:27757011

  10. Spatio-temporal dynamics of replication and transcription sites in the mammalian cell nucleus.

    PubMed

    Malyavantham, Kishore S; Bhattacharya, Sambit; Alonso, William D; Acharya, Raj; Berezney, Ronald

    2008-12-01

    To study when and where active genes replicated in early S phase are transcribed, a series of pulse-chase experiments are performed to label replicating chromatin domains (RS) in early S phase and subsequently transcription sites (TS) after chase periods of 0 to 24 h. Surprisingly, transcription activity throughout these chase periods did not show significant colocalization with early RS chromatin domains. Application of novel image segmentation and proximity algorithms, however, revealed close proximity of TS with the labeled chromatin domains independent of chase time. In addition, RNA polymerase II was highly proximal and showed significant colocalization with both TS and the chromatin domains. Based on these findings, we propose that chromatin activated for transcription dynamically unfolds or "loops out" of early RS chromatin domains where it can interact with RNA polymerase II and other components of the transcriptional machinery. Our results further suggest that the early RS chromatin domains are transcribing genes throughout the cell cycle and that multiple chromatin domains are organized around the same transcription factory.

  11. Discovering Echinococcus granulosus thioredoxin glutathione reductase inhibitors through site-specific dynamic combinatorial chemistry.

    PubMed

    Saiz, Cecilia; Castillo, Valerie; Fontán, Pablo; Bonilla, Mariana; Salinas, Gustavo; Rodríguez-Haralambides, Alejandra; Mahler, S Graciela

    2014-02-01

    In this study, we report a strategy using dynamic combinatorial chemistry for targeting the thioredoxin (Trx)-reductase catalytic site on Trx glutathione reductase (TGR), a pyridine nucleotide thiol-disulfide oxido-reductase. We chose Echinococcus granulosus TGR since it is a bottleneck enzyme of platyhelminth parasites and a validated pharmacological target. A dynamic combinatorial library (DCL) was constructed based on thiol-disulfide reversible exchange. We demonstrate the use of 5-thio-2-nitrobenzoic acid (TNB) as a non-covalent anchor fragment in a DCL templated by E. granulosus TGR. The heterodimer of TNB and bisthiazolidine (2af) was identified, upon library analysis by HPLC (IC50 = 24 μM). Furthermore, 14 analogs were synthetically prepared and evaluated against TGR. This allowed the study of a structure-activity relationship and the identification of a disulfide TNB-tricyclic bisthiazolidine (2aj) as the best enzyme inhibitor in these series, with an IC50 = 24 μM. Thus, our results validate the use of DCL for targeting thiol-disulfide oxido-reductases.

  12. Dynamic temperature fields under Mars landing sites and implications for supporting microbial life.

    PubMed

    Ulrich, Richard; Kral, Tim; Chevrier, Vincent; Pilgrim, Robert; Roe, Larry

    2010-01-01

    While average temperatures on Mars may be too low to support terrestrial life-forms or aqueous liquids, diurnal peak temperatures over most of the planet can be high enough to provide for both, down to a few centimeters beneath the surface for some fraction of the time. A thermal model was applied to the Viking 1, Viking 2, Pathfinder, Spirit, and Opportunity landing sites to demonstrate the dynamic temperature fields under the surface at these well-characterized locations. A benchmark temperature of 253 K was used as a lower limit for possible metabolic activity, which corresponds to the minimum found for specific terrestrial microorganisms. Aqueous solutions of salts known to exist on Mars can provide liquid solutions well below this temperature. Thermal modeling has shown that 253 K is reached beneath the surface at diurnal peak heating for at least some parts of the year at each of these landing sites. Within 40 degrees of the equator, 253 K beneath the surface should occur for at least some fraction of the year; and, within 20 degrees , it will be seen for most of the year. However, any life-form that requires this temperature to thrive must also endure daily excursions to far colder temperatures as well as periods of the year where 253 K is never reached at all.

  13. Current activities handbook: formerly utilized sites remedial action program

    SciTech Connect

    1981-02-27

    This volume is one of a series produced under contract with the DOE, by Politech Corporation to develop a legislative and regulatory data base to assist the FUSRAP management in addressing the institutional and socioeconomic issues involved in carrying out the Formerly Utilized Sites Remedial Action Program. This Information Handbook series contains information about all relevant government agencies at the Federal and state levels, the pertinent programs they administer, each affected state legislature, and current Federal and state legislative and regulatory initiatives. This volume is a compilation of information about the activities each of the thirteen state legislatures potentially affected by the Formerly Utilized Sites Remedial Action Program. It contains a description of the state legislative procedural rules and a schedule of each legislative session; a summary of pending relevant legislation; the name and telephone number of legislative and state agency contacts; and the full text of all bills identified.

  14. Dynamical quorum sensing and clustering dynamics in a population of spatially distributed active rotators

    NASA Astrophysics Data System (ADS)

    Sakaguchi, Hidetsugu; Maeyama, Satomi

    2013-02-01

    A model of clustering dynamics is proposed for a population of spatially distributed active rotators. A transition from excitable to oscillatory dynamics is induced by the increase of the local density of active rotators. It is interpreted as dynamical quorum sensing. In the oscillation regime, phase waves propagate without decay, which generates an effectively long-range interaction in the clustering dynamics. The clustering process becomes facilitated and only one dominant cluster appears rapidly as a result of the dynamical quorum sensing. An exact localized solution is found to a simplified model equation, and the competitive dynamics between two localized states is studied numerically.

  15. Electrostatic fields in the active sites of lysozymes.

    PubMed

    Sun, D P; Liao, D I; Remington, S J

    1989-07-01

    Considerable experimental evidence is in support of several aspects of the mechanism that has been proposed for the catalytic activity of lysozyme. However, the enzymatically catalyzed hydrolysis of polysaccharides proceeds over 5 orders of magnitude faster than that of model compounds that mimic the configuration of the substrate in the active site of the enzyme. Although several possible explanations for this rate enhancement have been discussed elsewhere, a definitive mechanism has not emerged. Here we report striking results obtained by classical electrodynamics, which suggest that bond breakage and the consequent separation of charge in lysozyme is promoted by a large electrostatic field across the active site cleft, produced in part by a very asymmetric distribution of charged residues on the enzyme surface. Lysozymes unrelated in amino acid sequence have similar distributions of charged residues and electric fields. The results reported here suggest that the electrostatic component of the rate enhancement is greater than 9 kcal.mol-1. Thus, electrostatic interactions may play a more important role in the enzymatic mechanism than has generally been appreciated.

  16. Histidine at the active site of Neurospora tyrosinase.

    PubMed

    Pfiffner, E; Lerch, K

    1981-10-13

    The involvement of histidyl residues as potential ligands to the binuclear active-site copper of Neurospora tyrosinase was explored by dye-sensitized photooxidation. The enzymatic activity of the holoenzyme was shown to be unaffected by exposure to light in the presence of methylene blue; however, irradiation of the apoenzyme under the same conditions led to a progressive loss of its ability to be reactivated with Cu2+. This photoinactivation was paralleled by a decrease in the histidine content whereas the number of histidyl residues in the holoenzyme remained constant. Copper measurements of photooxidized, reconstituted apoenzyme demonstrated the loss of binding of one copper atom per mole of enzyme as a consequence of photosensitized oxidation of three out of nine histidine residues. Their sequence positions were determined by a comparison of the relative yields of the histidine containing peptides of photooxidized holo- and apotyrosinases. The data obtained show the preferential modification of histidyl residues 188, 193, and 289 and suggest that they constitute metal ligands to one of the two active-site copper atoms. Substitution of copper by cobalt was found to afford complete protection of the histidyl residues from being modified by dye-sensitized photooxidation. PMID:6458322

  17. Effects of Pacing Site and Stimulation History on Alternans Dynamics and the Development of Complex Spatiotemporal Patterns in Cardiac Tissue

    PubMed Central

    Gizzi, Alessio; Cherry, Elizabeth M.; Gilmour, Robert F.; Luther, Stefan; Filippi, Simonetta; Fenton, Flavio H.

    2013-01-01

    Alternans of action potential duration has been associated with T wave alternans and the development of arrhythmias because it produces large gradients of repolarization. However, little is known about alternans dynamics in large mammalian hearts. Using optical mapping to record electrical activations simultaneously from the epicardium and endocardium of 9 canine right ventricles, we demonstrate novel arrhythmogenic complex spatiotemporal dynamics. (i) Alternans predominantly develops first on the endocardium. (ii) The postulated simple progression from normal rhythm to concordant to discordant alternans is not always observed; concordant alternans can develop from discordant alternans as the pacing period is decreased. (iii) In contrast to smaller tissue preparations, multiple stationary nodal lines may exist and need not be perpendicular to the pacing site or to each other. (iv) Alternans has fully three-dimensional dynamics and the epicardium and endocardium can show significantly different dynamics: multiple nodal surfaces can be transmural or intramural and can form concave/convex surfaces resulting in islands of discordant alternans. (v) The complex spatiotemporal patterns observed during alternans are very sensitive to both the site of stimulation and the stimulation history. Alternans in canine ventricles not only exhibit larger amplitudes and persist for longer cycle length regimes compared to those found in smaller mammalian hearts, but also show novel dynamics not previously described that enhance dispersion and show high sensitivity to initial conditions. This indicates some underlying predisposition to chaos and can help to guide the design of new drugs and devices controlling and preventing arrhythmic events. PMID:23637684

  18. Trichodiene synthase. Identification of active site residues by site-directed mutagenesis.

    PubMed

    Cane, D E; Shim, J H; Xue, Q; Fitzsimons, B C; Hohn, T M

    1995-02-28

    Derivatization of 5,5'-dithiobis(2-nitrobenzoic acid)-treated trichodiene synthase with [methyl-14C]methyl methanethiosulfonate and analysis of the derived tryptic peptides suggested the presence of two cysteine residues at the active site. The corresponding C146A and C190A mutants were constructed by site-directed mutagenesis. The C190A mutant displayed partial but significantly reduced activity, with a reduction in kcat/Km of 3000 compared to the wild-type trichodiene synthase, while the C146A mutant was essentially inactive. A hybrid trichodiene synthase, constructed from amino acids 1-309 of the Fusarium sporotrichioides enzyme and amino acids 310-383 of the Gibberella pulicaris cyclase, had steady state kinetic parameters nearly identical to those of the wild-type F. sporotrichioides enzyme. From this parent hybrid, a series of mutants was constructed by site-directed mutagenesis in which the amino acids in the base-rich region, 302-306 (DRRYR), were systematically modified. Three of these mutants were overexpressed and purified to homogeneity. The importance of Arg304 for catalysis was established by the observation that the R304K mutant showed a more than 25-fold increase in Km, as well as a 200-fold reduction in kcat. In addition, analysis of the incubation products of the R304K mutant by gas chromatography-mass spectrometry (GC-MS) indicated that farnesyl diphosphate was converted not only to trichodiene but to at least two additional C15H24 hydrocarbons, mle 204. Replacement of the Tyr305 residue of trichodiene synthase with Phe had little effect on kcat, while increasing the Km by a factor of ca. 7-8.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7873527

  19. The copper active site of CBM33 polysaccharide oxygenases.

    PubMed

    Hemsworth, Glyn R; Taylor, Edward J; Kim, Robbert Q; Gregory, Rebecca C; Lewis, Sally J; Turkenburg, Johan P; Parkin, Alison; Davies, Gideon J; Walton, Paul H

    2013-04-24

    The capacity of metal-dependent fungal and bacterial polysaccharide oxygenases, termed GH61 and CBM33, respectively, to potentiate the enzymatic degradation of cellulose opens new possibilities for the conversion of recalcitrant biomass to biofuels. GH61s have already been shown to be unique metalloenzymes containing an active site with a mononuclear copper ion coordinated by two histidines, one of which is an unusual τ-N-methylated N-terminal histidine. We now report the structural and spectroscopic characterization of the corresponding copper CBM33 enzymes. CBM33 binds copper with high affinity at a mononuclear site, significantly stabilizing the enzyme. X-band EPR spectroscopy of Cu(II)-CBM33 shows a mononuclear type 2 copper site with the copper ion in a distorted axial coordination sphere, into which azide will coordinate as evidenced by the concomitant formation of a new absorption band in the UV/vis spectrum at 390 nm. The enzyme's three-dimensional structure contains copper, which has been photoreduced to Cu(I) by the incident X-rays, confirmed by X-ray absorption/fluorescence studies of both aqueous solution and intact crystals of Cu-CBM33. The single copper(I) ion is ligated in a T-shaped configuration by three nitrogen atoms from two histidine side chains and the amino terminus, similar to the endogenous copper coordination geometry found in fungal GH61. PMID:23540833

  20. Activation of muscarinic acetylcholine receptors via their allosteric binding sites.

    PubMed Central

    Jakubík, J; Bacáková, L; Lisá, V; el-Fakahany, E E; Tucek, S

    1996-01-01

    Ligands that bind to the allosteric-binding sites on muscarinic acetylcholine receptors alter the conformation of the classical-binding sites of these receptors and either diminish or increase their affinity for muscarinic agonists and classical antagonists. It is not known whether the resulting conformational change also affects the interaction between the receptors and the G proteins. We have now found that the muscarinic receptor allosteric modulators alcuronium, gallamine, and strychnine (acting in the absence of an agonist) alter the synthesis of cAMP in Chinese hamster ovary (CHO) cells expressing the M2 or the M4 subtype of muscarinic receptors in the same direction as the agonist carbachol. In addition, most of their effects on the production of inositol phosphates in CHO cells expressing the M1 or the M3 muscarinic receptor subtypes are also similar to (although much weaker than) those of carbachol. The agonist-like effects of the allosteric modulators are not observed in CHO cells that have not been transfected with the gene for any of the subtypes of muscarinic receptors. The effects of alcuronium on the formation of cAMP and inositol phosphates are not prevented by the classical muscarinic antagonist quinuclidinyl benzilate. These observations demonstrate for the first time that the G protein-mediated functional responses of muscarinic receptors can be evoked not only from their classical, but also from their allosteric, binding sites. This represents a new mechanism of receptor activation. PMID:8710935

  1. Estimating groundwater dynamics at a Colorado River floodplain site using historical hydrological data and climate information

    NASA Astrophysics Data System (ADS)

    Chen, Jinsong; Hubbard, Susan S.; Williams, Kenneth H.; Ficklin, Darren L.

    2016-03-01

    Long-term prediction of groundwater dynamics is important for assessing water resources and their impacts on biogeochemical cycling. However, estimating future groundwater dynamics is challenging due to the wide range of spatiotemporal scales in hydrological processes and uncertainty in future climate conditions. In this study, we develop a Bayesian model to combine small-scale historical hydrological data with large-scale climate information to estimate groundwater dynamics at a floodplain site in Rifle, Colorado. Although we have only a few years of groundwater elevation measurements, we have 47 years of streamflow data from a gaging station approximately 43 km upstream and long-term climate prediction on the Upper Colorado River Basin. To estimate future daily groundwater dynamics, we first develop a time series model to downscale the monthly streamflow derived from climate information to daily streamflow, and then transform the daily streamflow to groundwater dynamics at the downstream floodplain site. We use Monte Carlo methods to estimate future groundwater dynamics at the site through sampling from the joint posterior probability distribution. The results suggest that although future groundwater levels are expected to be similar to the current levels, the timing of the high groundwater levels is predicted to occur about 1 month earlier. The developed framework is extendable to other sites to estimate future groundwater dynamics given disparate data sets and climate projections. Additionally, the obtained estimates are being used as input to a site-specific watershed reactive transport models to predict how climate-induced changes will influence future biogeochemical cycling relevant to a variety of ecosystem services.

  2. Radiation inactivation study of aminopeptidase: probing the active site

    NASA Astrophysics Data System (ADS)

    Jamadar, V. K.; Jamdar, S. N.; Mohan, Hari; Dandekar, S. P.; Harikumar, P.

    2004-04-01

    Ionizing radiation inactivated purified chicken intestinal aminopeptidase in media saturated with gases in the order N 2O>N 2>air. The D 37 values in the above conditions were 281, 210 and 198 Gy, respectively. OH radical scavengers such as t-butanol and isopropanol effectively nullified the radiation-induced damage in N 2O. The radicals (SCN) 2•-, Br 2•- and I 2•- inactivated the enzyme, pointing to the involvement of aromatic amino acids and cysteine in its catalytic activity. The enzyme exhibited fluorescence emission at 340 nm which is characteristic of tryptophan. The radiation-induced loss of activity was accompanied by a decrease in the fluorescence of the enzyme suggesting a predominant influence on tryptophan residues. The enzyme inhibition was associated with a marked increase in the Km and a decrease in the Vmax and kcat values, suggesting an irreversible alteration in the catalytic site. The above observations were confirmed by pulse radiolysis studies.

  3. Mimicking enzymatic active sites on surfaces for energy conversion chemistry.

    PubMed

    Gutzler, Rico; Stepanow, Sebastian; Grumelli, Doris; Lingenfelder, Magalí; Kern, Klaus

    2015-07-21

    Metal-organic supramolecular chemistry on surfaces has matured to a point where its underlying growth mechanisms are well understood and structures of defined coordination environments of metal atoms can be synthesized in a controlled and reproducible procedure. With surface-confined molecular self-assembly, scientists have a tool box at hand which can be used to prepare structures with desired properties, as for example a defined oxidation number and spin state of the transition metal atoms within the organic matrix. From a structural point of view, these coordination sites in the supramolecular structure resemble the catalytically active sites of metallo-enzymes, both characterized by metal centers coordinated to organic ligands. Several chemical reactions take place at these embedded metal ions in enzymes and the question arises whether these reactions also take place using metal-organic networks as catalysts. Mimicking the active site of metal atoms and organic ligands of enzymes in artificial systems is the key to understanding the selectivity and efficiency of enzymatic reactions. Their catalytic activity depends on various parameters including the charge and spin configuration in the metal ion, but also on the organic environment, which can stabilize intermediate reaction products, inhibits catalytic deactivation, and serves mostly as a transport channel for the reactants and products and therefore ensures the selectivity of the enzyme. Charge and spin on the transition metal in enzymes depend on the one hand on the specific metal element, and on the other hand on its organic coordination environment. These two parameters can carefully be adjusted in surface confined metal-organic networks, which can be synthesized by virtue of combinatorial mixing of building synthons. Different organic ligands with varying functional groups can be combined with several transition metals and spontaneously assemble into ordered networks. The catalytically active metal

  4. Spectroscopic Definition of the Ferroxidase Site in M Ferritin: Comparison of Binuclear Substrate vs. Cofactor Active Sites

    PubMed Central

    Schwartz, Jennifer K.; Liu, Xiaofeng S.; Tosha, Takehiko; Theil, Elizabeth C.; Solomon, Edward I.

    2008-01-01

    Maxi ferritins, 24 subunit protein nanocages, are essential in humans, plants, bacteria, and other animals for the concentration and storage of iron as hydrated ferric oxide, while minimizing free radical generation or use by pathogens. Formation of the precursors to these ferric oxides is catalyzed at a non-heme biferrous substrate site, which has some parallels with the cofactor sites in other biferrous enzymes. A combination of circular dichroism (CD), magnetic circular dichroism (MCD), and variable-temperature, variable-field MCD (VTVH MCD) has been used to probe Fe(II) binding to the substrate active site in frog M ferritin. These data determined that the active site within each subunit consists of two inequivalent five-coordinate (5C) ferrous centers that are weakly anti-ferromagnetically coupled, consistent with a μ-1,3 carboxylate bridge. The active site ligand set is unusual and likely includes a terminal water bound to each Fe(II) center. The Fe(II) ions bind to the active sites in a concerted manner, and cooperativity among the sites in each subunit is observed, potentially providing a mechanism for the control of ferritin iron loading. Differences in geometric and electronic structure – including a weak ligand field, availability of two water ligands at the biferrous substrate site, and the single carboxylate bridge in ferritin – coincide with the divergent reaction pathways observed between this substrate site and the previously studied cofactor active sites. PMID:18576633

  5. An active-site lysine in avian liver phosphoenolpyruvate carboxykinase

    SciTech Connect

    Guidinger, P.F.; Nowak, T. )

    1991-09-10

    The participation of lysine in the catalysis by avian liver phosphoenolpyruvate carboxykinase was studied by chemical modification and by a characterization of the modified enzyme. The rate of inactivation by 2,4-pentanedione is pseudo-first-order and linearly dependent on reagent concentration with a second-order rate constant of 0.36 {plus minus} 0.025 M{sup {minus}1} min{sup {minus}1}. Inactivation by pyridoxal 5{prime}-phosphate of the reversible reaction catalyzed by phosphoenolpyruvate carboxykinase follows bimolecular kinetics with a second-order rate constant of 7,700 {plus minus} 860 m{sup {minus}1} min{sup {minus}1}. Treatment of the enzyme or one lysine residue modified concomitant with 100% loss in activity. A stoichiometry of 1:1 is observed when either the reversible or the irreversible reactions catalyzed by the enzyme are monitored. A study of k{sub obs} vs pH suggests this active-site lysine has a pK{sub a} of 8.1 and a pH-independent rate constant of inactivation of 47,700 m{sup {minus}1} min{sup {minus}1}. Proton relaxation rate measurements suggest that pyridoxal 5{prime}-phosphate modification alters binding of the phosphate-containing substrates. {sup 31}P NMR relaxation rate measurements show altered binding of the substrates in the ternary enzyme {center dot}Mn{sup 2+}{center dot}substrate complex. Circular dichroism studies show little change in secondary structure of pyridoxal 5{prime}-phosphate modified phosphoenolpyruvate carboxykinase. These results indicate that avian liver phosphoenolpyruvate carboxykinase has one reactive lysine at the active site and it is involved in the binding and activation of the phosphate-containing substrates.

  6. Investigation of nonlinear dynamic soil property at the Savannah River Site

    SciTech Connect

    Lee, R.C.

    2000-01-17

    This document summarizes laboratory dynamic soil testing investigations conducted by the University of Texas at Austin (UTA) for the Savannah River Site (SRS) (Stokoe et al., 1995a, Stokoe et al., 1995b, Sponseller and Stokoe, 1995). The purpose of the investigation is to provide an evaluation of past testing results in the context of new test data and the development of consistent site wide models of material strain dependencies based upon geologic formation, depth, and relevant index properties.

  7. Overview af MSFC's Applied Fluid Dynamics Analysis Group Activities

    NASA Technical Reports Server (NTRS)

    Garcia, Roberto; Griffin, Lisa; Williams, Robert

    2004-01-01

    This paper presents viewgraphs on NASA Marshall Space Flight Center's Applied Fluid Dynamics Analysis Group Activities. The topics include: 1) Status of programs at MSFC; 2) Fluid Mechanics at MSFC; 3) Relevant Fluid Dynamics Activities at MSFC; and 4) Shuttle Return to Flight.

  8. Perchlorate Reductase Is Distinguished by Active Site Aromatic Gate Residues.

    PubMed

    Youngblut, Matthew D; Tsai, Chi-Lin; Clark, Iain C; Carlson, Hans K; Maglaqui, Adrian P; Gau-Pan, Phonchien S; Redford, Steven A; Wong, Alan; Tainer, John A; Coates, John D

    2016-04-22

    Perchlorate is an important ion on both Earth and Mars. Perchlorate reductase (PcrAB), a specialized member of the dimethylsulfoxide reductase superfamily, catalyzes the first step of microbial perchlorate respiration, but little is known about the biochemistry, specificity, structure, and mechanism of PcrAB. Here we characterize the biophysics and phylogeny of this enzyme and report the 1.86-Å resolution PcrAB complex crystal structure. Biochemical analysis revealed a relatively high perchlorate affinity (Km = 6 μm) and a characteristic substrate inhibition compared with the highly similar respiratory nitrate reductase NarGHI, which has a relatively much lower affinity for perchlorate (Km = 1.1 mm) and no substrate inhibition. Structural analysis of oxidized and reduced PcrAB with and without the substrate analog SeO3 (2-) bound to the active site identified key residues in the positively charged and funnel-shaped substrate access tunnel that gated substrate entrance and product release while trapping transiently produced chlorate. The structures suggest gating was associated with shifts of a Phe residue between open and closed conformations plus an Asp residue carboxylate shift between monodentate and bidentate coordination to the active site molybdenum atom. Taken together, structural and mutational analyses of gate residues suggest key roles of these gate residues for substrate entrance and product release. Our combined results provide the first detailed structural insight into the mechanism of biological perchlorate reduction, a critical component of the chlorine redox cycle on Earth.

  9. Eel calcitonin binding site distribution and antinociceptive activity in rats

    SciTech Connect

    Guidobono, F.; Netti, C.; Sibilia, V.; Villa, I.; Zamboni, A.; Pecile, A.

    1986-03-01

    The distribution of binding site for (/sup 125/I)-eel-calcitonin (ECT) to rat central nervous system, studied by an autoradiographic technique, showed concentrations of binding in the diencephalon, the brain stem and the spinal cord. Large accumulations of grains were seen in the hypothalamus, the amygdala, in the fasciculus medialis prosencephali, in the fasciculus longitudinalis medialis, in the ventrolateral part of the periventricular gray matter, in the lemniscus medialis and in the raphe nuclei. The density of grains in the reticular formation and in the nucleus tractus spinalis nervi trigemini was more moderate. In the spinal cord, grains were scattered throughout the dorsal horns. Binding of the ligand was displaced equally by cold ECT and by salmon CT(sCT), indicating that both peptides bind to the same receptors. Human CT was much weaker than sCT in displacing (/sup 125/I)-ECT binding. The administration of ECT into the brain ventricles of rats dose-dependently induced a significant and long-lasting enhancement of hot-plate latencies comparable with that obtained with sCT. The antinociceptive activity induced by ECT is compatible with the topographical distribution of binding sites for the peptide and is a further indication that fish CTs are active in the mammalian brain.

  10. Perchlorate Reductase Is Distinguished by Active Site Aromatic Gate Residues.

    PubMed

    Youngblut, Matthew D; Tsai, Chi-Lin; Clark, Iain C; Carlson, Hans K; Maglaqui, Adrian P; Gau-Pan, Phonchien S; Redford, Steven A; Wong, Alan; Tainer, John A; Coates, John D

    2016-04-22

    Perchlorate is an important ion on both Earth and Mars. Perchlorate reductase (PcrAB), a specialized member of the dimethylsulfoxide reductase superfamily, catalyzes the first step of microbial perchlorate respiration, but little is known about the biochemistry, specificity, structure, and mechanism of PcrAB. Here we characterize the biophysics and phylogeny of this enzyme and report the 1.86-Å resolution PcrAB complex crystal structure. Biochemical analysis revealed a relatively high perchlorate affinity (Km = 6 μm) and a characteristic substrate inhibition compared with the highly similar respiratory nitrate reductase NarGHI, which has a relatively much lower affinity for perchlorate (Km = 1.1 mm) and no substrate inhibition. Structural analysis of oxidized and reduced PcrAB with and without the substrate analog SeO3 (2-) bound to the active site identified key residues in the positively charged and funnel-shaped substrate access tunnel that gated substrate entrance and product release while trapping transiently produced chlorate. The structures suggest gating was associated with shifts of a Phe residue between open and closed conformations plus an Asp residue carboxylate shift between monodentate and bidentate coordination to the active site molybdenum atom. Taken together, structural and mutational analyses of gate residues suggest key roles of these gate residues for substrate entrance and product release. Our combined results provide the first detailed structural insight into the mechanism of biological perchlorate reduction, a critical component of the chlorine redox cycle on Earth. PMID:26940877

  11. How Force Might Activate Talin's Vinculin Binding Sites: SMD Reveals a Structural Mechanism

    PubMed Central

    Hytönen, Vesa P; Vogel, Viola

    2008-01-01

    Upon cell adhesion, talin physically couples the cytoskeleton via integrins to the extracellular matrix, and subsequent vinculin recruitment is enhanced by locally applied tensile force. Since the vinculin binding (VB) sites are buried in the talin rod under equilibrium conditions, the structural mechanism of how vinculin binding to talin is force-activated remains unknown. Taken together with experimental data, a biphasic vinculin binding model, as derived from steered molecular dynamics, provides high resolution structural insights how tensile mechanical force applied to the talin rod fragment (residues 486–889 constituting helices H1–H12) might activate the VB sites. Fragmentation of the rod into three helix subbundles is prerequisite to the sequential exposure of VB helices to water. Finally, unfolding of a VB helix into a completely stretched polypeptide might inhibit further binding of vinculin. The first events in fracturing the H1–H12 rods of talin1 and talin2 in subbundles are similar. The proposed force-activated α-helix swapping mechanism by which vinculin binding sites in talin rods are exposed works distinctly different from that of other force-activated bonds, including catch bonds. PMID:18282082

  12. An Active Site Water Network in the Plasminogen Activator Pla from Yersinia pestis

    SciTech Connect

    Eren, Elif; Murphy, Megan; Goguen, Jon; van den Berg, Bert

    2010-08-13

    The plasminogen activator Pla from Yersinia pestis is an outer membrane protease (omptin) that is important for the virulence of plague. Here, we present the high-resolution crystal structure of wild-type, enzymatically active Pla at 1.9 {angstrom}. The structure shows a water molecule located between active site residues D84 and H208, which likely corresponds to the nucleophilic water. A number of other water molecules are present in the active site, linking residues important for enzymatic activity. The R211 sidechain in loop L4 is close to the nucleophilic water and possibly involved in the stabilization of the oxyanion intermediate. Subtle conformational changes of H208 result from the binding of lipopolysaccharide to the outside of the barrel, explaining the unusual dependence of omptins on lipopolysaccharide for activity. The Pla structure suggests a model for the interaction with plasminogen substrate and provides a more detailed understanding of the catalytic mechanism of omptin proteases.

  13. Endolysosomes Are the Principal Intracellular Sites of Acid Hydrolase Activity.

    PubMed

    Bright, Nicholas A; Davis, Luther J; Luzio, J Paul

    2016-09-12

    The endocytic delivery of macromolecules from the mammalian cell surface for degradation by lysosomal acid hydrolases requires traffic through early endosomes to late endosomes followed by transient (kissing) or complete fusions between late endosomes and lysosomes. Transient or complete fusion results in the formation of endolysosomes, which are hybrid organelles from which lysosomes are re-formed. We have used synthetic membrane-permeable cathepsin substrates, which liberate fluorescent reporters upon proteolytic cleavage, as well as acid phosphatase cytochemistry to identify which endocytic compartments are acid hydrolase active. We found that endolysosomes are the principal organelles in which acid hydrolase substrates are cleaved. Endolysosomes also accumulated acidotropic probes and could be distinguished from terminal storage lysosomes, which were acid hydrolase inactive and did not accumulate acidotropic probes. Using live-cell microscopy, we have demonstrated that fusion events, which form endolysosomes, precede the onset of acid hydrolase activity. By means of sucrose and invertase uptake experiments, we have also shown that acid-hydrolase-active endolysosomes and acid-hydrolase-inactive, terminal storage lysosomes exist in dynamic equilibrium. We conclude that the terminal endocytic compartment is composed of acid-hydrolase-active, acidic endolysosomes and acid hydrolase-inactive, non-acidic, terminal storage lysosomes, which are linked and function in a lysosome regeneration cycle. PMID:27498570

  14. Active Sites Environmental Monitoring Program: Program plan. Revision 1

    SciTech Connect

    Ashwood, T.L.; Wickliff, D.S.; Morrissey, C.M.

    1992-02-01

    The Active Sites Environmental Monitoring Program (ASEMP), initiated in 1989, provides early detection and performance monitoring of transuranic (TRU) waste and active low-level waste (LLW) facilities at Oak Ridge National Laboratory (ORNL) in accordance with US Department of Energy (DOE) Order 5820.2A. Active LLW facilities in Solid Waste Storage Area (SWSA) 6 include Tumulus I and Tumulus II, the Interim Waste Management Facility (IWMF), LLW silos, high-range wells, asbestos silos, and fissile wells. The tumulus pads and IWMF are aboveground, high-strength concrete pads on which concrete vaults containing metal boxes of LLW are placed; the void space between the boxes and vaults is filled with grout. Eventually, these pads and vaults will be covered by an engineered multilayered cap. All other LLW facilities in SWSA 6 are below ground. In addition, this plan includes monitoring of the Hillcut Disposal Test Facility (HDTF) in SWSA 6, even though this facility was completed prior to the data of the DOE order. In SWSA 5 North, the TRU facilities include below-grade engineered caves, high-range wells, and unlined trenches. All samples from SWSA 6 are screened for alpha and beta activity, counted for gamma-emitting isotopes, and analyzed for tritium. In addition to these analytes, samples from SWSA 5 North are analyzed for specific transuranic elements.

  15. Individual and group dynamics in purchasing activity

    NASA Astrophysics Data System (ADS)

    Gao, Lei; Guo, Jin-Li; Fan, Chao; Liu, Xue-Jiao

    2013-01-01

    As a major part of the daily operation in an enterprise, purchasing frequency is in constant change. Recent approaches on the human dynamics can provide some new insights into the economic behavior of companies in the supply chain. This paper captures the attributes of creation times of purchase orders to an individual vendor, as well as to all vendors, and further investigates whether they have some kind of dynamics by applying logarithmic binning to the construction of distribution plots. It’s found that the former displays a power-law distribution with approximate exponent 2.0, while the latter is fitted by a mixture distribution with both power-law and exponential characteristics. Obviously, two distinctive characteristics are presented for the interval time distribution from the perspective of individual dynamics and group dynamics. Actually, this mixing feature can be attributed to the fitting deviations as they are negligible for individual dynamics, but those of different vendors are cumulated and then lead to an exponential factor for group dynamics. To better describe the mechanism generating the heterogeneity of the purchase order assignment process from the objective company to all its vendors, a model driven by product life cycle is introduced, and then the analytical distribution and the simulation result are obtained, which are in good agreement with the empirical data.

  16. Active Site and Laminarin Binding in Glycoside Hydrolase Family 55*

    PubMed Central

    Bianchetti, Christopher M.; Takasuka, Taichi E.; Deutsch, Sam; Udell, Hannah S.; Yik, Eric J.; Bergeman, Lai F.; Fox, Brian G.

    2015-01-01

    The Carbohydrate Active Enzyme (CAZy) database indicates that glycoside hydrolase family 55 (GH55) contains both endo- and exo-β-1,3-glucanases. The founding structure in the GH55 is PcLam55A from the white rot fungus Phanerochaete chrysosporium (Ishida, T., Fushinobu, S., Kawai, R., Kitaoka, M., Igarashi, K., and Samejima, M. (2009) Crystal structure of glycoside hydrolase family 55 β-1,3-glucanase from the basidiomycete Phanerochaete chrysosporium. J. Biol. Chem. 284, 10100–10109). Here, we present high resolution crystal structures of bacterial SacteLam55A from the highly cellulolytic Streptomyces sp. SirexAA-E with bound substrates and product. These structures, along with mutagenesis and kinetic studies, implicate Glu-502 as the catalytic acid (as proposed earlier for Glu-663 in PcLam55A) and a proton relay network of four residues in activating water as the nucleophile. Further, a set of conserved aromatic residues that define the active site apparently enforce an exo-glucanase reactivity as demonstrated by exhaustive hydrolysis reactions with purified laminarioligosaccharides. Two additional aromatic residues that line the substrate-binding channel show substrate-dependent conformational flexibility that may promote processive reactivity of the bound oligosaccharide in the bacterial enzymes. Gene synthesis carried out on ∼30% of the GH55 family gave 34 active enzymes (19% functional coverage of the nonredundant members of GH55). These active enzymes reacted with only laminarin from a panel of 10 different soluble and insoluble polysaccharides and displayed a broad range of specific activities and optima for pH and temperature. Application of this experimental method provides a new, systematic way to annotate glycoside hydrolase phylogenetic space for functional properties. PMID:25752603

  17. Simultaneous fast measurement of circuit dynamics at multiple sites across the mammalian brain.

    PubMed

    Kim, Christina K; Yang, Samuel J; Pichamoorthy, Nandini; Young, Noah P; Kauvar, Isaac; Jennings, Joshua H; Lerner, Talia N; Berndt, Andre; Lee, Soo Yeun; Ramakrishnan, Charu; Davidson, Thomas J; Inoue, Masatoshi; Bito, Haruhiko; Deisseroth, Karl

    2016-04-01

    Real-time activity measurements from multiple specific cell populations and projections are likely to be important for understanding the brain as a dynamical system. Here we developed frame-projected independent-fiber photometry (FIP), which we used to record fluorescence activity signals from many brain regions simultaneously in freely behaving mice. We explored the versatility of the FIP microscope by quantifying real-time activity relationships among many brain regions during social behavior, simultaneously recording activity along multiple axonal pathways during sensory experience, performing simultaneous two-color activity recording, and applying optical perturbation tuned to elicit dynamics that match naturally occurring patterns observed during behavior.

  18. Active site loop conformation regulates promiscuous activity in a lactonase from Geobacillus kaustophilus HTA426.

    PubMed

    Zhang, Yu; An, Jiao; Yang, Guang-Yu; Bai, Aixi; Zheng, Baisong; Lou, Zhiyong; Wu, Geng; Ye, Wei; Chen, Hai-Feng; Feng, Yan; Manco, Giuseppe

    2015-01-01

    Enzyme promiscuity is a prerequisite for fast divergent evolution of biocatalysts. A phosphotriesterase-like lactonase (PLL) from Geobacillus kaustophilus HTA426 (GkaP) exhibits main lactonase and promiscuous phosphotriesterase activities. To understand its catalytic and evolutionary mechanisms, we investigated a "hot spot" in the active site by saturation mutagenesis as well as X-ray crystallographic analyses. We found that position 99 in the active site was involved in substrate discrimination. One mutant, Y99L, exhibited 11-fold improvement over wild-type in reactivity (kcat/Km) toward the phosphotriesterase substrate ethyl-paraoxon, but showed 15-fold decrease toward the lactonase substrate δ-decanolactone, resulting in a 157-fold inversion of the substrate specificity. Structural analysis of Y99L revealed that the mutation causes a ∼6.6 Å outward shift of adjacent loop 7, which may cause increased flexibility of the active site and facilitate accommodation and/or catalysis of organophosphate substrate. This study provides for the PLL family an example of how the evolutionary route from promiscuity to specificity can derive from very few mutations, which promotes alteration in the conformational adjustment of the active site loops, in turn draws the capacity of substrate binding and activity.

  19. Active Site Loop Conformation Regulates Promiscuous Activity in a Lactonase from Geobacillus kaustophilus HTA426

    PubMed Central

    Zhang, Yu; An, Jiao; Yang, Guang-Yu; Bai, Aixi; Zheng, Baisong; Lou, Zhiyong; Wu, Geng; Ye, Wei; Chen, Hai-Feng; Feng, Yan; Manco, Giuseppe

    2015-01-01

    Enzyme promiscuity is a prerequisite for fast divergent evolution of biocatalysts. A phosphotriesterase-like lactonase (PLL) from Geobacillus kaustophilus HTA426 (GkaP) exhibits main lactonase and promiscuous phosphotriesterase activities. To understand its catalytic and evolutionary mechanisms, we investigated a “hot spot” in the active site by saturation mutagenesis as well as X-ray crystallographic analyses. We found that position 99 in the active site was involved in substrate discrimination. One mutant, Y99L, exhibited 11-fold improvement over wild-type in reactivity (kcat/Km) toward the phosphotriesterase substrate ethyl-paraoxon, but showed 15-fold decrease toward the lactonase substrate δ-decanolactone, resulting in a 157-fold inversion of the substrate specificity. Structural analysis of Y99L revealed that the mutation causes a ∼6.6 Å outward shift of adjacent loop 7, which may cause increased flexibility of the active site and facilitate accommodation and/or catalysis of organophosphate substrate. This study provides for the PLL family an example of how the evolutionary route from promiscuity to specificity can derive from very few mutations, which promotes alteration in the conformational adjustment of the active site loops, in turn draws the capacity of substrate binding and activity. PMID:25706379

  20. Dynamic HypA zinc site is essential for acid viability and proper urease maturation in Helicobacter pylori

    PubMed Central

    Johnson, Ryan C.; Hu, Heidi Q.; Merrell, D. Scott; Maroney, Michael J.

    2015-01-01

    Helicobacter pylori requires urease activity in order to survive in the acid environment of the human stomach. Urease is regulated in part by nickelation, a process that requires the HypA protein, which is a putative nickel metallochaperone that is generally associated with hydrogenase maturation. However, in H. pylori, HypA plays a dual role. In addition to an N-terminal nickel binding site, HypA proteins also contain a structural zinc site that is coordinated by two rigorously conserved CXXC sequences, which in H. pylori are flanked by His residues. These structural Zn sites are known to be dynamic, converting from Zn(Cys)4 centers at pH 7.2 to Zn(Cys)2(His)2 centers at pH 6.3 in the presence of Ni(II) ions. In this study, mutant strains of H. pylori that express zinc site variants of the HypA protein are used to show that the structural changes in the zinc site are important for the acid viability of the bacterium, and that a reduction in acid viability in these variants can be traced in large measure to deficient urease activity. This in turn leads to a model that connects the Zn(Cys)4 coordination to urease maturation. PMID:25608738

  1. Extensive site-directed mutagenesis reveals interconnected functional units in the alkaline phosphatase active site.

    PubMed

    Sunden, Fanny; Peck, Ariana; Salzman, Julia; Ressl, Susanne; Herschlag, Daniel

    2015-01-01

    Enzymes enable life by accelerating reaction rates to biological timescales. Conventional studies have focused on identifying the residues that have a direct involvement in an enzymatic reaction, but these so-called 'catalytic residues' are embedded in extensive interaction networks. Although fundamental to our understanding of enzyme function, evolution, and engineering, the properties of these networks have yet to be quantitatively and systematically explored. We dissected an interaction network of five residues in the active site of Escherichia coli alkaline phosphatase. Analysis of the complex catalytic interdependence of specific residues identified three energetically independent but structurally interconnected functional units with distinct modes of cooperativity. From an evolutionary perspective, this network is orders of magnitude more probable to arise than a fully cooperative network. From a functional perspective, new catalytic insights emerge. Further, such comprehensive energetic characterization will be necessary to benchmark the algorithms required to rationally engineer highly efficient enzymes. PMID:25902402

  2. Extensive site-directed mutagenesis reveals interconnected functional units in the alkaline phosphatase active site.

    PubMed

    Sunden, Fanny; Peck, Ariana; Salzman, Julia; Ressl, Susanne; Herschlag, Daniel

    2015-01-01

    Enzymes enable life by accelerating reaction rates to biological timescales. Conventional studies have focused on identifying the residues that have a direct involvement in an enzymatic reaction, but these so-called 'catalytic residues' are embedded in extensive interaction networks. Although fundamental to our understanding of enzyme function, evolution, and engineering, the properties of these networks have yet to be quantitatively and systematically explored. We dissected an interaction network of five residues in the active site of Escherichia coli alkaline phosphatase. Analysis of the complex catalytic interdependence of specific residues identified three energetically independent but structurally interconnected functional units with distinct modes of cooperativity. From an evolutionary perspective, this network is orders of magnitude more probable to arise than a fully cooperative network. From a functional perspective, new catalytic insights emerge. Further, such comprehensive energetic characterization will be necessary to benchmark the algorithms required to rationally engineer highly efficient enzymes.

  3. Hetero-site-specific X-ray pump-probe spectroscopy for femtosecond intramolecular dynamics

    DOE PAGES

    Picón, A.; Lehmann, C. S.; Bostedt, C.; Rudenko, A.; Marinelli, A.; Osipov, T.; Rolles, D.; Berrah, N.; Bomme, C.; Bucher, M.; et al

    2016-05-23

    New capabilities at X-ray free-electron laser facilities allow the generation of two-colour femtosecond X-ray pulses, opening the possibility of performing ultrafast studies of X-ray-induced phenomena. Specifically, the experimental realization of hetero-site-specific X-ray-pump/X-ray-probe spectroscopy is of special interest, in which an X-ray pump pulse is absorbed at one site within a molecule and an X-ray probe pulse follows the X-ray-induced dynamics at another site within the same molecule. In this paper, we show experimental evidence of a hetero-site pump-probe signal. By using two-colour 10-fs X-ray pulses, we are able to observe the femtosecond time dependence for the formation of F ionsmore » during the fragmentation of XeF2 molecules following X-ray absorption at the Xe site.« less

  4. Hetero-site-specific X-ray pump-probe spectroscopy for femtosecond intramolecular dynamics

    PubMed Central

    Picón, A.; Lehmann, C. S.; Bostedt, C.; Rudenko, A.; Marinelli, A.; Osipov, T.; Rolles, D.; Berrah, N.; Bomme, C.; Bucher, M.; Doumy, G.; Erk, B.; Ferguson, K. R.; Gorkhover, T.; Ho, P. J.; Kanter, E. P.; Krässig, B.; Krzywinski, J.; Lutman, A. A.; March, A. M.; Moonshiram, D.; Ray, D.; Young, L.; Pratt, S. T.; Southworth, S. H.

    2016-01-01

    New capabilities at X-ray free-electron laser facilities allow the generation of two-colour femtosecond X-ray pulses, opening the possibility of performing ultrafast studies of X-ray-induced phenomena. Particularly, the experimental realization of hetero-site-specific X-ray-pump/X-ray-probe spectroscopy is of special interest, in which an X-ray pump pulse is absorbed at one site within a molecule and an X-ray probe pulse follows the X-ray-induced dynamics at another site within the same molecule. Here we show experimental evidence of a hetero-site pump-probe signal. By using two-colour 10-fs X-ray pulses, we are able to observe the femtosecond time dependence for the formation of F ions during the fragmentation of XeF2 molecules following X-ray absorption at the Xe site. PMID:27212390

  5. Rate of hydrolysis in ATP synthase is fine-tuned by α-subunit motif controlling active site conformation

    PubMed Central

    Beke-Somfai, Tamás; Lincoln, Per; Nordén, Bengt

    2013-01-01

    Computer-designed artificial enzymes will require precise understanding of how conformation of active sites may control barrier heights of key transition states, including dependence on structure and dynamics at larger molecular scale. FoF1 ATP synthase is interesting as a model system: a delicate molecular machine synthesizing or hydrolyzing ATP using a rotary motor. Isolated F1 performs hydrolysis with a rate very sensitive to ATP concentration. Experimental and theoretical results show that, at low ATP concentrations, ATP is slowly hydrolyzed in the so-called tight binding site, whereas at higher concentrations, the binding of additional ATP molecules induces rotation of the central γ-subunit, thereby forcing the site to transform through subtle conformational changes into a loose binding site in which hydrolysis occurs faster. How the 1-Å-scale rearrangements are controlled is not yet fully understood. By a combination of theoretical approaches, we address how large macromolecular rearrangements may manipulate the active site and how the reaction rate changes with active site conformation. Simulations reveal that, in response to γ-subunit position, the active site conformation is fine-tuned mainly by small α-subunit changes. Quantum mechanics-based results confirm that the sub-Ångström gradual changes between tight and loose binding site structures dramatically alter the hydrolysis rate. PMID:23345443

  6. Rate of hydrolysis in ATP synthase is fine-tuned by α-subunit motif controlling active site conformation.

    PubMed

    Beke-Somfai, Tamás; Lincoln, Per; Nordén, Bengt

    2013-02-01

    Computer-designed artificial enzymes will require precise understanding of how conformation of active sites may control barrier heights of key transition states, including dependence on structure and dynamics at larger molecular scale. F(o)F(1) ATP synthase is interesting as a model system: a delicate molecular machine synthesizing or hydrolyzing ATP using a rotary motor. Isolated F(1) performs hydrolysis with a rate very sensitive to ATP concentration. Experimental and theoretical results show that, at low ATP concentrations, ATP is slowly hydrolyzed in the so-called tight binding site, whereas at higher concentrations, the binding of additional ATP molecules induces rotation of the central γ-subunit, thereby forcing the site to transform through subtle conformational changes into a loose binding site in which hydrolysis occurs faster. How the 1-Å-scale rearrangements are controlled is not yet fully understood. By a combination of theoretical approaches, we address how large macromolecular rearrangements may manipulate the active site and how the reaction rate changes with active site conformation. Simulations reveal that, in response to γ-subunit position, the active site conformation is fine-tuned mainly by small α-subunit changes. Quantum mechanics-based results confirm that the sub-Ångström gradual changes between tight and loose binding site structures dramatically alter the hydrolysis rate. PMID:23345443

  7. Monitoring and Modeling Carbon Dynamics at a Network of Intensive Sites in the USA and Mexico

    NASA Astrophysics Data System (ADS)

    Birdsey, R.; Wayson, C.; Johnson, K. D.; Pan, Y.; Angeles, G.; De Jong, B. H.; Andrade, J. L.; Dai, Z.

    2013-05-01

    The Forest Services of the USA and Mexico, supported by NASA and USAID, have begun to establish a network of intensive forest carbon monitoring sites. These sites are used for research and teaching, developing forest management practices, and forging links to the needs of communities. Several of the sites have installed eddy flux towers to basic meteorology data and daily estimates of forest carbon uptake and release, the processes that determine forest growth. Field sampling locations at each site provide estimates of forest biomass and carbon stocks, and monitor forest dynamic processes such as growth and mortality rates. Remote sensing facilitates scaling up to the surrounding landscapes. The sites support information requirements for implementing programs such as Reducing Emissions from Deforestation and Forest Degradation (REDD+), enabling communities to receive payments for ecosystem services such as reduced carbon emissions or improved forest management. In addition to providing benchmark data for REDD+ projects, the sites are valuable for validating state and national estimates from satellite remote sensing and the national forest inventory. Data from the sites provide parameters for forest models that support strategic management analysis, and support student training and graduate projects. The intensive monitoring sites may be a model for other countries in Latin America. Coordination among sites in the USA, Mexico and other Latin American countries can ensure harmonization of approaches and data, and share experiences and knowledge among countries with emerging opportunities for implementing REDD+ and other conservation programs.

  8. Overview of MSFC's Applied Fluid Dynamics Analysis Group Activities

    NASA Technical Reports Server (NTRS)

    Garcia, Roberto; Griffin, Lisa; Williams, Robert

    2002-01-01

    This viewgraph report presents an overview of activities and accomplishments of NASA's Marshall Space Flight Center's Applied Fluid Dynamics Analysis Group. Expertise in this group focuses on high-fidelity fluids design and analysis with application to space shuttle propulsion and next generation launch technologies. Topics covered include: computational fluid dynamics research and goals, turbomachinery research and activities, nozzle research and activities, combustion devices, engine systems, MDA development and CFD process improvements.

  9. Free energy simulations of active-site mutants of dihydrofolate reductase.

    PubMed

    Doron, Dvir; Stojković, Vanja; Gakhar, Lokesh; Vardi-Kilshtain, Alexandra; Kohen, Amnon; Major, Dan Thomas

    2015-01-22

    This study employs hybrid quantum mechanics-molecular mechanics (QM/MM) simulations to investigate the effect of mutations of the active-site residue I14 of E. coli dihydrofolate reductase (DHFR) on the hydride transfer. Recent kinetic measurements of the I14X mutants (X = V, A, and G) indicated slower hydride transfer rates and increasingly temperature-dependent kinetic isotope effects (KIEs) with systematic reduction of the I14 side chain. The QM/MM simulations show that when the original isoleucine residue is substituted in silico by valine, alanine, or glycine (I14V, I14A, and I14G DHFR, respectively), the free energy barrier height of the hydride transfer reaction increases relative to the wild-type enzyme. These trends are in line with the single-turnover rate measurements reported for these systems. In addition, extended dynamics simulations of the reactive Michaelis complex reveal enhanced flexibility in the mutants, and in particular for the I14G mutant, including considerable fluctuations of the donor-acceptor distance (DAD) and the active-site hydrogen bonding network compared with those detected in the native enzyme. These observations suggest that the perturbations induced by the mutations partly impair the active-site environment in the reactant state. On the other hand, the average DADs at the transition state of all DHFR variants are similar. Crystal structures of I14 mutants (V, A, and G) confirmed the trend of increased flexibility of the M20 and other loops. PMID:25382260

  10. Structural and dynamic effects of cholesterol at preferred sites of interaction with rhodopsin identified from microsecond length molecular dynamics simulations

    PubMed Central

    Khelashvili, George; Grossfield, Alan; Feller, Scott E.; Pitman, Michael C.; Weinstein, Harel

    2014-01-01

    An unresolved question about GPCR function is the role of membrane components in receptor stability and activation. In particular, cholesterol is known to affect the function of membrane proteins, but the details of its effect on GPCRs are still elusive. Here, we describe how cholesterol modulates the behavior of the TM1-TM2-TM7-helix 8(H8) functional network that comprises the highly conserved NPxxY(x)5,6F motif, through specific interactions with the receptor. The inferences are based on the analysis of microsecond length molecular dynamics (MD) simulations of rhodopsin in an explicit membrane environment. Three regions on the rhodopsin exhibit the highest cholesterol density throughout the trajectory: the extracellular end of TM7, a location resembling the high-density sterol area from the electron microscopy data; the intracellular parts of TM1, TM2, and TM4, a region suggested as the cholesterol binding site in the recent X-ray crystallography data on β2-adrenergic GPCR; and the intracellular ends of TM2-TM3, a location that was categorized as the high cholesterol density area in multiple independent 100 ns MD simulations of the same system. We found that cholesterol primarily affects specific local perturbations of the helical TM domains such as the kinks in TM1, TM2, and TM7. These local distortions, in turn, relate to rigid-body motions of the TMs in the TM1-TM2-TM7-H8 bundle. The specificity of the effects stems from the nonuniform distribution of cholesterol around the protein. Through correlation analysis we connect local effects of cholesterol on structural perturbations with a regulatory role of cholesterol in the structural rearrangements involved in GPCR function. PMID:19173312

  11. Metals in the active site of native protein phosphatase-1.

    PubMed

    Heroes, Ewald; Rip, Jens; Beullens, Monique; Van Meervelt, Luc; De Gendt, Stefan; Bollen, Mathieu

    2015-08-01

    Protein phosphatase-1 (PP1) is a major protein Ser/Thr phosphatase in eukaryotic cells. Its activity depends on two metal ions in the catalytic site, which were identified as manganese in the bacterially expressed phosphatase. However, the identity of the metal ions in native PP1 is unknown. In this study, total reflection X-ray fluorescence (TXRF) was used to detect iron and zinc in PP1 that was purified from rabbit skeletal muscle. Metal exchange experiments confirmed that the distinct substrate specificity of recombinant and native PP1 is determined by the nature of their associated metals. We also found that the iron level associated with native PP1 is decreased by incubation with inhibitor-2, consistent with a function of inhibitor-2 as a PP1 chaperone. PMID:25890482

  12. Local Pheromone Release from Dynamic Polarity Sites Underlies Cell-Cell Pairing during Yeast Mating.

    PubMed

    Merlini, Laura; Khalili, Bita; Bendezú, Felipe O; Hurwitz, Daniel; Vincenzetti, Vincent; Vavylonis, Dimitrios; Martin, Sophie G

    2016-04-25

    Cell pairing is central for many processes, including immune defense, neuronal connection, hyphal fusion, and sexual reproduction. How does a cell orient toward a partner, especially when faced with multiple choices? Fission yeast Schizosaccharomyces pombe P and M cells, which respectively express P and M factor pheromones [1, 2], pair during the mating process induced by nitrogen starvation. Engagement of pheromone receptors Map3 and Mam2 [3, 4] with their cognate pheromone ligands leads to activation of the Gα protein Gpa1 to signal sexual differentiation [3, 5, 6]. Prior to cell pairing, the Cdc42 GTPase, a central regulator of cell polarization, forms dynamic zones of activity at the cell periphery at distinct locations over time [7]. Here we show that Cdc42-GTP polarization sites contain the M factor transporter Mam1, the general secretion machinery, which underlies P factor secretion, and Gpa1, suggesting that these are sub-cellular zones of pheromone secretion and signaling. Zone lifetimes scale with pheromone concentration. Computational simulations of pair formation through a fluctuating zone show that the combination of local pheromone release and sensing, short pheromone decay length, and pheromone-dependent zone stabilization leads to efficient pair formation. Consistently, pairing efficiency is reduced in the absence of the P factor protease. Similarly, zone stabilization at reduced pheromone levels, which occurs in the absence of the predicted GTPase-activating protein for Ras, leads to reduction in pairing efficiency. We propose that efficient cell pairing relies on fluctuating local signal emission and perception, which become locked into place through stimulation. PMID:27020743

  13. Metavanadate at the active site of the phosphatase VHZ.

    PubMed

    Kuznetsov, Vyacheslav I; Alexandrova, Anastassia N; Hengge, Alvan C

    2012-09-01

    Vanadate is a potent modulator of a number of biological processes and has been shown by crystal structures and NMR spectroscopy to interact with numerous enzymes. Although these effects often occur under conditions where oligomeric forms dominate, the crystal structures and NMR data suggest that the inhibitory form is usually monomeric orthovanadate, a particularly good inhibitor of phosphatases because of its ability to form stable trigonal-bipyramidal complexes. We performed a computational analysis of a 1.14 Å structure of the phosphatase VHZ in complex with an unusual metavanadate species and compared it with two classical trigonal-bipyramidal vanadate-phosphatase complexes. The results support extensive delocalized bonding to the apical ligands in the classical structures. In contrast, in the VHZ metavanadate complex, the central, planar VO(3)(-) moiety has only one apical ligand, the nucleophilic Cys95, and a gap in electron density between V and S. A computational analysis showed that the V-S interaction is primarily ionic. A mechanism is proposed to explain the formation of metavanadate in the active site from a dimeric vanadate species that previous crystallographic evidence has shown to be able to bind to the active sites of phosphatases related to VHZ. Together, the results show that the interaction of vanadate with biological systems is not solely reliant upon the prior formation of a particular inhibitory form in solution. The catalytic properties of an enzyme may act upon the oligomeric forms primarily present in solution to generate species such as the metavanadate ion observed in the VHZ structure. PMID:22876963

  14. Zymogen Activation and Subcellular Activity of Subtilisin Kexin Isozyme 1/Site 1 Protease*

    PubMed Central

    da Palma, Joel Ramos; Burri, Dominique Julien; Oppliger, Joël; Salamina, Marco; Cendron, Laura; de Laureto, Patrizia Polverino; Seidah, Nabil Georges; Kunz, Stefan; Pasquato, Antonella

    2014-01-01

    The proprotein convertase subtilisin kexin isozyme 1 (SKI-1)/site 1 protease (S1P) plays crucial roles in cellular homeostatic functions and is hijacked by pathogenic viruses for the processing of their envelope glycoproteins. Zymogen activation of SKI-1/S1P involves sequential autocatalytic processing of its N-terminal prodomain at sites B′/B followed by the herein newly identified C′/C sites. We found that SKI-1/S1P autoprocessing results in intermediates whose catalytic domain remains associated with prodomain fragments of different lengths. In contrast to other zymogen proprotein convertases, all incompletely matured intermediates of SKI-1/S1P showed full catalytic activity toward cellular substrates, whereas optimal cleavage of viral glycoproteins depended on B′/B processing. Incompletely matured forms of SKI-1/S1P further process cellular and viral substrates in distinct subcellular compartments. Using a cell-based sensor for SKI-1/S1P activity, we found that 9 amino acid residues at the cleavage site (P1–P8) and P1′ are necessary and sufficient to define the subcellular location of processing and to determine to what extent processing of a substrate depends on SKI-1/S1P maturation. In sum, our study reveals novel and unexpected features of SKI-1/S1P zymogen activation and subcellular specificity of activity toward cellular and pathogen-derived substrates. PMID:25378398

  15. Searching target sites on DNA by proteins: Role of DNA dynamics under confinement

    PubMed Central

    Mondal, Anupam; Bhattacherjee, Arnab

    2015-01-01

    DNA-binding proteins (DBPs) rapidly search and specifically bind to their target sites on genomic DNA in order to trigger many cellular regulatory processes. It has been suggested that the facilitation of search dynamics is achieved by combining 3D diffusion with one-dimensional sliding and hopping dynamics of interacting proteins. Although, recent studies have advanced the knowledge of molecular determinants that affect one-dimensional search efficiency, the role of DNA molecule is poorly understood. In this study, by using coarse-grained simulations, we propose that dynamics of DNA molecule and its degree of confinement due to cellular crowding concertedly regulate its groove geometry and modulate the inter-communication with DBPs. Under weak confinement, DNA dynamics promotes many short, rotation-decoupled sliding events interspersed by hopping dynamics. While this results in faster 1D diffusion, associated probability of missing targets by jumping over them increases. In contrast, strong confinement favours rotation-coupled sliding to locate targets but lacks structural flexibility to achieve desired specificity. By testing under physiological crowding, our study provides a plausible mechanism on how DNA molecule may help in maintaining an optimal balance between fast hopping and rotation-coupled sliding dynamics, to locate target sites rapidly and form specific complexes precisely. PMID:26400158

  16. Chemotactic Activity of Cyclophilin A in the Skin Mucus of Yellow Catfish (Pelteobagrus fulvidraco) and Its Active Site for Chemotaxis.

    PubMed

    Dawar, Farman Ullah; Tu, Jiagang; Xiong, Yang; Lan, Jiangfeng; Dong, Xing Xing; Liu, Xiaoling; Khattak, Muhammad Nasir Khan; Mei, Jie; Lin, Li

    2016-01-01

    Fish skin mucus is a dynamic barrier for invading pathogens with a variety of anti-microbial enzymes, including cyclophilin A (CypA), a multi-functional protein with peptidyl-prolyl cis/trans isomerase (PPIase) activity. Beside various other immunological functions, CypA induces leucocytes migration in vitro in teleost. In the current study, we have discovered several novel immune-relevant proteins in yellow catfish skin mucus by mass spectrometry (MS). The CypA present among them was further detected by Western blot. Moreover, the CypA present in the skin mucus displayed strong chemotactic activity for yellow catfish leucocytes. Interestingly, asparagine (like arginine in mammals) at position 69 was the critical site in yellow catfish CypA involved in leucocyte attraction. These novel efforts do not only highlight the enzymatic texture of skin mucus, but signify CypA to be targeted for anti-inflammatory therapeutics. PMID:27589721

  17. Chemotactic Activity of Cyclophilin A in the Skin Mucus of Yellow Catfish (Pelteobagrus fulvidraco) and Its Active Site for Chemotaxis

    PubMed Central

    Dawar, Farman Ullah; Tu, Jiagang; Xiong, Yang; Lan, Jiangfeng; Dong, Xing Xing; Liu, Xiaoling; Khattak, Muhammad Nasir Khan; Mei, Jie; Lin, Li

    2016-01-01

    Fish skin mucus is a dynamic barrier for invading pathogens with a variety of anti-microbial enzymes, including cyclophilin A (CypA), a multi-functional protein with peptidyl-prolyl cis/trans isomerase (PPIase) activity. Beside various other immunological functions, CypA induces leucocytes migration in vitro in teleost. In the current study, we have discovered several novel immune-relevant proteins in yellow catfish skin mucus by mass spectrometry (MS). The CypA present among them was further detected by Western blot. Moreover, the CypA present in the skin mucus displayed strong chemotactic activity for yellow catfish leucocytes. Interestingly, asparagine (like arginine in mammals) at position 69 was the critical site in yellow catfish CypA involved in leucocyte attraction. These novel efforts do not only highlight the enzymatic texture of skin mucus, but signify CypA to be targeted for anti-inflammatory therapeutics. PMID:27589721

  18. Non-specific binding sites help to explain mixed inhibition in mushroom tyrosinase activities.

    PubMed

    Hassani, Sorour; Haghbeen, Kamahldin; Fazli, Mostafa

    2016-10-21

    Inhibition and activation studies of tyrosinase could prove beneficial to agricultural, food, cosmetic, and pharmaceutical industries. Although non-competitive and mixed-inhibition are frequent modes observed in kinetics studies on mushroom tyrosinase (MT) activities, the phenomena are left unexplained. In this study, dual effects of phthalic acid (PA) and cinnamic acid (CA) on MT during mono-phenolase activity were demonstrated. PA activated and inhibited MT at concentrations lower and higher than 150 μM, respectively. In contrast, CA inhibited and activated MT at concentrations lower and higher than 5 μM. The mode of inhibition for both effectors was mixed-type. Complex kinetics of MT in the presence of a modulator could partly be ascribed to its mixed-cooperativity. However, to explain mixed-inhibition mode, it is necessary to demonstrate how the ternary complex of substrate/enzyme/effector is formed. Therefore, we looked for possible non-specific binding sites using MT tropolone-bound PDB (2Y9X) in the computational studies. When tropolone was in MTPa (active site), PA and CA occupied different pockets (named MTPb and MTPc, respectively). The close Moldock scores of PA binding posed in MTPb and MTPa suggested that MTPb could be a secondary binding site for PA. Similar results were obtained for CA. Ensuing results from 10 ns molecular dynamics simulations for 2Y9X-effector complexes indicated that the structures were gradually stabilized during simulation. Tunnel analysis by using CAVER Analyst and CHEXVIS resulted in identifying two distinct channels that assumingly participate in exchanging the effectors when the direct channel to MTPa is not accessible.

  19. Non-specific binding sites help to explain mixed inhibition in mushroom tyrosinase activities.

    PubMed

    Hassani, Sorour; Haghbeen, Kamahldin; Fazli, Mostafa

    2016-10-21

    Inhibition and activation studies of tyrosinase could prove beneficial to agricultural, food, cosmetic, and pharmaceutical industries. Although non-competitive and mixed-inhibition are frequent modes observed in kinetics studies on mushroom tyrosinase (MT) activities, the phenomena are left unexplained. In this study, dual effects of phthalic acid (PA) and cinnamic acid (CA) on MT during mono-phenolase activity were demonstrated. PA activated and inhibited MT at concentrations lower and higher than 150 μM, respectively. In contrast, CA inhibited and activated MT at concentrations lower and higher than 5 μM. The mode of inhibition for both effectors was mixed-type. Complex kinetics of MT in the presence of a modulator could partly be ascribed to its mixed-cooperativity. However, to explain mixed-inhibition mode, it is necessary to demonstrate how the ternary complex of substrate/enzyme/effector is formed. Therefore, we looked for possible non-specific binding sites using MT tropolone-bound PDB (2Y9X) in the computational studies. When tropolone was in MTPa (active site), PA and CA occupied different pockets (named MTPb and MTPc, respectively). The close Moldock scores of PA binding posed in MTPb and MTPa suggested that MTPb could be a secondary binding site for PA. Similar results were obtained for CA. Ensuing results from 10 ns molecular dynamics simulations for 2Y9X-effector complexes indicated that the structures were gradually stabilized during simulation. Tunnel analysis by using CAVER Analyst and CHEXVIS resulted in identifying two distinct channels that assumingly participate in exchanging the effectors when the direct channel to MTPa is not accessible. PMID:27344491

  20. Can Dynamics Be Responsible for the Complex Multipeak Infrared Spectra of NO Adsorbed to Copper(II) Sites in Zeolites?

    PubMed

    Göltl, Florian; Sautet, Philippe; Hermans, Ive

    2015-06-26

    Copper-exchanged SSZ-13 is a very efficient material in the selective catalytic reduction of NO(x) using ammonia (deNO(x)-SCR) and characterizing the underlying distribution of copper sites in the material is of prime importance to understand its activity. The IR spectrum of NO adsorbed to divalent copper sites are modeled using ab initio molecular dynamics simulations. For most sites, complex multi-peak spectra induced by the thermal motion of the cation as well as the adsorbate are found. A finite temperature spectrum for a specific catalyst was constructed, which shows excellent agreement with previously reported data. Additionally these findings allow active and inactive species in deNO(x)-SCR to be identified. To the best of our knowledge, this is the first time such complex spectra for single molecules adsorbed to single active centers have been reported in heterogeneous catalysis, and we expect similar effects to be important in a large number of systems with mobile active centers. PMID:25966680

  1. Site-specific PEGylation of lidamycin and its antitumor activity.

    PubMed

    Li, Liang; Shang, Boyang; Hu, Lei; Shao, Rongguang; Zhen, Yongsu

    2015-05-01

    In this study, N-terminal site-specific mono-PEGylation of the recombinant lidamycin apoprotein (rLDP) of lidamycin (LDM) was prepared using a polyethyleneglycol (PEG) derivative (M w 20 kDa) through a reactive terminal aldehyde group under weak acidic conditions (pH 5.5). The biochemical properties of mPEG-rLDP-AE, an enediyne-integrated conjugate, were analyzed by SDS-PAGE, RP-HPLC, SEC-HPLC and MALDI-TOF. Meanwhile, in vitro and in vivo antitumor activity of mPEG-rLDP-AE was evaluated by MTT assays and in xenograft model. The results indicated that mPEG-rLDP-AE showed significant antitumor activity both in vitro and in vivo. After PEGylation, mPEG-rLDP still retained the binding capability to the enediyne AE and presented the physicochemical characteristics similar to that of native LDP. It is of interest that the PEGylation did not diminish the antitumor efficacy of LDM, implying the possibility that this derivative may function as a payload to deliver novel tumor-targeted drugs. PMID:26579455

  2. Hybrid [FeFe]-hydrogenases with modified active sites show remarkable residual enzymatic activity.

    PubMed

    Siebel, Judith F; Adamska-Venkatesh, Agnieszka; Weber, Katharina; Rumpel, Sigrun; Reijerse, Edward; Lubitz, Wolfgang

    2015-02-24

    [FeFe]-hydrogenases are to date the only enzymes for which it has been demonstrated that the native inorganic binuclear cofactor of the active site Fe2(adt)(CO)3(CN)2 (adt = azadithiolate = [S-CH2-NH-CH2-S](2-)) can be synthesized on the laboratory bench and subsequently inserted into the unmaturated enzyme to yield fully functional holo-enzyme (Berggren, G. et al. (2013) Nature 499, 66-70; Esselborn, J. et al. (2013) Nat. Chem. Biol. 9, 607-610). In the current study, we exploit this procedure to introduce non-native cofactors into the enzyme. Mimics of the binuclear subcluster with a modified bridging dithiolate ligand (thiodithiolate, N-methylazadithiolate, dimethyl-azadithiolate) and three variants containing only one CN(-) ligand were inserted into the active site of the enzyme. We investigated the activity of these variants for hydrogen oxidation as well as proton reduction and their structural accommodation within the active site was analyzed using Fourier transform infrared spectroscopy. Interestingly, the monocyanide variant with the azadithiolate bridge showed ∼50% of the native enzyme activity. This would suggest that the CN(-) ligands are not essential for catalytic activity, but rather serve to anchor the binuclear subsite inside the protein pocket through hydrogen bonding. The inserted artificial cofactors with a propanedithiolate and an N-methylazadithiolate bridge as well as their monocyanide variants also showed residual activity. However, these activities were less than 1% of the native enzyme. Our findings indicate that even small changes in the dithiolate bridge of the binuclear subsite lead to a rather strong decrease of the catalytic activity. We conclude that both the Brønsted base function and the conformational flexibility of the native azadithiolate amine moiety are essential for the high catalytic activity of the native enzyme. PMID:25633077

  3. Competing dynamic phases of active polymer networks

    NASA Astrophysics Data System (ADS)

    Freedman, Simon; Banerjee, Shiladitya; Dinner, Aaron R.

    Recent experiments on in-vitro reconstituted assemblies of F-actin, myosin-II motors, and cross-linking proteins show that tuning local network properties can changes the fundamental biomechanical behavior of the system. For example, by varying cross-linker density and actin bundle rigidity, one can switch between contractile networks useful for reshaping cells, polarity sorted networks ideal for directed molecular transport, and frustrated networks with robust structural properties. To efficiently investigate the dynamic phases of actomyosin networks, we developed a coarse grained non-equilibrium molecular dynamics simulation of model semiflexible filaments, molecular motors, and cross-linkers with phenomenologically defined interactions. The simulation's accuracy was verified by benchmarking the mechanical properties of its individual components and collective behavior against experimental results at the molecular and network scales. By adjusting the model's parameters, we can reproduce the qualitative phases observed in experiment and predict the protein characteristics where phase crossovers could occur in collective network dynamics. Our model provides a framework for understanding cells' multiple uses of actomyosin networks and their applicability in materials research. Supported by the Department of Defense (DoD) through the National Defense Science & Engineering Graduate Fellowship (NDSEG) Program.

  4. Site-level model intercomparison of high latitude and high altitude soil thermal dynamics in tundra and barren landscapes

    NASA Astrophysics Data System (ADS)

    Ekici, A.; Chadburn, S.; Chaudhary, N.; Hajdu, L. H.; Marmy, A.; Peng, S.; Boike, J.; Burke, E.; Friend, A. D.; Hauck, C.; Krinner, G.; Langer, M.; Miller, P. A.; Beer, C.

    2014-09-01

    Modelling soil thermal dynamics at high latitudes and altitudes requires representations of specific physical processes such as snow insulation, soil freezing/thawing, as well as subsurface conditions like soil water/ice content and soil texture type. We have compared six different land models (JSBACH, ORCHIDEE, JULES, COUP, HYBRID8, LPJ-GUESS) at four different sites with distinct cold region landscape types (i.e. Schilthorn-Alpine, Bayelva-high Arctic, Samoylov-wet polygonal tundra, Nuuk-non permafrost Arctic) to quantify the importance of physical processes in capturing observed temperature dynamics in soils. This work shows how a range of models can represent distinct soil temperature regimes in permafrost and non-permafrost soils. Snow insulation is of major importance for estimating topsoil conditions and must be combined with accurate subsoil temperature dynamics to correctly estimate active layer thicknesses. Analyses show that land models need more realistic surface processes (such as detailed snow dynamics and moss cover with changing thickness/wetness) as well as better representations of subsoil thermal dynamics (i.e. soil heat transfer mechanism and correct parameterization of heat conductivity/capacities).

  5. Evaluation of Dynamic Changes in Rating of Russian Information Sources of Medical Education Sites.

    PubMed

    Vasilyeva, Irina V; Arseniev, Sergey B

    2016-01-01

    The aim of the present study is to analyze dynamic changes in the rating of information sources of medical literature in the sites of the following electronic libraries (, , ) and the rating of information sources for electronic medical books (, ). While using the on-line programs Alexa and Cy-pr, we have analyzed their website's rating and identified basic data and time-varying site data obtained for fourteen months. Alexa Rank rating was calculated for each sitemonthly. Our study has shown that the most popular information sources of medical education among the six studied sites for Russian users is ; the site is at the second place. PMID:27350475

  6. Molecular Dynamics Simulations of Solvation and Kink Site Formation at the {001} Barite-Water Interface.

    SciTech Connect

    Stack, Andrew G

    2009-09-01

    Solvation and kink site formation on step edges are known to be controlling parameters in crystal growth and dissolution. However, links from classical crystal growth models to specific reactions at the mineral-water interface have remained elusive. Molecular dynamics is used here to examine the water structure on barium surface sites and kink site formation enthalpies for material adsorbed to and removed from the step parallel to the [120] direction on the {001} barite-water interface. The bariums at the interface are shown to be coordinatively unsaturated with respect to water, and it is suggested that this is due to a steric hindrance from the nature of the interface. Kink site detachment energies that include hydration energies are endothermic for barium and exothermic for sulfate. The implications and problems of using these parameters in a crystal growth model are discussed.

  7. Key Role of Active-Site Water Molecules in Bacteriorhodopsin Proton-Transfer Reactions

    SciTech Connect

    Bondar, A.N.; Baudry, Jerome Y; Suhai, Sandor; Fischer, S.; Smith, Jeremy C

    2008-10-01

    The functional mechanism of the light-driven proton pump protein bacteriorhodopsin depends on the location of water molecules in the active site at various stages of the photocycle and on their roles in the proton-transfer steps. Here, free energy computations indicate that electrostatic interactions favor the presence of a cytoplasmic-side water molecule hydrogen bonding to the retinal Schiff base in the state preceding proton transfer from the retinal Schiff base to Asp85. However, the nonequilibrium nature of the pumping process means that the probability of occupancy of a water molecule in a given site depends both on the free energies of insertion of the water molecule in this and other sites during the preceding photocycle steps and on the kinetic accessibility of these sites on the time scale of the reaction steps. The presence of the cytoplasmic-side water molecule has a dramatic effect on the mechanism of proton transfer: the proton is channeled on the Thr89 side of the retinal, whereas the transfer on the Asp212 side is hindered. Reaction-path simulations and molecular dynamics simulations indicate that the presence of the cytoplasmic-side water molecule permits a low-energy bacteriorhodopsin conformer in which the water molecule bridges the twisted retinal Schiff base and the proton acceptor Asp85. From this low-energy conformer, proton transfer occurs via a concerted mechanism in which the water molecule participates as an intermediate proton carrier.

  8. Annual dynamics within the active layer

    SciTech Connect

    Not Available

    1991-01-01

    We have continued our meteorological and hydrologic data collection in support of our process-oriented research. The six years of data collected to date is unique in its scope and continuity in a North Hemisphere Arctic setting. This valuable data base has allowed us to further our understanding of the interconnections and interactions between the atmosphere/hydrosphere/biosphere/lithosphere. The increased understanding of the heat and mass transfer processes has allowed us to increase our model-oriented research efforts. Examples of applications are the following. (1) Spring snowmelt on the North Slope of Alaska is the dominant hydrologic event of the year. This event provides most of the moisture for use by vegetation in the spring and early summer period. The mechanisms and timing of snowmelt are important factors in predicting runoff, the migrations of birds and large mammals and the diversity of plant communities. It is important globally due to the radical and abrupt change in the surface energy balance over vast areas. (2) We were able to explore the trends and differences in the snowmelt process along a transect from the Brooks Range to the Arctic Coastal plain. Snowpack ablation was monitored at three sites. These data were analyzed along with meteorologic data at each site. The initiation of ablation was site specific being largely controlled by the complementary addition of energy from radiation and sensible heat flux.

  9. Summary report of Hanford Site well remediation and decommissioning activities for fiscal year 1994

    SciTech Connect

    Reynolds, K.D.

    1994-12-30

    Remediation and decommissioning of Hanford Site wells has become an integral part of Hanford Site Environmental Restoration (ER) and Resource Conservation and Recovery Act of 1976 (RCRA) groundwater monitoring programs. A well remediation and decommissioning program was funded and implemented in fiscal year (FY) 1993 under the RCRA and Operational Monitoring (ROM) Program. Funding for this work increased in FY 1994. In FY 1994 well decommissioning activities conducted for the ROM program were centered around the 200 West Area; activities for the ER program were centered in the Fitzner/Eberhart Arid Land Ecology (ALE) (Reserve) unit and the Wahluke Slope (North Slope) area. A total of 116 wells and test borings were decommissioned between the two programs during FY 1994. Additionally, five wells were identified as in need of remediation and were successfully brought into compliance with regulatory requirements. As Hanford Site restoration and remediation efforts increase in scope, the well decommissioning program will remain dynamic. The program will aggressively seek to fulfill the needs of the various environmental cleanup and groundwater/vadose monitoring programs. Wells that do not meet regulatory requirements for preservation will continually be identified and remediated or decommissioned accordingly.

  10. The Mechanism by which 146-N-Glycan Affects the Active Site of Neuraminidase.

    PubMed

    Liu, Pi; Wang, Zhonghua; Zhang, Lijie; Li, Dongmei; Lin, Jianping

    2015-01-01

    One of the most conserved glycosylation sites of neuraminidase (NA) is 146-N-glycan. This site is adjacent to the 150-cavity of NA, which is found within the active site and thought to be a target for rational drug development against the antiviral resistance of influenza. Here, through a total of 2.4 μs molecular dynamics (MD) simulations, we demonstrated that 146-N-glycan can stabilize the conformation of the 150-loop that controls the volume of the 150-cavity. Moreover, with 146-N-glycan, our simulation result was more consistent with crystal structures of NAs than simulations conducted without glycans. Cluster analysis of the MD trajectories showed that 146-N-glycan adopted three distinct conformations: monomer-bridged, dimer-bridged and standing. Of these conformations, the dimer-bridged 146-N-glycan was the most stable one and contributed to stabilization of the 150-loop conformation. Furthermore, our simulation revealed that various standing conformations of 146-N-glycan could block the entrance of the binding pocket. This result was consistent with experimental data and explained the relatively low activity of inhibitors with flexible substituents toward the 150-cavity. Together, our results lead us to hypothesize that rigid and hydrophobic substituents could serve as better inhibitors targeting the 150-cavity. PMID:26267136

  11. New binding site conformations of the dengue virus NS3 protease accessed by molecular dynamics simulation.

    PubMed

    de Almeida, Hugo; Bastos, Izabela M D; Ribeiro, Bergmann M; Maigret, Bernard; Santana, Jaime M

    2013-01-01

    Dengue fever is caused by four distinct serotypes of the dengue virus (DENV1-4), and is estimated to affect over 500 million people every year. Presently, there are no vaccines or antiviral treatments for this disease. Among the possible targets to fight dengue fever is the viral NS3 protease (NS3PRO), which is in part responsible for viral processing and replication. It is now widely recognized that virtual screening campaigns should consider the flexibility of target protein by using multiple active conformational states. The flexibility of the DENV NS3PRO could explain the relatively low success of previous virtual screening studies. In this first work, we explore the DENV NS3PRO conformational states obtained from molecular dynamics (MD) simulations to take into account protease flexibility during the virtual screening/docking process. To do so, we built a full NS3PRO model by multiple template homology modeling. The model comprised the NS2B cofactor (essential to the NS3PRO activation), a glycine flexible link and the proteolytic domain. MD simulations had the purpose to sample, as closely as possible, the ligand binding site conformational landscape prior to inhibitor binding. The obtained conformational MD sample was clustered into four families that, together with principal component analysis of the trajectory, demonstrated protein flexibility. These results allowed the description of multiple binding modes for the Bz-Nle-Lys-Arg-Arg-H inhibitor, as verified by binding plots and pair interaction analysis. This study allowed us to tackle protein flexibility in our virtual screening campaign against the dengue virus NS3 protease.

  12. Dynamic strength of the quadriceps muscle and sports activity.

    PubMed

    Hahn, T; Foldspang, A; Ingemann-Hansen, T

    1999-04-01

    The study objectives were to examine the dynamic strength of the quadriceps muscle in athletes, and investigate its association with participation in sport. The study comprised 168 active competitive non-pregnant athletes, aged 14-24 years. The dynamic strength of their quadriceps muscle was measured, and they answered a questionnaire about sports activity and occupation. The dynamic strength of the quadriceps muscle was significantly higher in men than in women, and was positively associated with body weight, years of jogging, years of soccer, and weekly hours of basketball. In conclusion, the dynamic strength of the quadriceps muscle seems to be associated with sports activity. The results suggest sport specific adaptation, which may reflect high levels of running and jumping activity.

  13. Topological structure dynamics revealing collective evolution in active nematics

    PubMed Central

    Shi, Xia-qing; Ma, Yu-qiang

    2013-01-01

    Topological defects frequently emerge in active matter like bacterial colonies, cytoskeleton extracts on substrates, self-propelled granular or colloidal layers and so on, but their dynamical properties and the relations to large-scale organization and fluctuations in these active systems are seldom touched. Here we reveal, through a simple model for active nematics using self-driven hard elliptic rods, that the excitation, annihilation and transportation of topological defects differ markedly from those in non-active media. These dynamical processes exhibit strong irreversibility in active nematics in the absence of detailed balance. Moreover, topological defects are the key factors in organizing large-scale dynamic structures and collective flows, resulting in multi-spatial temporal effects. These findings allow us to control the self-organization of active matter through topological structures. PMID:24346733

  14. Estimating groundwater dynamics at a Colorado floodplain site using historical hydrological data and climate information

    NASA Astrophysics Data System (ADS)

    Chen, J.; Hubbard, S. S.; Williams, K. H.

    2014-12-01

    Developing a predictive understanding of how hydrological variations impact biogeochemical cycles in terrestrial environments is challenging due to the wide range of processes occurring across vast spatiotemporal scales and due to the uncertainty associated with future climate conditions. In this study, we develop a multi-scale time series model to estimate groundwater dynamics at a floodplain site using historical hydrological data and climate information. Our study is focused at Rifle, CO, where the US DOE Sustainable Systems 2.0 project is developing a genome-to-watershed reactive transport simulation capability and where snowmelt annually delivers a hydrological pulse to the floodplain system that significantly influences the water table and subsurface cycles of carbon and nitrogen. Although long-term predictions of biogeochemical cycling at the site require estimates of hydrological conditions, hydrological data include only a few years of groundwater elevation measurements, with river gage data available from a station located approximately 26 miles upstream. To project future hydrological conditions at the site, we developed a multi-scale statistical model to combine both datasets. We first analyzed 47 years of hydrological data from the gage station to identify multi-frequency temporal patterns in the river stage and its relationship to climate factors (e.g., precipitation or temperature). We then developed empirical models to downscale the estimated hydrological information to daily discharge and subsequently transform them to groundwater dynamics at the downstream floodplain site. Our model provides a probabilistic estimation that is conditioned to the multi-scale hydrological and climate information. With the developed approach, we retrospectively estimate groundwater dynamics at the site for the past five decades as well as the associated uncertainty. Based on Colorado River Basin climate projections, we also predict mean and extreme hydrological

  15. Probing the active site loop motif of murine ferrochelatase by random mutagenesis.

    PubMed

    Shi, Zhen; Ferreira, Gloria C

    2004-05-01

    Ferrochelatase catalyzes the terminal step of the heme biosynthetic pathway by inserting ferrous iron into protoporphyrin IX. A conserved loop motif was shown to form part of the active site and contact the bound porphyrin by molecular dynamics calculations and structural analysis. We applied a random mutagenesis approach and steady-state kinetic analysis to assess the role of the loop motif in murine ferrochelatase function, particularly with respect to porphyrin interaction. Functional substitutions in the 10 consecutive loop positions Gln(248)-Leu(257) were identified by genetic complementation in Escherichia coli strain Deltavis. Lys(250), Val(251), Pro(253), Val(254), and Pro(255) tolerated a variety of replacements including single substitutions and contained low informational content. Gln(248), Ser(249), Gly(252), Trp(256), and Leu(257) possessed high informational content, since permissible replacements were limited and only observed in multiply substituted mutants. Selected active loop variants exhibited k(cat) values comparable with or higher than that of wild-type murine ferrochelatase. The K(m) values for porphyrin increased, except for the single mutant V251L. Other than a moderate increase observed in the triple mutant S249A/K250Q/V251C, the K(m) values for Fe(2+) were lowered. The k(cat)/K(m) for porphyrin remained largely unchanged, with the exception of a 10-fold reduction in the triple mutant K250M/V251L/W256Y. The k(cat)/K(m) for Fe(2+) was improved. Molecular modeling of these active loop variants indicated that loop mutations resulted in alterations of the active site architecture. However, despite the plasticity of the loop primary structure, the relative spatial positioning of the loop in the active site appeared to be maintained in functional variants, supporting a role for the loop in ferrochelatase function. PMID:14981080

  16. Active cage model of glassy dynamics.

    PubMed

    Fodor, Étienne; Hayakawa, Hisao; Visco, Paolo; van Wijland, Frédéric

    2016-07-01

    We build up a phenomenological picture in terms of the effective dynamics of a tracer confined in a cage experiencing random hops to capture some characteristics of glassy systems. This minimal description exhibits scale invariance properties for the small-displacement distribution that echo experimental observations. We predict the existence of exponential tails as a crossover between two Gaussian regimes. Moreover, we demonstrate that the onset of glassy behavior is controlled only by two dimensionless numbers: the number of hops occurring during the relaxation of the particle within a local cage and the ratio of the hopping length to the cage size. PMID:27575182

  17. Active cage model of glassy dynamics

    NASA Astrophysics Data System (ADS)

    Fodor, Étienne; Hayakawa, Hisao; Visco, Paolo; van Wijland, Frédéric

    2016-07-01

    We build up a phenomenological picture in terms of the effective dynamics of a tracer confined in a cage experiencing random hops to capture some characteristics of glassy systems. This minimal description exhibits scale invariance properties for the small-displacement distribution that echo experimental observations. We predict the existence of exponential tails as a crossover between two Gaussian regimes. Moreover, we demonstrate that the onset of glassy behavior is controlled only by two dimensionless numbers: the number of hops occurring during the relaxation of the particle within a local cage and the ratio of the hopping length to the cage size.

  18. Neural network with formed dynamics of activity

    SciTech Connect

    Dunin-Barkovskii, V.L.; Osovets, N.B.

    1995-03-01

    The problem of developing a neural network with a given pattern of the state sequence is considered. A neural network structure and an algorithm, of forming its bond matrix which lead to an approximate but robust solution of the problem are proposed and discussed. Limiting characteristics of the serviceability of the proposed structure are studied. Various methods of visualizing dynamic processes in a neural network are compared. Possible applications of the results obtained for interpretation of neurophysiological data and in neuroinformatics systems are discussed.

  19. Dynamic MRI of small electrical activity.

    PubMed

    Song, Allen W; Truong, Trong-Kha; Woldorff, Marty

    2009-01-01

    Neuroscience methods entailing in vivo measurements of brain activity have greatly contributed to our understanding of brain function for the past decades, from the invasive early studies in animals using single-cell electrical recordings, to the noninvasive techniques in humans of scalp-recorded electroencephalography (EEG) and magnetoencephalography (MEG), positron emission tomography (PET), and, most recently, blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI). A central objective of these techniques is to measure neuronal activities with high spatial and temporal resolution. Each of these methods, however, has substantial limitations in this regard. Single-cell recording is invasive and only typically records cellular activity in a single location; EEG/MEG cannot generally provide accurate and unambiguous delineations of neuronal activation spatially; and the most sophisticated BOLD-based fMRI methods are still fundamentally limited by their dependence on the very slow hemodynamic responses upon which they are based. Even the latest neuroimaging methodology (e.g., multimodal EEG/fMRI) does not yet unambiguously provide accurate localization of neuronal activation spatially and temporally. There is hence a need to further develop noninvasive imaging methods that can directly image neuroelectric activity and thus truly achieve a high temporal resolution and spatial specificity in humans. Here, we discuss the theory, implementation, and potential utility of an MRI technique termed Lorentz effect imaging (LEI) that can detect spatially incoherent yet temporally synchronized, minute electrical activities in the neural amplitude range (microamperes) when they occur in a strong magnetic field. Moreover, we demonstrate with our preliminary results in phantoms and in vivo, the feasibility of imaging such activities with a temporal resolution on the order of milliseconds.

  20. Binding site identification and role of permanent water molecule of PIM-3 kinase: A molecular dynamics study.

    PubMed

    Ul-Haq, Zaheer; Gul, Sana; Usmani, Saman; Wadood, Abdul; Khan, Waqasuddin

    2015-11-01

    The kinome is a protein kinase complement of the human genome, categorized as serine/threonine and tyrosine kinases. These kinases catalyze phosphorylation reaction by using ATP as phosphoryl donor. Proviral Integration Site for Moloney Murine Leukemia Virus (PIM) kinase encodes serine/threonine protein kinases that recognized as proto-oncogene, responsible for rapid growth of cancerous cells. It is implicated in cell survival and function via cell cycle progression and its metabolism. PIM-3, sub-member of PIM kinases is a proto-oncogene, its overexpression inhibits apoptosis, and results in progression of hepatocellular carcinoma. PIM-3 is considered as a promising drug target but attempts to develop its specific inhibitors is slowed down due to the lack of 3D structure by any experimental technique. In silico techniques generally facilitate scientist to explore hidden structural features in order to improve drug discovery. In the present study, homology modeling, molecular docking and MD simulation techniques were utilized to explore the structure and dynamics of PIM-3 kinase. Induction of water molecules during molecular docking simulation explored differences in the hinge region between PIM-1 and PIM-3 kinases that may be responsible for specificity. Furthermore, role of water molecules in the active site was also explored via radial distribution function (RDF) after a 10 ns molecular dynamics (MD) simulations. Generated RDF plots exhibited the importance of water for inhibitor binding through their bridging capability that links the ligand with binding site residues.

  1. Binding site identification and role of permanent water molecule of PIM-3 kinase: A molecular dynamics study.

    PubMed

    Ul-Haq, Zaheer; Gul, Sana; Usmani, Saman; Wadood, Abdul; Khan, Waqasuddin

    2015-11-01

    The kinome is a protein kinase complement of the human genome, categorized as serine/threonine and tyrosine kinases. These kinases catalyze phosphorylation reaction by using ATP as phosphoryl donor. Proviral Integration Site for Moloney Murine Leukemia Virus (PIM) kinase encodes serine/threonine protein kinases that recognized as proto-oncogene, responsible for rapid growth of cancerous cells. It is implicated in cell survival and function via cell cycle progression and its metabolism. PIM-3, sub-member of PIM kinases is a proto-oncogene, its overexpression inhibits apoptosis, and results in progression of hepatocellular carcinoma. PIM-3 is considered as a promising drug target but attempts to develop its specific inhibitors is slowed down due to the lack of 3D structure by any experimental technique. In silico techniques generally facilitate scientist to explore hidden structural features in order to improve drug discovery. In the present study, homology modeling, molecular docking and MD simulation techniques were utilized to explore the structure and dynamics of PIM-3 kinase. Induction of water molecules during molecular docking simulation explored differences in the hinge region between PIM-1 and PIM-3 kinases that may be responsible for specificity. Furthermore, role of water molecules in the active site was also explored via radial distribution function (RDF) after a 10 ns molecular dynamics (MD) simulations. Generated RDF plots exhibited the importance of water for inhibitor binding through their bridging capability that links the ligand with binding site residues. PMID:26529487

  2. Characterization of the active site of chloroperoxidase using physical techniques

    SciTech Connect

    Hall, K.S.

    1986-01-01

    Chloroperoxidase (CPO) and Cytochrome P-450, two very different hemeproteins, have been shown to have similar active sites by several techniques. Recent work has demonstrated thiolate ligation from a cysteine residue to the iron in P-450. A major portion of this research has been devoted to obtaining direct evidence that CPO also has a thiolate 5th ligand from a cysteine residue. This information will provide the framework for a detailed analysis of the structure-function relationships between peroxidases, catalase and cytochrome P-450 hemeproteins. To determine whether the 5th ligand is a cysteine, methionine or a unique amino acid, specific isotope enrichment experiments were used. Preliminary /sup 1/H-NMR studies show that the carbon monoxide-CPO complex has a peak in the upfield region corresponding to alpha-protons of a thiolate amino acid. C. fumago was grown on 95% D/sub 2/O media with a small amount of /sup 1/H-cysteine added. Under these conditions C. fumago slows down the biosynthesis of cysteine by at least 50% and utilizes the exogenous cysteine in the media. GC-MS was able to show that the methylene protons next to the sulfur atom in cysteine are 80-90% protonated while these positions in methionine are approximately 73% deuterated. Comparison of the /sup 1/H-NMR spectra of CO-CPO and CO-CPO indicate the presence of a cysteine ligand in chloroperoxidase.

  3. N6-Methyldeoxyadenosine Marks Active Transcription Start Sites in Chlamydomonas

    PubMed Central

    Chen, Kai; Deng, Xin; Yu, Miao; Han, Dali; Hao, Ziyang; Liu, Jianzhao; Lu, Xingyu; Dore, Louis C; Weng, Xiaocheng; Ji, Quanjiang; Mets, Laurens; He, Chuan

    2015-01-01

    SUMMARY N6-methyldeoxyadenosine (6mA or m6A) is a DNA modification preserved in prokaryotes to eukaryotes. It is widespread in bacteria, and functions in DNA mismatch repair, chromosome segregation, and virulence regulation. In contrast, the distribution and function of 6mA in eukaryotes have been unclear. Here we present a comprehensive analysis of the 6mA landscape in the genome of Chlamydomonas using new sequencing approaches. We identified the 6mA modification in 84% of genes in Chlamydomonas. We found that 6mA mainly locates at ApT dinucleotides around transcription start sites (TSS) with a bimodal distribution, and appears to mark active genes. A periodic pattern of 6mA deposition was also observed at base resolution, which is associated with nucleosome distribution near the TSS, suggesting a possible role in nucleosome positioning. The new genome-wide mapping of 6mA and its unique distribution in the Chlamydomonas genome suggest potential regulatory roles of 6mA in gene expression in eukaryotic organisms. PMID:25936837

  4. Detection limit for activation measurements in ultralow background sites

    NASA Astrophysics Data System (ADS)

    Trache, Livius; Chesneanu, D.; Margineanu, R.; Pantelica, A.; Ghita, D. G.; Burducea, I.; Straticiuc, M.; Tang, X. D.

    2014-09-01

    We used 12C +13C fusion at the beam energies E = 6, 7 and 8 MeV to determine the sensitivity and the limits of activation method measurements in ultralow background sites. A 13C beam of 0.5 μA from the 3 MV Tandem accelerator of the Horia Hulubei National Institute of Physics and Nuclear Engineering - IFIN HH impinged on thick graphite targets. After about 24 hrs of irradiation targets were measured in two different laboratories: one with a heavy shielded Ge detector in the institute (at the surface) and one located underground in the microBequerel laboratory, in the salt mine of Slanic-Prahova, Romania. The 1369- and 2754 keV peaks from 24Na deactivation were clearly observed in the γ-ray spectra obtained for acquisitions lasting a few hours, or a few days. Determination of the detection limit in evaluating the cross sections for the target irradiated at Ec . m = 3 MeV indicates the fact that it is possible to measure gamma spectrum in underground laboratory down to Ec . m = 2 . 6 MeV. Cleaning the spectra with beta-gamma coincidences and increasing beam intensity 20 times will take as further down. The measurements are motivated by the study of the 12 C +12 C reaction at astrophysical energies.

  5. Disturbance opens recruitment sites for bacterial colonization in activated sludge.

    PubMed

    Vuono, David C; Munakata-Marr, Junko; Spear, John R; Drewes, Jörg E

    2016-01-01

    Little is known about the role of immigration in shaping bacterial communities or the factors that may dictate success or failure of colonization by bacteria from regional species pools. To address these knowledge gaps, the influence of bacterial colonization into an ecosystem (activated sludge bioreactor) was measured through a disturbance gradient (successive decreases in the parameter solids retention time) relative to stable operational conditions. Through a DNA sequencing approach, we show that the most abundant bacteria within the immigrant community have a greater probability of colonizing the receiving ecosystem, but mostly as low abundance community members. Only during the disturbance do some of these bacterial populations significantly increase in abundance beyond background levels and in few cases become dominant community members post-disturbance. Two mechanisms facilitate the enhanced enrichment of immigrant populations during disturbance: (i) the availability of resources left unconsumed by established species and (ii) the increased availability of niche space for colonizers to establish and displace resident populations. Thus, as a disturbance decreases local diversity, recruitment sites become available to promote colonization. This work advances our understanding of microbial resource management and diversity maintenance in complex ecosystems. PMID:25727891

  6. Active Site Characterization of Proteases Sequences from Different Species of Aspergillus.

    PubMed

    Morya, V K; Yadav, Virendra K; Yadav, Sangeeta; Yadav, Dinesh

    2016-09-01

    A total of 129 proteases sequences comprising 43 serine proteases, 36 aspartic proteases, 24 cysteine protease, 21 metalloproteases, and 05 neutral proteases from different Aspergillus species were analyzed for the catalytically active site residues using MEROPS database and various bioinformatics tools. Different proteases have predominance of variable active site residues. In case of 24 cysteine proteases of Aspergilli, the predominant active site residues observed were Gln193, Cys199, His364, Asn384 while for 43 serine proteases, the active site residues namely Asp164, His193, Asn284, Ser349 and Asp325, His357, Asn454, Ser519 were frequently observed. The analysis of 21 metalloproteases of Aspergilli revealed Glu298 and Glu388, Tyr476 as predominant active site residues. In general, Aspergilli species-specific active site residues were observed for different types of protease sequences analyzed. The phylogenetic analysis of these 129 proteases sequences revealed 14 different clans representing different types of proteases with diverse active site residues.

  7. A proposed definition of the 'activity' of surface sites on lactose carriers for dry powder inhalation.

    PubMed

    Grasmeijer, Floris; Frijlink, Henderik W; de Boer, Anne H

    2014-06-01

    A new definition of the activity of surface sites on lactose carriers for dry powder inhalation is proposed which relates to drug detachment during dispersion. The new definition is expected to improve the understanding of 'carrier surface site activity', which stimulates the unambiguous communication about this subject and may aid in the rational design and interpretation of future formulation studies. In contrast to the currently prevailing view on carrier surface site activity, it follows from the newly proposed definition that carrier surface site activity depends on more variables than just the physicochemical properties of the carrier surface. Because the term 'active sites' is ambiguous, it is recommended to use the term 'highly active sites' instead to denote carrier surface sites with a relatively high activity. PMID:24613490

  8. Tunable dynamics of microtubule-based active isotropic gels

    PubMed Central

    Henkin, Gil; DeCamp, Stephen J.; Chen, Daniel T. N.; Sanchez, Tim; Dogic, Zvonimir

    2014-01-01

    We investigate the dynamics of an active gel of bundled microtubules (MTs) that is driven by clusters of kinesin molecular motors. Upon the addition of ATP, the coordinated action of thousands of molecular motors drives the gel to a highly dynamical turbulent-like state that persists for hours and is only limited by the stability of constituent proteins and the availability of the chemical fuel. We characterize how enhanced transport and emergent macroscopic flows of active gels depend on relevant molecular parameters, including ATP, kinesin motor and depletant concentrations, MT volume fraction, as well as the stoichiometry of the constituent motor clusters. Our results show that the dynamical and structural properties of MT-based active gels are highly tunable. They also indicate existence of an optimal concentration of molecular motors that maximize far-from-equilibrium activity of active isotropic MT gels. PMID:25332391

  9. Defect Dynamics in Active 2D Nematic Liquid Crystals

    NASA Astrophysics Data System (ADS)

    Decamp, Stephen; Redner, Gabriel; Hagan, Michael; Dogic, Zvonimir

    2014-03-01

    Active materials are assemblies of animate, energy-consuming objects that exhibit continuous dynamics. As such, they have properties that are dramatically different from those found in conventional materials made of inanimate objects. We present a 2D active nematic liquid crystal composed of bundled microtubules and kinesin motor proteins that exists in a dynamic steady-state far from equilibrium. The active nematic exhibits spontaneous binding and unbinding of charge +1/2 and -1/2 disclination defects as well as streaming of +1/2 defects. By tuning ATP concentration, we precisely control the amount of activity, a key parameter of the system. We characterize the dynamics of streaming defects on a large, flat, 2D interface using quantitative polarization light microscopy. We report fundamental characteristics of the active nematics such as defect velocities, defect creation and annihilation rates, and emergent length scales in the system.

  10. Site-specific dissociation dynamics of H{sub 2}/D{sub 2} on Ag(111) and Co(0001) and the validity of the site-averaging model

    SciTech Connect

    Hu, Xixi; Jiang, Bin; Xie, Daiqian E-mail: hguo@unm.edu; Guo, Hua E-mail: hguo@unm.edu

    2015-09-21

    Dissociative chemisorption of polyatomic molecules on metal surfaces involves high-dimensional dynamics, of which quantum mechanical treatments are computationally challenging. A promising reduced-dimensional approach approximates the full-dimensional dynamics by a weighted average of fixed-site results. To examine the performance of this site-averaging model, we investigate two distinct reactions, namely, hydrogen dissociation on Co(0001) and Ag(111), using accurate first principles potential energy surfaces (PESs). The former has a very low barrier of ∼0.05 eV while the latter is highly activated with a barrier of ∼1.15 eV. These two systems allow the investigation of not only site-specific dynamical behaviors but also the validity of the site-averaging model. It is found that the reactivity is not only controlled by the barrier height but also by the topography of the PES. Moreover, the agreement between the site-averaged and full-dimensional results is much better on Ag(111), though quantitative in neither system. Further quasi-classical trajectory calculations showed that the deviations can be attributed to dynamical steering effects, which are present in both reactions at all energies.

  11. A subharmonic dynamical bifurcation during in vitro epileptiform activity

    NASA Astrophysics Data System (ADS)

    Perez Velazquez, Jose L.; Khosravani, Houman

    2004-06-01

    Epileptic seizures are considered to result from a sudden change in the synchronization of firing neurons in brain neural networks. We have used an in vitro model of status epilepticus (SE) to characterize dynamical regimes underlying the observed seizure-like activity. Time intervals between spikes or bursts were used as the variable to construct first-return interpeak or interburst interval plots, for studying neuronal population activity during the transition to seizure, as well as within seizures. Return maps constructed for a brief epoch before seizures were used for approximating the local system dynamics during that time window. Analysis of the first-return maps suggests that intermittency is a dynamical regime underlying the observed epileptic activity. This type of analysis may be useful for understanding the collective dynamics of neuronal populations in the normal and pathological brain.

  12. Seismic characterization and dynamic site response of a municipal solid waste landfill in Bangalore, India.

    PubMed

    Anbazhagan, P; SivakumarBabu, G L; Lakshmikanthan, P; VivekAnand, K S

    2016-03-01

    Seismic design of landfills requires an understanding of the dynamic properties of municipal solid waste (MSW) and the dynamic site response of landfill waste during seismic events. The dynamic response of the Mavallipura landfill situated in Bangalore, India, is investigated using field measurements, laboratory studies and recorded ground motions from the intraplate region. The dynamic shear modulus values for the MSW were established on the basis of field measurements of shear wave velocities. Cyclic triaxial testing was performed on reconstituted MSW samples and the shear modulus reduction and damping characteristics of MSW were studied. Ten ground motions were selected based on regional seismicity and site response parameters have been obtained considering one-dimensional non-linear analysis in the DEEPSOIL program. The surface spectral response varied from 0.6 to 2 g and persisted only for a period of 1 s for most of the ground motions. The maximum peak ground acceleration (PGA) obtained was 0.5 g and the minimum and maximum amplifications are 1.35 and 4.05. Amplification of the base acceleration was observed at the top surface of the landfill underlined by a composite soil layer and bedrock for all ground motions. Dynamic seismic properties with amplification and site response parameters for MSW landfill in Bangalore, India, are presented in this paper. This study shows that MSW has less shear stiffness and more amplification due to loose filling and damping, which need to be accounted for seismic design of MSW landfills in India. PMID:26759434

  13. Seismic characterization and dynamic site response of a municipal solid waste landfill in Bangalore, India.

    PubMed

    Anbazhagan, P; SivakumarBabu, G L; Lakshmikanthan, P; VivekAnand, K S

    2016-03-01

    Seismic design of landfills requires an understanding of the dynamic properties of municipal solid waste (MSW) and the dynamic site response of landfill waste during seismic events. The dynamic response of the Mavallipura landfill situated in Bangalore, India, is investigated using field measurements, laboratory studies and recorded ground motions from the intraplate region. The dynamic shear modulus values for the MSW were established on the basis of field measurements of shear wave velocities. Cyclic triaxial testing was performed on reconstituted MSW samples and the shear modulus reduction and damping characteristics of MSW were studied. Ten ground motions were selected based on regional seismicity and site response parameters have been obtained considering one-dimensional non-linear analysis in the DEEPSOIL program. The surface spectral response varied from 0.6 to 2 g and persisted only for a period of 1 s for most of the ground motions. The maximum peak ground acceleration (PGA) obtained was 0.5 g and the minimum and maximum amplifications are 1.35 and 4.05. Amplification of the base acceleration was observed at the top surface of the landfill underlined by a composite soil layer and bedrock for all ground motions. Dynamic seismic properties with amplification and site response parameters for MSW landfill in Bangalore, India, are presented in this paper. This study shows that MSW has less shear stiffness and more amplification due to loose filling and damping, which need to be accounted for seismic design of MSW landfills in India.

  14. 10 CFR 63.16 - Review of site characterization activities. 2

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of... conduct of site characterization activities at the Yucca Mountain site, DOE shall report the nature and... activities at the Yucca Mountain site, NRC staff shall be permitted to visit and inspect the locations...

  15. 10 CFR 63.16 - Review of site characterization activities. 2

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of... conduct of site characterization activities at the Yucca Mountain site, DOE shall report the nature and... activities at the Yucca Mountain site, NRC staff shall be permitted to visit and inspect the locations...

  16. 10 CFR 63.16 - Review of site characterization activities. 2

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of... conduct of site characterization activities at the Yucca Mountain site, DOE shall report the nature and... activities at the Yucca Mountain site, NRC staff shall be permitted to visit and inspect the locations...

  17. 10 CFR 63.16 - Review of site characterization activities. 2

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of... conduct of site characterization activities at the Yucca Mountain site, DOE shall report the nature and... activities at the Yucca Mountain site, NRC staff shall be permitted to visit and inspect the locations...

  18. 10 CFR 63.16 - Review of site characterization activities. 2

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of... conduct of site characterization activities at the Yucca Mountain site, DOE shall report the nature and... activities at the Yucca Mountain site, NRC staff shall be permitted to visit and inspect the locations...

  19. Pervasive and dynamic protein binding sites of the mRNA transcriptome in Saccharomyces cerevisiae

    PubMed Central

    2013-01-01

    Background Protein-RNA interactions are integral components of nearly every aspect of biology, including regulation of gene expression, assembly of cellular architectures, and pathogenesis of human diseases. However, studies in the past few decades have only uncovered a small fraction of the vast landscape of the protein-RNA interactome in any organism, and even less is known about the dynamics of protein-RNA interactions under changing developmental and environmental conditions. Results Here, we describe the gPAR-CLIP (global photoactivatable-ribonucleoside-enhanced crosslinking and immunopurification) approach for capturing regions of the untranslated, polyadenylated transcriptome bound by RNA-binding proteins (RBPs) in budding yeast. We report over 13,000 RBP crosslinking sites in untranslated regions (UTRs) covering 72% of protein-coding transcripts encoded in the genome, confirming 3' UTRs as major sites for RBP interaction. Comparative genomic analyses reveal that RBP crosslinking sites are highly conserved, and RNA folding predictions indicate that secondary structural elements are constrained by protein binding and may serve as generalizable modes of RNA recognition. Finally, 38% of 3' UTR crosslinking sites show changes in RBP occupancy upon glucose or nitrogen deprivation, with major impacts on metabolic pathways as well as mitochondrial and ribosomal gene expression. Conclusions Our study offers an unprecedented view of the pervasiveness and dynamics of protein-RNA interactions in vivo. PMID:23409723

  20. Static and dynamic stress analyses of the prototype high head Francis runner based on site measurement

    NASA Astrophysics Data System (ADS)

    Huang, X.; Oram, C.; Sick, M.

    2014-03-01

    More efforts are put on hydro-power to balance voltage and frequency within seconds for primary control in modern smart grids. This requires hydraulic turbines to run at off-design conditions. especially at low load or speed-no load. Besides. the tendency of increasing power output and decreasing weight of the turbine runners has also led to the high level vibration problem of the runners. especially high head Francis runners. Therefore. it is important to carry out the static and dynamic stress analyses of prototype high head Francis runners. This paper investigates the static and dynamic stresses on the prototype high head Francis runner based on site measurements and numerical simulations. The site measurements are performed with pressure transducers and strain gauges. Based on the measured results. computational fluid dynamics (CFD) simulations for the flow channel from stay vane to draft tube cone are performed. Static pressure distributions and dynamic pressure pulsations caused by rotor-stator interaction (RSI) are obtained under various operating conditions. With the CFD results. static and dynamic stresses on the runner at different operating points are calculated by means of the finite element method (FEM). The agreement between simulation and measurement is analysed with linear regression method. which indicates that the numerical result agrees well with that of measurement. Furthermore. the maximum static and dynamic stresses on the runner blade are obtained at various operating points. The relations of the maximum stresses and the power output are discussed in detail. The influences of the boundary conditions on the structural behaviour of the runner are also discussed.

  1. Exchange and redistribution dynamics of the cytoskeleton of the active zone molecule bassoon.

    PubMed

    Tsuriel, Shlomo; Fisher, Arava; Wittenmayer, Nina; Dresbach, Thomas; Garner, Craig C; Ziv, Noam E

    2009-01-14

    Presynaptic sites typically appear as varicosities (boutons) distributed along axons. Ultrastructurally, presynaptic boutons lack obvious physical barriers that separate them from the axon proper, yet activity-related and constitutive dynamics continuously promote the "reshuffling" of presynaptic components and even their dispersal into flanking axonal segments. How presynaptic sites manage to maintain their organization and individual characteristics over long durations is thus unclear. Conceivably, presynaptic tenacity might depend on the active zone (AZ), an electron-dense specialization of the presynaptic membrane, and particularly on the cytoskeletal matrix associated with the AZ (CAZ) that could act as a relatively stable "core scaffold" that conserves and dictates presynaptic organization. At present, however, little is known on the molecular dynamics of CAZ molecules, and thus, the factual basis for this hypothesis remains unclear. To examine the stability of the CAZ, we studied the molecular dynamics of the major CAZ molecule Bassoon in cultured hippocampal neurons. Fluorescence recovery after photobleaching and photoactivation experiments revealed that exchange rates of green fluorescent protein and photoactivatable green fluorescent protein-tagged Bassoon at individual presynaptic sites are very low (tau > 8 h). Exchange rates varied between boutons and were only slightly accelerated by stimulation. Interestingly, photoactivation experiments revealed that Bassoon lost from one synapse was occasionally assimilated into neighboring presynaptic sites. Our findings indicate that Bassoon is engaged in relatively stable associations within the CAZ and thus support the notion that the CAZ or some of its components might constitute a relatively stable presynaptic core scaffold.

  2. Computational Prediction of Metabolism: Sites, Products, SAR, P450 Enzyme Dynamics, and Mechanisms

    PubMed Central

    2012-01-01

    Metabolism of xenobiotics remains a central challenge for the discovery and development of drugs, cosmetics, nutritional supplements, and agrochemicals. Metabolic transformations are frequently related to the incidence of toxic effects that may result from the emergence of reactive species, the systemic accumulation of metabolites, or by induction of metabolic pathways. Experimental investigation of the metabolism of small organic molecules is particularly resource demanding; hence, computational methods are of considerable interest to complement experimental approaches. This review provides a broad overview of structure- and ligand-based computational methods for the prediction of xenobiotic metabolism. Current computational approaches to address xenobiotic metabolism are discussed from three major perspectives: (i) prediction of sites of metabolism (SOMs), (ii) elucidation of potential metabolites and their chemical structures, and (iii) prediction of direct and indirect effects of xenobiotics on metabolizing enzymes, where the focus is on the cytochrome P450 (CYP) superfamily of enzymes, the cardinal xenobiotics metabolizing enzymes. For each of these domains, a variety of approaches and their applications are systematically reviewed, including expert systems, data mining approaches, quantitative structure–activity relationships (QSARs), and machine learning-based methods, pharmacophore-based algorithms, shape-focused techniques, molecular interaction fields (MIFs), reactivity-focused techniques, protein–ligand docking, molecular dynamics (MD) simulations, and combinations of methods. Predictive metabolism is a developing area, and there is still enormous potential for improvement. However, it is clear that the combination of rapidly increasing amounts of available ligand- and structure-related experimental data (in particular, quantitative data) with novel and diverse simulation and modeling approaches is accelerating the development of effective tools for

  3. GAS HYDRATES AT TWO SITES OF AN ACTIVE CONTINENTAL MARGIN.

    USGS Publications Warehouse

    Kvenvolden, K.A.

    1985-01-01

    Sediment containing gas hydrates from two distant Deep Sea Drilling Project sites (565 and 568), located about 670 km apart on the landward flank of the Middle America Trench, was studied to determine the geochemical conditions that characterize the occurrence of gas hydrates. Site 565 was located in the Pacific Ocean offshore the Nicoya Peninsula of Costa Rica in 3,111 m of water. The depth of the hole at this site was 328 m, and gas hydrates were recovered from 285 and 319 m. Site 568 was located about 670 km to the northwest offshore Guatemala in 2,031 m of water. At this site the hole penetrated to 418 m, and gas hydrates were encountered at 404 m.

  4. Stochastic dynamics of coupled active particles in an overdamped limit

    NASA Astrophysics Data System (ADS)

    Ann, Minjung; Lee, Kong-Ju-Bock; Park, Pyeong Jun

    2015-10-01

    We introduce a model for Brownian dynamics of coupled active particles in an overdamped limit. Our system consists of several identical active particles and one passive particle. Each active particle is elastically coupled to the passive particle and there is no direct coupling among the active particles. We investigate the dynamics of the system with respect to the number of active particles, viscous friction, and coupling between the active and passive particles. For this purpose, we consider an intracellular transport process as an application of our model and perform a Brownian dynamics simulation using realistic parameters for processive molecular motors such as kinesin-1. We determine an adequate energy conversion function for molecular motors and study the dynamics of intracellular transport by multiple motors. The results show that the average velocity of the coupled system is not affected by the number of active motors and that the stall force increases linearly as the number of motors increases. Our results are consistent with well-known experimental observations. We also examine the effects of coupling between the motors and the cargo, as well as of the spatial distribution of the motors around the cargo. Our model might provide a physical explanation of the cooperation among active motors in the cellular transport processes.

  5. Control of active sites in selective flocculation: III -- Mechanism of site blocking

    SciTech Connect

    Behl, S.; Moudgil, B.M. . Dept. of Materials Science and Engineering)

    1993-12-01

    It has been shown in Parts I and II of this paper that heteroflocculation can be controlled by poisoning the sites for flocculant adsorption using a site blocking agent (SBA). An efficient SBA was determined to be the lower molecular weight fraction of the flocculant. In this paper, the underlying mechanism of SBA action is described. Also, the mathematical model detailed in Part I is used to determine the effect of different SBAs on apatite-dolomite separation efficiency. It has been demonstrated that the depression in flocculation is directly related to the site blocking parameter ([bar [Phi

  6. Variation in body mass dynamics among sites in Black Brant Branta bernicla nigricans supports adaptivity of mass loss during moult

    USGS Publications Warehouse

    Fondell, Thomas F.; Flint, Paul L.; Schmutz, Joel A.; Schamber, Jason L.; Nicolai, Christopher A.

    2013-01-01

    Birds employ varying strategies to accommodate the energetic demands of moult, one important example being changes in body mass. To understand better their physiological and ecological significance, we tested three hypotheses concerning body mass dynamics during moult. We studied Black Brant in 2006 and 2007 moulting at three sites in Alaska which varied in food availability, breeding status and whether geese undertook a moult migration. First we predicted that if mass loss during moult were simply the result of inadequate food resources then mass loss would be highest where food was least available. Secondly, we predicted that if mass loss during moult were adaptive, allowing birds to reduce activity during moult, then birds would gain mass prior to moult where feeding conditions allowed and mass loss would be positively related to mass at moult initiation. Thirdly, we predicted that if mass loss during moult were adaptive, allowing birds to regain flight sooner, then across sites and groups, mass at the end of the flightless period would converge on a theoretical optimum, i.e. the mass that permits the earliest possible return to flight. Mass loss was greatest where food was most available and thus our results did not support the prediction that mass loss resulted from inadequate food availability. Mass at moult initiation was positively related to both food availability and mass loss. In addition, among sites and years, variation in mass was high at moult initiation but greatly reduced at the end of the flightless period, appearing to converge. Thus, our results supported multiple predictions that mass loss during moult was adaptive and that the optimal moulting strategy was to gain mass prior to the flightless period, then through behavioural modifications use these body reserves to reduce activity and in so doing also reduce wing loading. Geese that undertook a moult migration initiated moult at the highest mass, indicating that they were more than able to

  7. Dynamical downscaling of present climate extremal episodes for the BINGO research site of Cyprus

    NASA Astrophysics Data System (ADS)

    Zittis, George; Hadjinicolaou, Panos; Bruggeman, Adriana; Camera, Corrado; Lelieveld, Jos

    2016-04-01

    Besides global warming, climate change is expected to cause alterations in precipitation amounts and distribution than can be linked to extreme events such as floods or prolonged droughts. This will have a significant impact in strategic societal sectors that base their activities on water resources. While the global climate projections inform us about the long-term and weather forecasts can give useful information only for a few days or weeks, decision-makers and end-users also need guidance on inter-annual to decadal time scales. In this context, the BINGO (Bringing INnovation to onGOing water management - a better future under climate change) H2020 project aims both at reducing the uncertainty of near-term climate predictions and developing response strategies in order to better manage the remaining uncertainty. One of the project's main objectives is to develop improved decadal predictions, in adequate spatiotemporal scales, with a specific focus on extreme precipitation events. The projected rainfall will be eventually used to drive hydrological impact models. BINGO focuses on research sites that encompass river basins, watersheds and urban areas of six European countries including Norway, Cyprus, Germany, Portugal, The Netherlands and Spain. In this study we present the dynamical downscaling of the ERA-Interim dataset for validation purposes and for the research site of Cyprus. Five extreme rainfall periods were identified from the observed precipitation archives and were simulated in very high horizontal resolutions (4~1 km) using the WRF limited area atmospheric model. To optimize the performance of the model we have tested a combination of three cumulus and five microphysics parameterization schemes that resulted in 15 simulations for each extreme precipitation event. The model output was compared with daily or hourly (where available) representative rain gauge data. A set of statistical metrics was applied in order to objectively select the best

  8. Lethal Factor Active-Site Mutations Affect Catalytic Activity In Vitro

    PubMed Central

    Hammond, S. E.; Hanna, P. C.

    1998-01-01

    The lethal factor (LF) protein of Bacillus anthracis lethal toxin contains the thermolysin-like active-site and zinc-binding consensus motif HEXXH (K. R. Klimpel, N. Arora, and S. H. Leppla, Mol. Microbiol. 13:1093–1100, 1994). LF is hypothesized to act as a Zn2+ metalloprotease in the cytoplasm of macrophages, but no proteolytic activities have been previously shown on any target substrate. Here, synthetic peptides are hydrolyzed by LF in vitro. Mass spectroscopy and peptide sequencing of isolated cleavage products separated by reverse-phase high-pressure liquid chromatography indicate that LF seems to prefer proline-containing substrates. Substitution mutations within the consensus active-site residues completely abolish all in vitro catalytic functions, as does addition of 1,10-phenanthroline, EDTA, and certain amino acid hydroxamates, including the novel zinc metalloprotease inhibitor ZINCOV. In contrast, the protease inhibitors bestatin and lysine CMK, previously shown to block LF activity on macrophages, did not block LF activity in vitro. These data provide the first direct evidence that LF may act as an endopeptidase. PMID:9573135

  9. Monoclonal antibody against the active site of caeruloplasmin and the ELISA system detecting active caeruloplasmin.

    PubMed

    Hiyamuta, S; Ito, K

    1994-04-01

    Serum caeruloplasmin deficiency is a characteristic biochemical abnormality found in patients with Wilson's disease, but the mechanism of this disease is unknown. Although the phenylenediamine oxidase activity of serum caeruloplasmin is markedly low in patients with Wilson's disease, mRNA of caeruloplasmin exists to some extent. To investigate the deficiency of caeruloplasmin oxidase activity in Wilson's disease, we generated 14 monoclonal antibodies (MAbs) and selected ID1, which had the strongest reactivity, and ID2, which had neutralizing ability. We also established a system to measure active caeruloplasmin specifically using these MAbs. These MAbs and the system will be useful tools in analyzing the active site of caeruloplasmin in patients with Wilson's disease.

  10. Intramembrane Proton Binding Site Linked to Activation of Bacterial Pentameric Ion Channel*

    PubMed Central

    Wang, Hai-Long; Cheng, Xiaolin; Sine, Steven M.

    2012-01-01

    Prokaryotic orthologs of eukaryotic Cys-loop receptor channels recently emerged as structural and mechanistic surrogates to investigate this superfamily of intercellular signaling proteins. Here, we examine proton activation of the prokaryotic ortholog GLIC using patch clamp electrophysiology, mutagenesis, and molecular dynamics (MD) simulations. Whole-cell current recordings from human embryonic kidney (HEK) 293 cells expressing GLIC show half-maximal activation at pH 6, close to the pKa of histidine, implicating the three native His residues in proton sensing linked to activation. The mutation H235F abolishes proton activation, H277Y is without effect, and all nine mutations of His-127 prevent expression on the cell surface. In the GLIC crystal structure, His-235 on transmembrane (TM) α-helix 2, hydrogen bonds to the main chain carbonyl oxygen of Ile-259 on TM α-helix 3. MD simulations show that when His-235 is protonated, the hydrogen bond persists, and the channel remains in the open conformation, whereas when His-235 is deprotonated, the hydrogen bond dissociates, and the channel closes. Mutations of the proximal Tyr-263, which also links TM α-helices 2 and 3 via a hydrogen bond, alter proton sensitivity over a 1.5 pH unit range. MD simulations show that mutations of Tyr-263 alter the hydrogen bonding capacity of His-235. The overall findings show that His-235 in the TM region of GLIC is a novel proton binding site linked to channel activation. PMID:22084238

  11. Hysteretic dynamics of active particles in a periodic orienting field

    PubMed Central

    Romensky, Maksym; Scholz, Dimitri; Lobaskin, Vladimir

    2015-01-01

    Active motion of living organisms and artificial self-propelling particles has been an area of intense research at the interface of biology, chemistry and physics. Significant progress in understanding these phenomena has been related to the observation that dynamic self-organization in active systems has much in common with ordering in equilibrium condensed matter such as spontaneous magnetization in ferromagnets. The velocities of active particles may behave similar to magnetic dipoles and develop global alignment, although interactions between the individuals might be completely different. In this work, we show that the dynamics of active particles in external fields can also be described in a way that resembles equilibrium condensed matter. It follows simple general laws, which are independent of the microscopic details of the system. The dynamics is revealed through hysteresis of the mean velocity of active particles subjected to a periodic orienting field. The hysteresis is measured in computer simulations and experiments on unicellular organisms. We find that the ability of the particles to follow the field scales with the ratio of the field variation period to the particles' orientational relaxation time, which, in turn, is related to the particle self-propulsion power and the energy dissipation rate. The collective behaviour of the particles due to aligning interactions manifests itself at low frequencies via increased persistence of the swarm motion when compared with motion of an individual. By contrast, at high field frequencies, the active group fails to develop the alignment and tends to behave like a set of independent individuals even in the presence of interactions. We also report on asymptotic laws for the hysteretic dynamics of active particles, which resemble those in magnetic systems. The generality of the assumptions in the underlying model suggests that the observed laws might apply to a variety of dynamic phenomena from the motion of

  12. Hysteretic dynamics of active particles in a periodic orienting field.

    PubMed

    Romensky, Maksym; Scholz, Dimitri; Lobaskin, Vladimir

    2015-07-01

    Active motion of living organisms and artificial self-propelling particles has been an area of intense research at the interface of biology, chemistry and physics. Significant progress in understanding these phenomena has been related to the observation that dynamic self-organization in active systems has much in common with ordering in equilibrium condensed matter such as spontaneous magnetization in ferromagnets. The velocities of active particles may behave similar to magnetic dipoles and develop global alignment, although interactions between the individuals might be completely different. In this work, we show that the dynamics of active particles in external fields can also be described in a way that resembles equilibrium condensed matter. It follows simple general laws, which are independent of the microscopic details of the system. The dynamics is revealed through hysteresis of the mean velocity of active particles subjected to a periodic orienting field. The hysteresis is measured in computer simulations and experiments on unicellular organisms. We find that the ability of the particles to follow the field scales with the ratio of the field variation period to the particles' orientational relaxation time, which, in turn, is related to the particle self-propulsion power and the energy dissipation rate. The collective behaviour of the particles due to aligning interactions manifests itself at low frequencies via increased persistence of the swarm motion when compared with motion of an individual. By contrast, at high field frequencies, the active group fails to develop the alignment and tends to behave like a set of independent individuals even in the presence of interactions. We also report on asymptotic laws for the hysteretic dynamics of active particles, which resemble those in magnetic systems. The generality of the assumptions in the underlying model suggests that the observed laws might apply to a variety of dynamic phenomena from the motion of

  13. Robotics and Automation Activities at the Savannah River Site: A Site Report for SUBWOG 39F

    SciTech Connect

    Teese, G.D.

    1995-09-28

    The Savannah River Site has successfully used robots, teleoperators, and remote video to reduce exposure to ionizing radiation, improve worker safety, and improve the quality of operations. Previous reports have described the use of mobile teleoperators in coping with a high level liquid waste spill, the removal of highly contaminated equipment, and the inspection of nuclear reactor vessels. This report will cover recent applications at the Savannah River, as well as systems which SRS has delivered to other DOE site customers.

  14. Control of active sites in selective flocculation: II -- Role of site blocking agents

    SciTech Connect

    Behl, S.; Moudgil, B.M. . Dept. of Materials Science and Engineering)

    1993-12-01

    Control of heteroflocculation using a lower molecular weight fraction of the flocculant as a site blocking agent is demonstrated in the apatite-dolomite-polyethylene oxide system. The most effective SBA (site blocking agent) was determined to be the highest molecular weight fraction of the flocculant itself which was not capable of flocculating any of the components of the mixture. In the presence of the SBA, flocculant adsorption decreased significantly on apatite particles, thereby inhibiting coflocculation.

  15. Fine root dynamics in lodgepole pine and white spruce stands along productivity gradients in reclaimed oil sands sites.

    PubMed

    Jamro, Ghulam Murtaza; Chang, Scott X; Naeth, M Anne; Duan, Min; House, Jason

    2015-10-01

    Open-pit mining activities in the oil sands region of Alberta, Canada, create disturbed lands that, by law, must be reclaimed to a land capability equivalent to that existed before the disturbance. Re-establishment of forest cover will be affected by the production and turnover rate of fine roots. However, the relationship between fine root dynamics and tree growth has not been studied in reclaimed oil sands sites. Fine root properties (root length density, mean surface area, total root biomass, and rates of root production, turnover, and decomposition) were assessed from May to October 2011 and 2012 using sequential coring and ingrowth core methods in lodgepole pine (Pinus contorta Dougl.) and white spruce (Picea glauca (Moench.) Voss) stands. The pine and spruce stands were planted on peat mineral soil mix placed over tailings sand and overburden substrates, respectively, in reclaimed oil sands sites in Alberta. We selected stands that form a productivity gradient (low, medium, and high productivities) of each tree species based on differences in tree height and diameter at breast height (DBH) increments. In lodgepole pine stands, fine root length density and fine root production, and turnover rates were in the order of high > medium > low productivity sites and were positively correlated with tree height and DBH and negatively correlated with soil salinity (P < 0.05). In white spruce stands, fine root surface area was the only parameter that increased along the productivity gradient and was negatively correlated with soil compaction. In conclusion, fine root dynamics along the stand productivity gradients were closely linked to stand productivity and were affected by limiting soil properties related to the specific substrate used for reconstructing the reclaimed soil. Understanding the impact of soil properties on fine root dynamics and overall stand productivity will help improve land reclamation outcomes. PMID:26668730

  16. Fine root dynamics in lodgepole pine and white spruce stands along productivity gradients in reclaimed oil sands sites.

    PubMed

    Jamro, Ghulam Murtaza; Chang, Scott X; Naeth, M Anne; Duan, Min; House, Jason

    2015-10-01

    Open-pit mining activities in the oil sands region of Alberta, Canada, create disturbed lands that, by law, must be reclaimed to a land capability equivalent to that existed before the disturbance. Re-establishment of forest cover will be affected by the production and turnover rate of fine roots. However, the relationship between fine root dynamics and tree growth has not been studied in reclaimed oil sands sites. Fine root properties (root length density, mean surface area, total root biomass, and rates of root production, turnover, and decomposition) were assessed from May to October 2011 and 2012 using sequential coring and ingrowth core methods in lodgepole pine (Pinus contorta Dougl.) and white spruce (Picea glauca (Moench.) Voss) stands. The pine and spruce stands were planted on peat mineral soil mix placed over tailings sand and overburden substrates, respectively, in reclaimed oil sands sites in Alberta. We selected stands that form a productivity gradient (low, medium, and high productivities) of each tree species based on differences in tree height and diameter at breast height (DBH) increments. In lodgepole pine stands, fine root length density and fine root production, and turnover rates were in the order of high > medium > low productivity sites and were positively correlated with tree height and DBH and negatively correlated with soil salinity (P < 0.05). In white spruce stands, fine root surface area was the only parameter that increased along the productivity gradient and was negatively correlated with soil compaction. In conclusion, fine root dynamics along the stand productivity gradients were closely linked to stand productivity and were affected by limiting soil properties related to the specific substrate used for reconstructing the reclaimed soil. Understanding the impact of soil properties on fine root dynamics and overall stand productivity will help improve land reclamation outcomes.

  17. Human activity recognition based on human shape dynamics

    NASA Astrophysics Data System (ADS)

    Cheng, Zhiqing; Mosher, Stephen; Cheng, Huaining; Webb, Timothy

    2013-05-01

    Human activity recognition based on human shape dynamics was investigated in this paper. The shape dynamics describe the spatial-temporal shape deformation of a human body during its movement and thus provide important information about the identity of a human subject and the motions performed by the subject. The dynamic shapes of four subjects in five activities (digging, jogging, limping, throwing, and walking) were created via 3-D motion replication. The Paquet Shape Descriptor (PSD) was used to describe subject shapes in each frame. The principal component analysis was performed on the calculated PSDs and principal components (PCs) were used to characterize PSDs. The PSD calculation was then reasonably approximated by its significant projections in the eigen-space formed by PCs and represented by the corresponding projection coefficients. As such, the dynamic human shapes for each activity were described by these projection coefficients, which in turn, along with their derivatives were used to form the feature vectors (attribute sets) for activity classification. Data mining technology was employed with six classification methods used. Seven attribute sets were evaluated with high classification accuracy attained for most of them. The results from this investigation illustrate the great potential of human shape dynamics for activity recognition.

  18. Overview of MSFC's Applied Fluid Dynamics Analysis Group Activities

    NASA Technical Reports Server (NTRS)

    Garcia, Roberto; Wang, Tee-See; Griffin, Lisa; Turner, James E. (Technical Monitor)

    2001-01-01

    This document is a presentation graphic which reviews the activities of the Applied Fluid Dynamics Analysis Group at Marshall Space Flight Center (i.e., Code TD64). The work of this group focused on supporting the space transportation programs. The work of the group is in Computational Fluid Dynamic tool development. This development is driven by hardware design needs. The major applications for the design and analysis tools are: turbines, pumps, propulsion-to-airframe integration, and combustion devices.

  19. Collective dynamics of soft active particles.

    PubMed

    van Drongelen, Ruben; Pal, Anshuman; Goodrich, Carl P; Idema, Timon

    2015-03-01

    We present a model of soft active particles that leads to a rich array of collective behavior found also in dense biological swarms of bacteria and other unicellular organisms. Our model uses only local interactions, such as Vicsek-type nearest-neighbor alignment, short-range repulsion, and a local boundary term. Changing the relative strength of these interactions leads to migrating swarms, rotating swarms, and jammed swarms, as well as swarms that exhibit run-and-tumble motion, alternating between migration and either rotating or jammed states. Interestingly, although a migrating swarm moves slower than an individual particle, the diffusion constant can be up to three orders of magnitude larger, suggesting that collective motion can be highly advantageous, for example, when searching for food. PMID:25871143

  20. Collective dynamics of soft active particles

    NASA Astrophysics Data System (ADS)

    van Drongelen, Ruben; Pal, Anshuman; Goodrich, Carl P.; Idema, Timon

    2015-03-01

    We present a model of soft active particles that leads to a rich array of collective behavior found also in dense biological swarms of bacteria and other unicellular organisms. Our model uses only local interactions, such as Vicsek-type nearest-neighbor alignment, short-range repulsion, and a local boundary term. Changing the relative strength of these interactions leads to migrating swarms, rotating swarms, and jammed swarms, as well as swarms that exhibit run-and-tumble motion, alternating between migration and either rotating or jammed states. Interestingly, although a migrating swarm moves slower than an individual particle, the diffusion constant can be up to three orders of magnitude larger, suggesting that collective motion can be highly advantageous, for example, when searching for food.

  1. Characterization of the Dielectric Constant in the Trichoderma reesei Cel7B Active Site.

    PubMed

    Song, Xiangfei; Wang, Yefei; Zhang, Shujun; Yan, Shihai; Li, Tong; Yao, Lishan

    2015-07-27

    An attempt is made to evaluate the dielectric constant of the Trichoderma reesei Cel7B active site. Through kinetic measurements, the pKa value of the catalytic acid E201 is determined. Mutations (away from E201) with net charge changes are introduced to perturb the E201 pKa. It is shown that the mutation with a +1 charge change (including G225R, G230R, and A335R) decreases the pKa of E201, whereas the mutation with a -1 charge change (including Q149E, A222D, G225D, and G230D) increases the pKa. This effect is consistent with the electrostatic interaction between the changed charge and the E201 side chain. The fitting of the experimental data yields an apparent dielectric constant of 25-80. Molecular dynamics simulations with explicit water molecules indicate that the high solvent accessibility of the active site contributes largely to the high dielectric constant. ONIOM calculations show that high dielectric constant benefits the catalysis through decreasing the energy of the transition state relative to that of the enzyme substrate complex. PMID:26114648

  2. Multiple, Ligand-Dependent Routes from the Active Site of Cytochrome P450 2C9

    SciTech Connect

    Cojocaru, Vlad; Winn, Peter J.; Wade, Rebecca C.

    2012-02-13

    The active site of liver-specific, drug-metabolizing cytochrome P450 (CYP) monooxygenases is deeply buried in the protein and is connected to the protein surface through multiple tunnels, many of which were found open in different CYP crystal structures. It has been shown that different tunnels could serve as ligand passage routes in different CYPs. However, it is not understood whether one CYP uses multiple routes for substrate access and product release and whether these routes depend on ligand properties. From 300 ns of molecular dynamics simulations of CYP2C9, the second most abundant CYP in the human liver we found four main ligand exit routes, the occurrence of each depending on the ligand type and the conformation of the F-G loop, which is likely to be affected by the CYP-membrane interaction. A non-helical F-G loop favored exit towards the putative membrane-embedded region. Important protein features that direct ligand exit include aromatic residues that divide the active site and whose motions control access to two pathways. The ligands interacted with positively charged residues on the protein surface through hydrogen bonds that appear to select for acidic substrates. The observation of multiple, ligand-dependent routes in a CYP aids understanding of how CYP mutations affect drug metabolism and provides new possibilities for CYP inhibition.

  3. Characterization of the Dielectric Constant in the Trichoderma reesei Cel7B Active Site.

    PubMed

    Song, Xiangfei; Wang, Yefei; Zhang, Shujun; Yan, Shihai; Li, Tong; Yao, Lishan

    2015-07-27

    An attempt is made to evaluate the dielectric constant of the Trichoderma reesei Cel7B active site. Through kinetic measurements, the pKa value of the catalytic acid E201 is determined. Mutations (away from E201) with net charge changes are introduced to perturb the E201 pKa. It is shown that the mutation with a +1 charge change (including G225R, G230R, and A335R) decreases the pKa of E201, whereas the mutation with a -1 charge change (including Q149E, A222D, G225D, and G230D) increases the pKa. This effect is consistent with the electrostatic interaction between the changed charge and the E201 side chain. The fitting of the experimental data yields an apparent dielectric constant of 25-80. Molecular dynamics simulations with explicit water molecules indicate that the high solvent accessibility of the active site contributes largely to the high dielectric constant. ONIOM calculations show that high dielectric constant benefits the catalysis through decreasing the energy of the transition state relative to that of the enzyme substrate complex.

  4. Benzene Probes in Molecular Dynamics Simulations Reveal Novel Binding Sites for Ligand Design.

    PubMed

    Tan, Yaw Sing; Reeks, Judith; Brown, Christopher J; Thean, Dawn; Ferrer Gago, Fernando Jose; Yuen, Tsz Ying; Goh, Eunice Tze Leng; Lee, Xue Er Cheryl; Jennings, Claire E; Joseph, Thomas L; Lakshminarayanan, Rajamani; Lane, David P; Noble, Martin E M; Verma, Chandra S

    2016-09-01

    Protein flexibility poses a major challenge in binding site identification. Several computational pocket detection methods that utilize small-molecule probes in molecular dynamics (MD) simulations have been developed to address this issue. Although they have proven hugely successful at reproducing experimental structural data, their ability to predict new binding sites that are yet to be identified and characterized has not been demonstrated. Here, we report the use of benzenes as probe molecules in ligand-mapping MD (LMMD) simulations to predict the existence of two novel binding sites on the surface of the oncoprotein MDM2. One of them was serendipitously confirmed by biophysical assays and X-ray crystallography to be important for the binding of a new family of hydrocarbon stapled peptides that were specifically designed to target the other putative site. These results highlight the predictive power of LMMD and suggest that predictions derived from LMMD simulations can serve as a reliable basis for the identification of novel ligand binding sites in structure-based drug design. PMID:27532490

  5. Carbon dynamics within agricultural and native sites in the loess region of Western lowa

    USGS Publications Warehouse

    Manies, K.L.; Harden, J.W.; Kramer, L.; Parton, W.J.

    2001-01-01

    In order to quantify the historical changes in carbon storage that result from agricultural conversion, this study compared the carbon dynamics of two sites in the loess region of Iowa: a native prairie and a cropland. Field data were obtained to determine present-day carbon storage and its variability within a landscape (a stable ridgetop vs. eroding upper-midslope vs. depositional lower slope). Models were used to recreate the historical carbon budget of these sites and determine the cropland's potential to be a net CO2 source or sink, relative to the atmosphere. Regardless of slope position, the cropland site contains approximately half the amount of carbon as prairie. Variability in soil carbon storage within a site as a consequence of slope position is as large or larger (variations of 200-300%) than temporal variation (???200% at all slope positions). The most extreme difference in soil carbon storage between the cropland and prairie sites is found in the soil at the upper-midslope, which is the area of greatest erosion. The models estimate that 93-172% of the carbon in the original topsoil has been lost from the cropland's eroding midslope. Much of this carbon is derived from deeper soil horizons. Either a small sink or strong source of carbon to the atmosphere is created, depending on the fate of the eroded sediment and its associated carbon.

  6. Identifying ligand binding sites and poses using GPU-accelerated Hamiltonian replica exchange molecular dynamics

    PubMed Central

    Wang, Kai; Yang, Yanzhi; Chodera, John D.; Shirts, Michael R.

    2014-01-01

    We present a method to identify small molecule ligand binding sites and orientations to a given protein crystal structure using GPU-accelerated Hamiltonian replica exchange molecular dynamics simulations. The Hamiltonians used vary from the physical end state of protein interacting with the ligand to a unphysical end state where the ligand does not interact with the protein. As replicas explore the space of Hamiltonians interpolating between these states the ligand can rapidly escape local minima and explore potential binding sites. Geometric restraints keep the ligands within the protein volume, and a potential energy pathway designed to increase phase space overlap between intermediates ensures good mixing. Because of the rigorous statistical mechanical nature of the Hamiltonian exchange framework, we can also extract binding free energy estimates at all putative binding sites, which agree well with free energies computed from occupation probabilities. We present results of this methodology on the T4 lysozyme L99A model system with four ligands, including one non-binder as a control. We find that our methodology identifies the crystallographic binding sites consistently and accurately for the small number of ligands considered here and gives free energies consistent with experiment. We are also able to analyze the contribution of individual binding sites on the overall binding affinity. Our methodology points to near term potential applications in early-stage drug discovery. PMID:24297454

  7. Promoting the Adsorption of Metal Ions on Kaolinite by Defect Sites: A Molecular Dynamics Study

    PubMed Central

    Li, Xiong; Li, Hang; Yang, Gang

    2015-01-01

    Defect sites exist abundantly in minerals and play a crucial role for a variety of important processes. Here molecular dynamics simulations are used to comprehensively investigate the adsorption behaviors, stabilities and mechanisms of metal ions on defective minerals, considering different ionic concentrations, defect sizes and contents. Outer-sphere adsorbed Pb2+ ions predominate for all models (regular and defective), while inner-sphere Na+ ions, which exist sporadically only at concentrated solutions for regular models, govern the adsorption for all defective models. Adsorption quantities and stabilities of metal ions on kaolinite are fundamentally promoted by defect sites, thus explaining the experimental observations. Defect sites improve the stabilities of both inner- and outer-sphere adsorption, and (quasi) inner-sphere Pb2+ ions emerge only at defect sites that reinforce the interactions. Adsorption configurations are greatly altered by defect sites but respond weakly by changing defect sizes or contents. Both adsorption quantities and stabilities are enhanced by increasing defect sizes or contents, while ionic concentrations mainly affect adsorption quantities. We also find that adsorption of metal ions and anions can be promoted by each other and proceeds in a collaborative mechanism. Results thus obtained are beneficial to comprehend related processes for all types of minerals. PMID:26403873

  8. The effects of spatial patterns in habitat quality on community dynamics within a site

    PubMed Central

    Clarke, R. T.; Thomas, J. A.; Elmes, G. W.; Hochberg, M. E.

    1997-01-01

    Metapopulation studies of single species have shown that the size and spatial arrangement of patches of assumed uniformly 'suitable' habitat can influence their population dynamics and persistence. We investigated whether variation in the spatial arrangement of 'suitable' habitat of varied quality within a single site can affect the abundance and persistence of interacting species. We accomplished this by extending a field-based spatial simulation model of four interacting species at two trophic levels (an endangered butterfly, its larval food-plant, and two ants). The habitat on sites with the same average and range of qualities was rearranged to give varying degrees of local spatial heterogeneity or 'site ruggedness'. We found that the ant species that acts as host to the butterfly caterpillars decreased with site ruggedness. The impact on the butterfly was more substantial: it often failed to persist on very rugged sites. Despite being free-ranging over the whole area, the butterfly's persistence depends on the arrangement of habitat quality at a finer spatial scale, due to its interactions with species possessing narrower habitat niches and more localized dispersal. Ruggedness also influenced the rate of recovery of the host ant, and hence community structure, for more than a century following the butterfly's extinction.

  9. Locating the nucleation sites for protein encapsulated gold nanoclusters: a molecular dynamics and fluorescence study.

    PubMed

    Russell, B A; Kubiak-Ossowska, K; Mulheran, P A; Birch, D J S; Chen, Y

    2015-09-14

    Fluorescent gold nanoclusters encapsulated by proteins have attracted considerable attention in recent years for their unique properties as new fluorescence probes for biological sensing and imaging. However, fundamental questions, such as the nucleation sites of gold nanoclusters within proteins and the fluorescence mechanism remain unsolved. Here we present a study of the location of gold nanoclusters within bovine serum albumin (BSA) combining both fully atomistic molecular dynamic (MD) simulations and fluorescence spectroscopic studies. The MD simulations show gold clusters growing close to a number of cysteine sites across all three domains of BSA, although just two major sites in domains IIB and IA were found to accommodate large clusters comprising more than 12 atoms. The dependence of the fluorescence on pH is found to be compatible with possible nucleation sites in domains IIB and IA. Furthermore, the energy transfer between tryptophan and gold nanoclusters reveals a separation of 29.7 Å, further indicating that gold nanoclusters were most likely located in the major nucleation site in domain IIB. The disclosure of the precise location of the gold nanoclusters and their surrounding amino acid residues should help better understanding of their fluorescence mechanism and aid their optimization as fluorescent nanoprobes.

  10. Following [FeFe] Hydrogenase Active Site Intermediates by Time-Resolved Mid-IR Spectroscopy.

    PubMed

    Mirmohades, Mohammad; Adamska-Venkatesh, Agnieszka; Sommer, Constanze; Reijerse, Edward; Lomoth, Reiner; Lubitz, Wolfgang; Hammarström, Leif

    2016-08-18

    Time-resolved nanosecond mid-infrared spectroscopy is for the first time employed to study the [FeFe] hydrogenase from Chlamydomonas reinhardtii and to investigate relevant intermediates of the enzyme active site. An actinic 355 nm, 10 ns laser flash triggered photodissociation of a carbonyl group from the CO-inhibited state Hox-CO to form the state Hox, which is an intermediate of the catalytic proton reduction cycle. Time-resolved infrared spectroscopy allowed us to directly follow the subsequent rebinding of the carbonyl, re-forming Hox-CO, and determine the reaction half-life to be t1/2 ≈ 13 ± 5 ms at room temperature. This gives direct information on the dynamics of CO inhibition of the enzyme. PMID:27494400

  11. Simulation analysis of formycin 5'-monophosphate analog substrates in the ricin A-chain active site.

    PubMed

    Olson, M A; Scovill, J P; Hack, D C

    1995-06-01

    Ricin is an RNA N-glycosidase that hydrolyzes a single adenine base from a conserved loop of 28S ribosomal RNA, thus inactivating protein synthesis. Molecular-dynamics simulation methods are used to analyze the structural interactions and thermodynamics that govern the binding of formycin 5'-monophosphate (FMP) and several of its analogs to the active site of ricin A-chain. Simulations are carried out initiated from the X-ray crystal structure of the ricin-FMP complex with the ligand modeled as a dianion, monoanion and zwitterion. Relative changes in binding free energies are estimated for FMP analogs constructed from amino substitutions at the 2- and 2'-positions, and from hydroxyl substitution at the 2'-position.

  12. Implication of crystal water molecules in inhibitor binding at ALR2 active site.

    PubMed

    Hymavati; Kumar, Vivek; Sobhia, M Elizabeth

    2012-01-01

    Water molecules play a crucial role in mediating the interaction between a ligand and a macromolecule. The solvent environment around such biomolecule controls their structure and plays important role in protein-ligand interactions. An understanding of the nature and role of these water molecules in the active site of a protein could greatly increase the efficiency of rational drug design approaches. We have performed the comparative crystal structure analysis of aldose reductase to understand the role of crystal water in protein-ligand interaction. Molecular dynamics simulation has shown the versatile nature of water molecules in bridge H bonding during interaction. Occupancy and life time of water molecules depend on the type of cocrystallized ligand present in the structure. The information may be useful in rational approach to customize the ligand, and thereby longer occupancy and life time for bridge H-bonding. PMID:22649481

  13. Following [FeFe] Hydrogenase Active Site Intermediates by Time-Resolved Mid-IR Spectroscopy.

    PubMed

    Mirmohades, Mohammad; Adamska-Venkatesh, Agnieszka; Sommer, Constanze; Reijerse, Edward; Lomoth, Reiner; Lubitz, Wolfgang; Hammarström, Leif

    2016-08-18

    Time-resolved nanosecond mid-infrared spectroscopy is for the first time employed to study the [FeFe] hydrogenase from Chlamydomonas reinhardtii and to investigate relevant intermediates of the enzyme active site. An actinic 355 nm, 10 ns laser flash triggered photodissociation of a carbonyl group from the CO-inhibited state Hox-CO to form the state Hox, which is an intermediate of the catalytic proton reduction cycle. Time-resolved infrared spectroscopy allowed us to directly follow the subsequent rebinding of the carbonyl, re-forming Hox-CO, and determine the reaction half-life to be t1/2 ≈ 13 ± 5 ms at room temperature. This gives direct information on the dynamics of CO inhibition of the enzyme.

  14. Protein-water dynamics in antifreeze protein III activity

    NASA Astrophysics Data System (ADS)

    Xu, Yao; Bäumer, Alexander; Meister, Konrad; Bischak, Connor G.; DeVries, Arthur L.; Leitner, David M.; Havenith, Martina

    2016-03-01

    We combine Terahertz absorption spectroscopy (THz) and molecular dynamics (MD) simulations to investigate the underlying molecular mechanism for the antifreeze activity of one class of antifreeze protein, antifreeze protein type III (AFP-III) with a focus on the collective water hydrogen bond dynamics near the protein. After summarizing our previous work on AFPs, we present a new investigation of the effects of cosolutes on protein antifreeze activity by adding sodium citrate to the protein solution of AFP-III. Our results reveal that for AFP-III, unlike some other AFPs, the addition of the osmolyte sodium citrate does not affect the hydrogen bond dynamics at the protein surface significantly, as indicated by concentration dependent THz measurements. The present data, in combination with our previous THz measurements and molecular simulations, confirm that while long-range solvent perturbation is a necessary condition for the antifreeze activity of AFP-III, the local binding affinity determines the size of the hysteresis.

  15. Influences of Dynamics on Convection at the Tropical Pacific ARM Sites

    SciTech Connect

    Mather, Jim H.

    2004-02-15

    Convection in the Tropical Western Pacific (TWP) region plays an important role in driving global-scale atmospheric circulation. While the TWP is very important, climate models have a difficult time capturing the intricacies of convection in this region. For these reasons, the Atmospheric Radiation Measurement (ARM) program has made the TWP one of its concentrated areas of study. The ARM program now has three sites in the tropical Pacific region: Manus, Nauru, and Darwin. An examination of cloud frequency and radiation data from theses sites reveals variability on a wide range of time scales. At Manus, variability due to intraseasonal processes, including the Madden-Julian Oscillation, is most pronounced but intraseasonal effects are modulated by the annual monsoon cycle as well as by ENSO. In particular, the relative frequency of convection over the large islands of the maritime continent and over the ocean depends on the stage of the monsoon cycle. An important element of ARM's interest in tropical Pacific convection is to characterize cloud optical properties. Because cloud properties are likely to be different for maritime and island-based convection, it is important to understand the dynamical processes that govern the frequency with which each occurs. This paper attempts to attribute variability observed at the tropical Pacific ARM sites to dynamical processes on a range of spatial scales.

  16. Dynamic, Rho1p-dependent localization of Pkc1p to sites of polarized growth.

    PubMed

    Andrews, P D; Stark, M J

    2000-08-01

    In eukaryotes, the Rho GTPases and their effectors are key regulators of the actin cytoskeleton, membrane trafficking and secretion, cell growth, cell cycle progression and cytokinesis. Budding yeast Pkc1p, a protein kinase C-like enzyme involved in cell wall biosynthesis and cytoskeletal polarity, is structurally and functionally related to the Rho-associated kinases (PRK/ROCK) of mammalian cells. In this study, localization of Pkc1p was monitored in live cells using a GFP fusion (Pkc1p-GFP). Pkc1p-GFP showed dynamic spatial and temporal localization at sites of polarized growth. Early in the cell cycle, Pkc1p-GFP was found at the pre-bud site and bud tips, becoming delocalized as the cell progressed further and finally relocalizing around the mother-daughter bud neck in an incomplete ring, which persisted until cell separation. Bud localization was actin-dependent but stability of Pkc1p-GFP at the neck was actin-independent, although localization at both sites required functional Rho1p. In addition, Pkc1p-GFP showed rapid relocalization after cell wall damage. These results suggest that the roles of Pkc1p in both polarized growth and the response to cell wall stress are mediated by dynamic changes in its localization, and suggest an additional potential role in cytokinesis.

  17. Mapping membrane protein backbone dynamics: a comparison of site-directed spin labeling with NMR 15N-relaxation measurements.

    PubMed

    Lo, Ryan H; Kroncke, Brett M; Solomon, Tsega L; Columbus, Linda

    2014-10-01

    The ability to detect nanosecond backbone dynamics with site-directed spin labeling (SDSL) in soluble proteins has been well established. However, for membrane proteins, the nitroxide appears to have more interactions with the protein surface, potentially hindering the sensitivity to backbone motions. To determine whether membrane protein backbone dynamics could be mapped with SDSL, a nitroxide was introduced at 55 independent sites in a model polytopic membrane protein, TM0026. Electron paramagnetic resonance spectral parameters were compared with NMR (15)N-relaxation data. Sequential scans revealed backbone dynamics with the same trends observed for the R1 relaxation rate, suggesting that nitroxide dynamics remain coupled to the backbone on membrane proteins.

  18. In-situ coupling between kinase activities and protein dynamics within single focal adhesions

    PubMed Central

    Wu, Yiqian; Zhang, Kaiwen; Seong, Jihye; Fan, Jason; Chien, Shu; Wang, Yingxiao; Lu, Shaoying

    2016-01-01

    The dynamic activation of oncogenic kinases and regulation of focal adhesions (FAs) are crucial molecular events modulating cell adhesion in cancer metastasis. However, it remains unclear how these events are temporally coordinated at single FA sites. Therefore, we targeted fluorescence resonance energy transfer (FRET)-based biosensors toward subcellular FAs to report local molecular events during cancer cell adhesion. Employing single FA tracking and cross-correlation analysis, we quantified the dynamic coupling characteristics between biochemical kinase activities and structural FA within single FAs. We show that kinase activations and FA assembly are strongly and sequentially correlated, with the concurrent FA assembly and Src activation leading focal adhesion kinase (FAK) activation by 42.6 ± 12.6 sec. Strikingly, the temporal coupling between kinase activation and individual FA assembly reflects the fate of FAs at later stages. The FAs with a tight coupling tend to grow and mature, while the less coupled FAs likely disassemble. During FA disassembly, however, kinase activations lead the disassembly, with FAK being activated earlier than Src. Therefore, by integrating subcellularly targeted FRET biosensors and computational analysis, our study reveals intricate interplays between Src and FAK in regulating the dynamic life of single FAs in cancer cells. PMID:27383747

  19. In-situ coupling between kinase activities and protein dynamics within single focal adhesions.

    PubMed

    Wu, Yiqian; Zhang, Kaiwen; Seong, Jihye; Fan, Jason; Chien, Shu; Wang, Yingxiao; Lu, Shaoying

    2016-01-01

    The dynamic activation of oncogenic kinases and regulation of focal adhesions (FAs) are crucial molecular events modulating cell adhesion in cancer metastasis. However, it remains unclear how these events are temporally coordinated at single FA sites. Therefore, we targeted fluorescence resonance energy transfer (FRET)-based biosensors toward subcellular FAs to report local molecular events during cancer cell adhesion. Employing single FA tracking and cross-correlation analysis, we quantified the dynamic coupling characteristics between biochemical kinase activities and structural FA within single FAs. We show that kinase activations and FA assembly are strongly and sequentially correlated, with the concurrent FA assembly and Src activation leading focal adhesion kinase (FAK) activation by 42.6 ± 12.6 sec. Strikingly, the temporal coupling between kinase activation and individual FA assembly reflects the fate of FAs at later stages. The FAs with a tight coupling tend to grow and mature, while the less coupled FAs likely disassemble. During FA disassembly, however, kinase activations lead the disassembly, with FAK being activated earlier than Src. Therefore, by integrating subcellularly targeted FRET biosensors and computational analysis, our study reveals intricate interplays between Src and FAK in regulating the dynamic life of single FAs in cancer cells. PMID:27383747

  20. Mutation at a Strictly Conserved, Active Site Tyrosine in the Copper Amine Oxidase Leads to Uncontrolled Oxygenase Activity

    SciTech Connect

    Chen, Zhi-wei; Datta, Saumen; DuBois, Jennifer L.; Klinman, Judith P.; Mathews, F. Scott

    2010-09-07

    The copper amine oxidases carry out two copper-dependent processes: production of their own redox-active cofactor (2,4,5-trihydroxyphenylalanine quinone, TPQ) and the subsequent oxidative deamination of substrate amines. Because the same active site pocket must facilitate both reactions, individual active site residues may serve multiple roles. We have examined the roles of a strictly conserved active site tyrosine Y305 in the copper amine oxidase from Hansenula polymorpha kinetically, spetroscopically (Dubois and Klinman (2006) Biochemistry 45, 3178), and, in the present work, structurally. While the Y305A enzyme is almost identical to the wild type, a novel, highly oxygenated species replaces TPQ in the Y305F active sites. This new structure not only provides the first direct detection of peroxy intermediates in cofactor biogenesis but also indicates the critical control of oxidation chemistry that can be conferred by a single active site residue.

  1. Single, competitive, and dynamic adsorption on activated carbon of compounds used as plasticizers and herbicides.

    PubMed

    Abdel daiem, Mahmoud M; Rivera-Utrilla, José; Sánchez-Polo, Manuel; Ocampo-Pérez, Raúl

    2015-12-15

    The main aim of this study was to investigate the single, competitive, and dynamic adsorption of phthalic acid (PA), bisphenol A (BPA), diphenolic acid (DPA), 2,4-dichlorophenoxy-acetic acid (2,4-D), and 4-chloro-2-methylphenoxyacetic acid (MCPA) on two activated carbons with different chemical natures and similar textural characteristics. The adsorption mechanism was also elucidated by analyzing the influence of solution pH and ionic strength. The activated carbons demonstrated high adsorption capacity to remove all micropollutants due to the presence of active sites on their surfaces, which increase dispersive interactions between the activated carbon graphene layers and the aromatic ring of pollutants. The adsorption capacity of the activated carbons increased in the order: DPAdynamic regime, the amount of pollutant adsorbed was much higher for PA, followed by DPA, and was approximately similar for BPA, 2,4-D, and MCPA. Finally, the amount of BPA and DPA adsorbed on activated carbon decreased by around 50% and 70% in the presence of DPA and BPA, respectively, indicating that both compounds are adsorbed on the same adsorption sites of the activated carbon.

  2. Single, competitive, and dynamic adsorption on activated carbon of compounds used as plasticizers and herbicides.

    PubMed

    Abdel daiem, Mahmoud M; Rivera-Utrilla, José; Sánchez-Polo, Manuel; Ocampo-Pérez, Raúl

    2015-12-15

    The main aim of this study was to investigate the single, competitive, and dynamic adsorption of phthalic acid (PA), bisphenol A (BPA), diphenolic acid (DPA), 2,4-dichlorophenoxy-acetic acid (2,4-D), and 4-chloro-2-methylphenoxyacetic acid (MCPA) on two activated carbons with different chemical natures and similar textural characteristics. The adsorption mechanism was also elucidated by analyzing the influence of solution pH and ionic strength. The activated carbons demonstrated high adsorption capacity to remove all micropollutants due to the presence of active sites on their surfaces, which increase dispersive interactions between the activated carbon graphene layers and the aromatic ring of pollutants. The adsorption capacity of the activated carbons increased in the order: DPAdynamic regime, the amount of pollutant adsorbed was much higher for PA, followed by DPA, and was approximately similar for BPA, 2,4-D, and MCPA. Finally, the amount of BPA and DPA adsorbed on activated carbon decreased by around 50% and 70% in the presence of DPA and BPA, respectively, indicating that both compounds are adsorbed on the same adsorption sites of the activated carbon. PMID:26282767

  3. An Essential Viral Transcription Activator Modulates Chromatin Dynamics

    PubMed Central

    Gibeault, Rebecca L.; Bildersheim, Michael D.

    2016-01-01

    Although ICP4 is the only essential transcription activator of herpes simplex virus 1 (HSV-1), its mechanisms of action are still only partially understood. We and others propose a model in which HSV-1 genomes are chromatinized as a cellular defense to inhibit HSV-1 transcription. To counteract silencing, HSV-1 would have evolved proteins that prevent or destabilize chromatinization to activate transcription. These proteins should act as HSV-1 transcription activators. We have shown that HSV-1 genomes are organized in highly dynamic nucleosomes and that histone dynamics increase in cells infected with wild type HSV-1. We now show that whereas HSV-1 mutants encoding no functional ICP0 or VP16 partially enhanced histone dynamics, mutants encoding no functional ICP4 did so only minimally. Transient expression of ICP4 was sufficient to enhance histone dynamics in the absence of other HSV-1 proteins or HSV-1 DNA. The dynamics of H3.1 were increased in cells expressing ICP4 to a greater extent than those of H3.3. The dynamics of H2B were increased in cells expressing ICP4, whereas those of canonical H2A were not. ICP4 preferentially targets silencing H3.1 and may also target the silencing H2A variants. In infected cells, histone dynamics were increased in the viral replication compartments, where ICP4 localizes. These results suggest a mechanism whereby ICP4 activates transcription by disrupting, or preventing the formation of, stable silencing nucleosomes on HSV-1 genomes. PMID:27575707

  4. Identification of promiscuous ene-reductase activity by mining structural databases using active site constellations

    PubMed Central

    Steinkellner, Georg; Gruber, Christian C.; Pavkov-Keller, Tea; Binter, Alexandra; Steiner, Kerstin; Winkler, Christoph; Łyskowski, Andrzej; Schwamberger, Orsolya; Oberer, Monika; Schwab, Helmut; Faber, Kurt; Macheroux, Peter; Gruber, Karl

    2014-01-01

    The exploitation of catalytic promiscuity and the application of de novo design have recently opened the access to novel, non-natural enzymatic activities. Here we describe a structural bioinformatic method for predicting catalytic activities of enzymes based on three-dimensional constellations of functional groups in active sites (‘catalophores’). As a proof-of-concept we identify two enzymes with predicted promiscuous ene-reductase activity (reduction of activated C–C double bonds) and compare them with known ene-reductases, that is, members of the Old Yellow Enzyme family. Despite completely different amino acid sequences, overall structures and protein folds, high-resolution crystal structures reveal equivalent binding modes of typical Old Yellow Enzyme substrates and ligands. Biochemical and biocatalytic data show that the two enzymes indeed possess ene-reductase activity and reveal an inverted stereopreference compared with Old Yellow Enzymes for some substrates. This method could thus be a tool for the identification of viable starting points for the development and engineering of novel biocatalysts. PMID:24954722

  5. Atomistic insights into rhodopsin activation from a dynamic model.

    PubMed

    Tikhonova, Irina G; Best, Robert B; Engel, Stanislav; Gershengorn, Marvin C; Hummer, Gerhard; Costanzi, Stefano

    2008-08-01

    Rhodopsin, the light sensitive receptor responsible for blue-green vision, serves as a prototypical G protein-coupled receptor (GPCR). Upon light absorption, it undergoes a series of conformational changes that lead to the active form, metarhodopsin II (META II), initiating a signaling cascade through binding to the G protein transducin (G(t)). Here, we first develop a structural model of META II by applying experimental distance restraints to the structure of lumi-rhodopsin (LUMI), an earlier intermediate. The restraints are imposed by using a combination of biased molecular dynamics simulations and perturbations to an elastic network model. We characterize the motions of the transmembrane helices in the LUMI-to-META II transition and the rearrangement of interhelical hydrogen bonds. We then simulate rhodopsin activation in a dynamic model to study the path leading from LUMI to our META II model for wild-type rhodopsin and a series of mutants. The simulations show a strong correlation between the transition dynamics and the pharmacological phenotypes of the mutants. These results help identify the molecular mechanisms of activation in both wild type and mutant rhodopsin. While static models can provide insights into the mechanisms of ligand recognition and predict ligand affinity, a dynamic model of activation could be applicable to study the pharmacology of other GPCRs and their ligands, offering a key to predictions of basal activity and ligand efficacy.

  6. Site-specific probing of charge transfer dynamics in organic photovoltaics

    SciTech Connect

    Arion, Tiberiu; Roth, Friedrich; Hussain, Zahid; Eberhardt, Wolfgang

    2015-03-23

    We report the site-specific probing of charge-transfer dynamics in a prototype system for organic photovoltaics (OPVs) by picosecond time-resolved X-ray photoelectron spectroscopy. A layered system consisting of approximately two monolayers of C{sub 60} deposited on top of a thin film of Copper-Phthalocyanine (CuPC) is excited by an optical pump pulse and the induced electronic dynamics are probed with 590 eV X-ray pulses. Charge transfer from the electron donor (CuPC) to the acceptor (C{sub 60}) and subsequent charge carrier dynamics are monitored by recording the time-dependent C 1s core level photoemission spectrum of the system. The arrival of electrons in the C{sub 60} layer is readily observed as a completely reversible, transient shift of the C{sub 60} associated C 1s core level, while the C 1s level of the CuPC remains unchanged. The capability to probe charge transfer and recombination dynamics in OPV assemblies directly in the time domain and from the perspective of well-defined domains is expected to open additional pathways to better understand and optimize the performance of this emerging technology.

  7. An ionizable active-site tryptophan imparts catalase activity to a peroxidase core.

    PubMed

    Loewen, Peter C; Carpena, Xavi; Vidossich, Pietro; Fita, Ignacio; Rovira, Carme

    2014-05-21

    Catalase peroxidases (KatG's) are bifunctional heme proteins that can disproportionate hydrogen peroxide (catalatic reaction) despite their structural dissimilarity with monofunctional catalases. Using X-ray crystallography and QM/MM calculations, we demonstrate that the catalatic reaction of KatG's involves deprotonation of the active-site Trp, which plays a role similar to that of the distal His in monofunctional catalases. The interaction of a nearby mobile arginine with the distal Met-Tyr-Trp essential adduct (in/out) acts as an electronic switch, triggering deprotonation of the adduct Trp.

  8. Structural Origins of Nitroxide Side Chain Dynamics on Membrane Protein [alpha]-Helical Sites

    SciTech Connect

    Kroncke, Brett M.; Horanyi, Peter S.; Columbus, Linda

    2010-12-07

    Understanding the structure and dynamics of membrane proteins in their native, hydrophobic environment is important to understanding how these proteins function. EPR spectroscopy in combination with site-directed spin labeling (SDSL) can measure dynamics and structure of membrane proteins in their native lipid environment; however, until now the dynamics measured have been qualitative due to limited knowledge of the nitroxide spin label's intramolecular motion in the hydrophobic environment. Although several studies have elucidated the structural origins of EPR line shapes of water-soluble proteins, EPR spectra of nitroxide spin-labeled proteins in detergents or lipids have characteristic differences from their water-soluble counterparts, suggesting significant differences in the underlying molecular motion of the spin label between the two environments. To elucidate these differences, membrane-exposed {alpha}-helical sites of the leucine transporter, LeuT, from Aquifex aeolicus, were investigated using X-ray crystallography, mutational analysis, nitroxide side chain derivatives, and spectral simulations in order to obtain a motional model of the nitroxide. For each crystal structure, the nitroxide ring of a disulfide-linked spin label side chain (R1) is resolved and makes contacts with hydrophobic residues on the protein surface. The spin label at site I204 on LeuT makes a nontraditional hydrogen bond with the ortho-hydrogen on its nearest neighbor F208, whereas the spin label at site F177 makes multiple van der Waals contacts with a hydrophobic pocket formed with an adjacent helix. These results coupled with the spectral effect of mutating the i {+-} 3, 4 residues suggest that the spin label has a greater affinity for its local protein environment in the low dielectric than on a water-soluble protein surface. The simulations of the EPR spectra presented here suggest the spin label oscillates about the terminal bond nearest the ring while maintaining weak contact

  9. Molecular dynamics study of liquid methanol with a flexible three-site model

    SciTech Connect

    Palinkas, G.; Hawlicka, E.; Heinzinger, K.

    1987-07-30

    A new potential is presented which describes the methanol-methanol interactions on the basis of a flexible three-site model. The intramolecular part of the potential has been derived from spectroscopic data. A molecular dynamics study has been performed with this potential at 286 K. The structural properties of liquid methanol calculated from the simulations are in good agreement with X-ray measurements. The average geometrical arrangement of nearest neighbors and their hydrogen bonding are discussed. The potential describes correctly the gas-liquid frequency shifts of the intramolecular vibrations. Several thermodynamic properties calculated from the simulation compare favorably with experimental results.

  10. Fast-growing species and sustainability (productivity and site dynamics of three fast-growing species)

    SciTech Connect

    Reddy, A.N.; Sugur, G.V.

    1992-12-31

    Growth of three fast-growing species, raised in a high rainfall zone (2000-2500 mm per annum) has been compared, and the associated site dynamics studies in the Western Ghat area of Karnataka State. Two fast-growing exotics, Acacia auriculiformis and Castuarina equisitifolia, were planted on degraded, open sites at high planting densities (5000 plants ha{sup {minus}1}), and one native fast-growing species. Dendrocalamus strictus, was planted on a good site under seasonal irrigation and wider spacing (500 plants ha{sup {minus}1}). These were studies at the age of 5 years for their comparative productivity, quantity of litter fall and changes in nutrient and microbial status. Among these species, A. auriculiformis recorded the highest total productivity closely followed by D. strictus. However, the MAI after 5 years indicated a higher productivity for D. strictus, when culm production attained harvestable size. C. equisitifolia was a close third. It was also found that D. strictus produced higher biomass at lower planting densities, under better sites and management. The litter fall and changes in nutrient status indicated the highest efficiency in A. auriculiformis, followed by C. equisitifolia. It was concluded that the higher planting density was the major contributing factor; the values were comparatively low for D. strictus mainly owing to a lower stocking density of plants.

  11. Food chain dynamics and potential ecological risks of mercury at the Carson River site

    SciTech Connect

    Peterson, S.C.

    1995-12-31

    The USEPA is conducting a remedial investigation of mercury contamination in the Carson River watershed, located near Carson City in central west Nevada. As a component of this investigation, water, sediment, and tissue samples were collected for mercury speciation and other analyses. Tissues analyses from the seven site-investigation areas and four background areas include: whole-body and fillet analyses of five species of fish, composite and individual analyses of three species of benthic macroinvertebrates, blood, feather and liver analyses of two bird species, composite analyses of zooplankton, and whole-body analyses of lizards. The data are used to develop site-specific estimates of mercury bioaccumulation in aquatic food chains of riverine/riparian, open-water, and mudflat habitats at the Carson River site. Because the behavior and food chain dynamics of mercury in semi-arid ecosystems of the southwestern US is poorly understood, these data can be compared and contrasted with bioaccumulation estimates derived from well-studied ecosystems such as northern temperate lakes. Potential ecological risks of mercury exposure through the food chain and through ingestion of and contact with contaminated media are evaluated for important wildlife receptors occurring at the Carson River site.

  12. Dynamic multistate site occupancy models to evaluate hypotheses relevant to conservation of Golden Eagles in Denali National Park, Alaska

    USGS Publications Warehouse

    Martin, Julien; McIntyre, Carol L.; Hines, James E.; Nichols, James D.; Schmutz, Joel A.; MacCluskie, Maggie C.

    2009-01-01

    The recent development of multistate site occupancy models offers great opportunities to frame and solve decision problems for conservation that can be viewed in terms of site occupancy. These models have several characteristics (e.g., they account for detectability) that make them particularly well suited for addressing management and conservation problems. We applied multistate site occupancy models to evaluate hypotheses related to the conservation and management of Golden Eagles (Aquila chrysaetos) in Denali National Park, Alaska, and provided estimates of transition probabilities among three occupancy states for nesting areas (occupied with successful reproduction, occupied with unsuccessful reproduction, and unoccupied). Our estimation models included the effect of potential recreational activities (hikers) and environmental covariates such as a snowshoe hare (Lepus americanus) index on transition probabilities among the three occupancy states. Based on the most parsimonious model, support for the hypothesis of an effect of potential human disturbance on site occupancy dynamics was equivocal. There was some evidence that potential human disturbance negatively affected local colonization of territories, but there was no evidence of an effect on reproductive performance parameters. In addition, models that assume a positive relationship between the hare index and successful reproduction were well supported by the data. The statistical approach that we used is particularly useful to parameterize management models that can then be used to make optimal decisions related to the management of Golden Eagles in Denali. Although in our case we were particularly interested in managing recreational activities, we believe that such models should be useful to for a broad class of management and conservation problems.

  13. Nuclear Site Security in the Event of Terrorist Activity

    SciTech Connect

    Thomson, M.L.; Sims, J.

    2008-07-01

    This paper, presented as a poster, identifies why ballistic protection should now be considered at nuclear sites to counter terrorist threats. A proven and flexible form of multi purpose protection is described in detail with identification of trial results that show its suitability for this role. (authors)

  14. Controlled Activation of Protein Rotational Dynamics Using Smart Hydrogel Tethering

    SciTech Connect

    Beech, Brenda M.; Xiong, Yijia; Boschek, Curt B.; Baird, Cheryl L.; Bigelow, Diana J.; Mcateer, Kathleen; Squier, Thomas C.

    2014-09-05

    Stimulus-responsive hydrogel materials that stabilize and control protein dynamics have the potential to enable a range of applications to take advantage of the inherent specificity and catalytic efficiencies of proteins. Here we describe the modular construction of a hydrogel using an engineered calmodulin (CaM) within a polyethylene glycol (PEG) matrix that involves the reversible tethering of proteins through an engineered CaM-binding sequence. For these measurements, maltose binding protein (MBP) was isotopically labeled with [13C] and [15N], permitting dynamic structural measurements using TROSY-HSQC NMR spectroscopy. Upon initial formation of hydrogels protein dynamics are suppressed, with concomitant increases in protein stability. Relaxation of the hydrogel matrix following transient heating results in the activation of protein dynamics and restoration of substrate-induced large-amplitude domain motions necessary for substrate binding.

  15. Dynamics of a polymer in an active and viscoelastic bath

    NASA Astrophysics Data System (ADS)

    Vandebroek, Hans; Vanderzande, Carlo

    2015-12-01

    We study the dynamics of an ideal polymer chain in a viscoelastic medium and in the presence of active forces. The motion of the center of mass and of individual monomers is calculated. On time scales that are comparable to the persistence time of the active forces, monomers can move superdiffusively, while on larger time scales subdiffusive behavior occurs. The difference between this subdiffusion and that in the absence of active forces is quantified. We show that the polymer swells in response to active processes and determine how this swelling depends on the viscoelastic properties of the environment. Our results are compared to recent experiments on the motion of chromosomal loci in bacteria.

  16. Preliminary siting activities for new waste handling facilities at the Idaho National Engineering Laboratory

    SciTech Connect

    Taylor, D.D.; Hoskinson, R.L.; Kingsford, C.O.; Ball, L.W.

    1994-09-01

    The Idaho Waste Processing Facility, the Mixed and Low-Level Waste Treatment Facility, and the Mixed and Low-Level Waste Disposal Facility are new waste treatment, storage, and disposal facilities that have been proposed at the Idaho National Engineering Laboratory (INEL). A prime consideration in planning for such facilities is the selection of a site. Since spring of 1992, waste management personnel at the INEL have been involved in activities directed to this end. These activities have resulted in the (a) identification of generic siting criteria, considered applicable to either treatment or disposal facilities for the purpose of preliminary site evaluations and comparisons, (b) selection of six candidate locations for siting,and (c) site-specific characterization of candidate sites relative to selected siting criteria. This report describes the information gathered in the above three categories for the six candidate sites. However, a single, preferred site has not yet been identified. Such a determination requires an overall, composite ranking of the candidate sites, which accounts for the fact that the sites under consideration have different advantages and disadvantages, that no single site is superior to all the others in all the siting criteria, and that the criteria should be assigned different weighing factors depending on whether a site is to host a treatment or a disposal facility. Stakeholder input should now be solicited to help guide the final selection. This input will include (a) siting issues not already identified in the siting, work to date, and (b) relative importances of the individual siting criteria. Final site selection will not be completed until stakeholder input (from the State of Idaho, regulatory agencies, the public, etc.) in the above areas has been obtained and a strategy has been developed to make a composite ranking of all candidate sites that accounts for all the siting criteria.

  17. Active Layer and Moisture Measurements for Intensive Site 0 and 1, Barrow, Alaska

    DOE Data Explorer

    John Peterson

    2015-04-17

    These are measurements of Active Layer Thickness collected along several lines beginning in September, 2011 to the present. The data were collected at several time periods along the Site0 L2 Line, the Site1 AB Line, and an ERT Monitoring Line near Area A in Site1.

  18. Structural mechanism of RuBisCO activation by carbamylation of the active site lysine

    PubMed Central

    Stec, Boguslaw

    2012-01-01

    Ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is a crucial enzyme in carbon fixation and the most abundant protein on earth. It has been studied extensively by biochemical and structural methods; however, the most essential activation step has not yet been described. Here, we describe the mechanistic details of Lys carbamylation that leads to RuBisCO activation by atmospheric CO2. We report two crystal structures of nitrosylated RuBisCO from the red algae Galdieria sulphuraria with O2 and CO2 bound at the active site. G. sulphuraria RuBisCO is inhibited by cysteine nitrosylation that results in trapping of these gaseous ligands. The structure with CO2 defines an elusive, preactivation complex that contains a metal cation Mg2+ surrounded by three H2O/OH molecules. Both structures suggest the mechanism for discriminating gaseous ligands by their quadrupole electric moments. We describe conformational changes that allow for intermittent binding of the metal ion required for activation. On the basis of these structures we propose the individual steps of the activation mechanism. Knowledge of all these elements is indispensable for engineering RuBisCO into a more efficient enzyme for crop enhancement or as a remedy to global warming. PMID:23112176

  19. Experimental dynamic deformation analysis of active stressed lap.

    PubMed

    Zhao, Hongshen; Li, Xiaojin; Fan, Bin; Zeng, Zhige

    2016-02-20

    We introduce a method to measure the dynamic surface deformation of an active stressed lap for fabricating a 4  mf/1.5  mirror. Lap surface accuracy working in some typical deformation velocities is put forward. Experimental results indicate that dynamic lap surface accuracy is worse than that of a static surface, and dynamic surface accuracy gets worse if deformation velocity increases, while the difference of lap surface error RMS is less than 1 μm. An optimization of the processing strategy is feasible through changing the deformation velocity of the active stressed lap depending on the processing schedule. After optimizing the grinding and polishing strategy, efficiency is expected to have a significant increase. PMID:26906568

  20. Conditions of activity bubble uniqueness in dynamic neural fields.

    PubMed

    Mikhailova, Inna; Goerick, Christian

    2005-02-01

    Dynamic neural fields (DNFs) offer a rich spectrum of dynamic properties like hysteresis, spatiotemporal information integration, and coexistence of multiple attractors. These properties make DNFs more and more popular in implementations of sensorimotor loops for autonomous systems. Applications often imply that DNFs should have only one compact region of firing neurons (activity bubble), whereas the rest of the field should not fire (e.g., if the field represents motor commands). In this article we prove the conditions of activity bubble uniqueness in the case of locally symmetric input bubbles. The qualitative condition on inhomogeneous inputs used in earlier work on DNFs is transfered to a quantitative condition of a balance between the internal dynamics and the input. The mathematical analysis is carried out for the two-dimensional case with methods that can be extended to more than two dimensions. The article concludes with an example of how our theoretical results facilitate the practical use of DNFs. PMID:15685393

  1. Using catalytic atom maps to predict the catalytic functions present in enzyme active sites.

    PubMed

    Nosrati, Geoffrey R; Houk, K N

    2012-09-18

    Catalytic atom maps (CAMs) are minimal models of enzyme active sites. The structures in the Protein Data Bank (PDB) were examined to determine if proteins with CAM-like geometries in their active sites all share the same catalytic function. We combined the CAM-based search protocol with a filter based on the weighted contact number (WCN) of the catalytic residues, a measure of the "crowdedness" of the microenvironment around a protein residue. Using this technique, a CAM based on the Ser-His-Asp catalytic triad of trypsin was able to correctly identify catalytic triads in other enzymes within 0.5 Å rmsd of the CAM with 96% accuracy. A CAM based on the Cys-Arg-(Asp/Glu) active site residues from the tyrosine phosphatase active site achieved 89% accuracy in identifying this type of catalytic functionality. Both of these CAMs were able to identify active sites across different fold types. Finally, the PDB was searched to locate proteins with catalytic functionality similar to that present in the active site of orotidine 5'-monophosphate decarboxylase (ODCase), whose mechanism is not known with certainty. A CAM, based on the conserved Lys-Asp-Lys-Asp tetrad in the ODCase active site, was used to search the PDB for enzymes with similar active sites. The ODCase active site has a geometry similar to that of Schiff base-forming Class I aldolases, with lowest aldolase rmsd to the ODCase CAM at 0.48 Å. The similarity between this CAM and the aldolase active site suggests that ODCase has the correct catalytic functionality present in its active site for the generation of a nucleophilic lysine. PMID:22909276

  2. Using Catalytic Atom Maps to Predict the Catalytic Functions Present in Enzyme Active Sites

    PubMed Central

    Nosrati, Geoffrey R.; Houk, K. N.

    2012-01-01

    Catalytic Atom Maps (CAMs) are minimal models of enzyme active sites. The structures in the Protein Data Bank (PDB) were examined to determine if proteins with CAM-like geometries in their active sites all share the same catalytic function. We combined the CAM-based search protocol with a filter based on the weighted contact number (WCN) of the catalytic residues, a measure of the “crowdedness” of the microenvironment around a protein residue. Using this technique, a CAM based on the Ser-His-Asp catalytic triad of trypsin was able to correctly identify catalytic triads in other enzymes within 0.5 Å RMSD of the Catalytic Atom Map with 96% accuracy. A CAM based on the Cys-Arg-(Asp/Glu) active site residues from the tyrosine phosphatase active site achieved 89% accuracy in identifying this type of catalytic functionality. Both of these Catalytic Atom Maps were able to identify active sites across different fold types. Finally, the PDB was searched to locate proteins with catalytic functionality similar to that present in the active site of orotidine 5′-monophosphate decarboxylase (ODCase), whose mechanism is not known with certainty. A CAM, based on the conserved Lys-Asp-Lys-Asp tetrad in the ODCase active site, was used to search the PDB for enzymes with similar active sites. The ODCase active site has a geometry similar to that of Schiff base-forming Class I aldolases, with lowest aldolase RMSD to the ODCase CAM at 0.48 Å. The similarity between this CAM and the aldolase active site suggests that ODCase has the correct catalytic functionality present in its active site for the generation of a nucleophilic lysine. PMID:22909276

  3. Coordination of the Filament Stabilizing Versus Destabilizing Activities of Cofilin Through its Secondary Binding Site on Actin

    PubMed Central

    Aggeli, Dimitra; Kish-Trier, Erik; Lin, Meng Chi; Haarer, Brian; Cingolani, Gino; Cooper, John A.; Wilkens, Stephan; Amberg, David C.

    2014-01-01

    Cofilin is a ubiquitous modulator of actin cytoskeleton dynamics that can both stabilize and destabilize actin filaments depending on its concentration and/or the presence of regulatory co-factors. Three charge-reversal mutants of yeast cofilin, located in cofilin’s filament-specific secondary binding site, were characterized in order to understand why disruption of this site leads to enhanced filament disassembly. Crystal structures of the mutants showed that the mutations specifically affect the secondary actin-binding interface, leaving the primary binding site unaltered. The mutant cofilins show enhanced activity compared to wild-type cofilin in severing and disassembling actin filaments. Electron microscopy and image analysis revealed long actin filaments in the presence of wild-type cofilin, while the mutants induced many short filaments, consistent with enhanced severing. Real-time fluorescence microscopy of labeled actin filaments confirmed that the mutants, unlike wild-type cofilin, were functioning as constitutively active severing proteins. In cells, the mutant cofilins delayed endocytosis, which depends on rapid actin turnover. We conclude that mutating cofilin’s secondary actin-binding site increases cofilin’s ability to sever and depolymerize actin filaments. We hypothesize that activators of cofilin severing, like Aip1p, may act by disrupting the interface between cofilin’s secondary actin-binding site and the actin filament. PMID:24943913

  4. Active synchronization between two different chaotic dynamical system

    NASA Astrophysics Data System (ADS)

    Maheri, M.; Arifin, N. Md; Ismail, F.

    2015-05-01

    In this paper we investigate on the synchronization problem between two different chaotic dynamical system based on the Lyapunov stability theorem by using nonlinear control functions. Active control schemes are used for synchronization Liu system as drive and Rossler system as response. Numerical simulation by using Maple software are used to show effectiveness of the proposed schemes.

  5. Active synchronization between two different chaotic dynamical system

    SciTech Connect

    Maheri, M.; Arifin, N. Md; Ismail, F.

    2015-05-15

    In this paper we investigate on the synchronization problem between two different chaotic dynamical system based on the Lyapunov stability theorem by using nonlinear control functions. Active control schemes are used for synchronization Liu system as drive and Rossler system as response. Numerical simulation by using Maple software are used to show effectiveness of the proposed schemes.

  6. Heterogeneous dynamics in DNA site discrimination by the structurally homologous DNA-binding domains of ETS-family transcription factors.

    PubMed

    He, Gaofei; Tolic, Ana; Bashkin, James K; Poon, Gregory M K

    2015-04-30

    The ETS family of transcription factors exemplifies current uncertainty in how eukaryotic genetic regulators with overlapping DNA sequence preferences achieve target site specificity. PU.1 and Ets-1 represent archetypes for studying site discrimination by ETS proteins because their DNA-binding domains are the most divergent in sequence, yet they share remarkably superimposable DNA-bound structures. To gain insight into the contrasting thermodynamics and kinetics of DNA recognition by these two proteins, we investigated the structure and dynamics of site discrimination by their DNA-binding domains. Electrophoretic mobilities of complexes formed by the two homologs with circularly permuted binding sites showed significant dynamic differences only for DNA complexes of PU.1. Free solution measurements by dynamic light scattering showed PU.1 to be more dynamic than Ets-1; moreover, dynamic changes are strongly coupled to site discrimination by PU.1, but not Ets-1. Interrogation of the protein/DNA interface by DNA footprinting showed similar accessibility to dimethyl sulfate for PU.1/DNA and Ets-1/DNA complexes, indicating that the dynamics of PU.1/DNA complexes reside primarily outside that interface. An information-based analysis of the two homologs' binding motifs suggests a role for dynamic coupling in PU.1's ability to enforce a more stringent sequence preference than Ets-1 and its proximal sequence homologs.

  7. Heterogeneous dynamics in DNA site discrimination by the structurally homologous DNA-binding domains of ETS-family transcription factors.

    PubMed

    He, Gaofei; Tolic, Ana; Bashkin, James K; Poon, Gregory M K

    2015-04-30

    The ETS family of transcription factors exemplifies current uncertainty in how eukaryotic genetic regulators with overlapping DNA sequence preferences achieve target site specificity. PU.1 and Ets-1 represent archetypes for studying site discrimination by ETS proteins because their DNA-binding domains are the most divergent in sequence, yet they share remarkably superimposable DNA-bound structures. To gain insight into the contrasting thermodynamics and kinetics of DNA recognition by these two proteins, we investigated the structure and dynamics of site discrimination by their DNA-binding domains. Electrophoretic mobilities of complexes formed by the two homologs with circularly permuted binding sites showed significant dynamic differences only for DNA complexes of PU.1. Free solution measurements by dynamic light scattering showed PU.1 to be more dynamic than Ets-1; moreover, dynamic changes are strongly coupled to site discrimination by PU.1, but not Ets-1. Interrogation of the protein/DNA interface by DNA footprinting showed similar accessibility to dimethyl sulfate for PU.1/DNA and Ets-1/DNA complexes, indicating that the dynamics of PU.1/DNA complexes reside primarily outside that interface. An information-based analysis of the two homologs' binding motifs suggests a role for dynamic coupling in PU.1's ability to enforce a more stringent sequence preference than Ets-1 and its proximal sequence homologs. PMID:25824951

  8. Hydrogen production by the naked active site of the di-iron hydrogenases in water.

    PubMed

    Zipoli, Federico; Car, Roberto; Cohen, Morrel H; Selloni, Annabella

    2009-10-01

    We explored the reactivity of the active center of the [FeFe]-hydrogenases detached from the enzyme and immersed in acidified water by first-principles Car-Parrinello molecular-dynamics simulations. We focused on the identification of the structures that are stable and metastable in acidified water and on their activity for hydrogen production. Our calculations revealed that the naked active center could be an efficient catalyst provided that electrons are transferred to the cluster. We found that both bridging and terminal isomers are present at equilibrium and that the bridging configuration is essential for efficient hydrogen production. The formation of the hydrogen molecule occurs via sequential protonations of the distal iron and of the N-atom of the S-CH(2)-NH-CH(2)-S chelating group. H(2) desorption does not involve a significant energy barrier, making the process very efficient at room temperature. We established that the bottleneck in the reaction is the direct proton transfer from water to the vacant site of the distal iron. Moreover, we found that even if the terminal isomer is present at the equilibrium, its strong local hydrophobicity prevents poisoning of the cluster. PMID:19737003

  9. Hydrogen production by the naked active site of the di-iron hydrogenases in water.

    PubMed

    Zipoli, Federico; Car, Roberto; Cohen, Morrel H; Selloni, Annabella

    2009-10-01

    We explored the reactivity of the active center of the [FeFe]-hydrogenases detached from the enzyme and immersed in acidified water by first-principles Car-Parrinello molecular-dynamics simulations. We focused on the identification of the structures that are stable and metastable in acidified water and on their activity for hydrogen production. Our calculations revealed that the naked active center could be an efficient catalyst provided that electrons are transferred to the cluster. We found that both bridging and terminal isomers are present at equilibrium and that the bridging configuration is essential for efficient hydrogen production. The formation of the hydrogen molecule occurs via sequential protonations of the distal iron and of the N-atom of the S-CH(2)-NH-CH(2)-S chelating group. H(2) desorption does not involve a significant energy barrier, making the process very efficient at room temperature. We established that the bottleneck in the reaction is the direct proton transfer from water to the vacant site of the distal iron. Moreover, we found that even if the terminal isomer is present at the equilibrium, its strong local hydrophobicity prevents poisoning of the cluster.

  10. Self-assembly of active colloidal molecules with dynamic function

    NASA Astrophysics Data System (ADS)

    Soto, Rodrigo; Golestanian, Ramin

    Catalytically active colloids maintain non-equilibrium conditions in which they produce and deplete chemicals at their surface. While individual colloids that are symmetrically coated do not exhibit dynamical activity, the concentration fields resulting from their chemical activity decay as 1/r and produce gradients that attract or repel other colloids depending on their surface chemistry and ambient variables. This results in a non-equilibrium analogue of ionic systems, but with the remarkable novel feature of action-reaction symmetry breaking. In dilute conditions these active colloids join up to form molecules via generalized ionic bonds. Colloids are found to join up to form self-assembled molecules that could be inert or have spontaneous activity in the form of net translational velocity and spin depending on their symmetry properties and their constituents. As the interactions do not satisfy detailed-balance, it is possible to achieve structures with time dependent functionality. We study a molecule that adopts spontaneous oscillations and another that exhibits a run-and-tumble dynamics similar to bacteria. Our study shows that catalytically active colloids could be used for designing self-assembled structures that posses dynamical functionalities.

  11. Deformation invariant bounding spheres for dynamic active constraints in surgery.

    PubMed

    Bowyer, Stuart A; Rodriguez Y Baena, Ferdinando

    2014-04-01

    Active constraints are collaborative robot control strategies, which can be used to guide a surgeon or protect delicate tissue structures during robot-assisted surgery. Tissue structures of interest often move and deform throughout a surgical intervention, and therefore, dynamic active constraints, which adapt and conform to these changes, are required. A fundamental element of an active constraint controller is the computation of the geometric relationship between the constraint geometry and the surgical instrument. For a static active constraint, there are a variety of computationally efficient methods for computing this relative configuration; however, for a dynamic active constraint, it becomes significantly more challenging. Deformation invariant bounding spheres are a novel bounding volume formulation, which can be used within a hierarchy to allow efficient proximity queries within dynamic active constraints. These bounding spheres are constructed in such a way that as the surface deforms, they do not require time-consuming rebuilds or updates, rather they are implicitly updated and continue to represent the underlying geometry as it changes. Experimental results show that performing proximity queries with deformation invariant bounding sphere hierarchies is faster than common methods from the literature when the deformation rate is within the range expected from conventional imaging systems. PMID:24622983

  12. Probing Binding Sites and Mechanisms of Action of an IKs Activator by Computations and Experiments

    PubMed Central

    Xu, Yu; Wang, Yuhong; Zhang, Mei; Jiang, Min; Rosenhouse-Dantsker, Avia; Wassenaar, Tsjerk; Tseng, Gea-Ny

    2015-01-01

    The slow delayed rectifier (IKs) channel is composed of the KCNQ1 channel and KCNE1 auxiliary subunit, and functions to repolarize action potentials in the human heart. IKs activators may provide therapeutic efficacy for treating long QT syndromes. Here, we show that a new KCNQ1 activator, ML277, can enhance IKs amplitude in adult guinea pig and canine ventricular myocytes. We probe its binding site and mechanism of action by computational analysis based on our recently reported KCNQ1 and KCNQ1/KCNE1 3D models, followed by experimental validation. Results from a pocket analysis and docking exercise suggest that ML277 binds to a side pocket in KCNQ1 and the KCNE1-free side pocket of KCNQ1/KCNE1. Molecular-dynamics (MD) simulations based on the most favorable channel/ML277 docking configurations reveal a well-defined ML277 binding space surrounded by the S2-S3 loop and S4-S5 helix on the intracellular side, and by S4–S6 transmembrane helices on the lateral sides. A detailed analysis of MD trajectories suggests two mechanisms of ML277 action. First, ML277 restricts the conformational dynamics of the KCNQ1 pore, optimizing K+ ion coordination in the selectivity filter and increasing current amplitudes. Second, ML277 binding induces global motions in the channel, including regions critical for KCNQ1 gating transitions. We conclude that ML277 activates IKs by binding to an intersubunit space and allosterically influencing pore conductance and gating transitions. KCNE1 association protects KCNQ1 from an arrhythmogenic (constitutive current-inducing) effect of ML277, but does not preclude its current-enhancing effect. PMID:25564853

  13. Blogs and Social Network Sites as Activity Systems: Exploring Adult Informal Learning Process through Activity Theory Framework

    ERIC Educational Resources Information Center

    Heo, Gyeong Mi; Lee, Romee

    2013-01-01

    This paper uses an Activity Theory framework to explore adult user activities and informal learning processes as reflected in their blogs and social network sites (SNS). Using the assumption that a web-based space is an activity system in which learning occurs, typical features of the components were investigated and each activity system then…

  14. Site-Specific DNA Structural and Dynamic Features Revealed by Nucleotide-Independent Nitroxide Probes†

    PubMed Central

    Popova, Anna M.; Kálai, Tamás; Hideg, Kálmán; Qin, Peter Z.

    2009-01-01

    In site-directed spin labeling, a covalently attached nitroxide probe containing a chemically inert unpaired electron is utilized to obtain information on the local environment of the parent macromolecule. Studies presented here examine the feasibility of probing local DNA structural and dynamic features using a class of nitroxide probes that are linked to chemically substituted phosphorothioate positions at the DNA backbone. Two members of this family, designated as R5 and R5a, were attached to eight different sites of a dodecameric DNA duplex without severely perturbing the native B-form conformation. Measured X-band electron paramagnetic resonance (EPR) spectra, which report on nitroxide rotational motions, were found to vary depending on the location of the label (e.g., duplex center vs termini) and the surrounding DNA sequence. This indicates that R5 and R5a can provide information on the DNA local environment at the level of an individual nucleotide. As these probes can be attached to arbitrary nucleotides within a nucleic acid sequence, they may provide a means to “scan” a given DNA molecule in order to interrogate its local structural and dynamic features. PMID:19650666

  15. A fragment-based approach to probing adenosine recognition sites by using dynamic combinatorial chemistry.

    PubMed

    Scott, Duncan E; Dawes, Gwen J; Ando, Michiyo; Abell, Chris; Ciulli, Alessio

    2009-11-23

    A new strategy that combines the concepts of fragment-based drug design and dynamic combinatorial chemistry (DCC) for targeting adenosine recognition sites on enzymes is reported. We demonstrate the use of 5'-deoxy-5'-thioadenosine as a noncovalent anchor fragment in dynamic combinatorial libraries templated by Mycobacterium tuberculosis pantothenate synthetase. A benzyl disulfide derivative was identified upon library analysis by HPLC. Structural and binding studies of protein-ligand complexes by X-ray crystallography and isothermal titration calorimetry informed the subsequent optimisation of the DCC hit into a disulfide containing the novel meta-nitrobenzyl fragment that targets the pantoate binding site of pantothenate synthetase. Given the prevalence of adenosine-recognition motifs in enzymes, our results provide a proof-of-concept for using this strategy to probe adjacent pockets for a range of adenosine binding enzymes, including other related adenylate-forming ligases, kinases, and ATPases, as well as NAD(P)(H), CoA and FAD(H2) binding proteins.

  16. Site-Specific DNA Structural and Dynamic Features Revealed by Nucleotide-Independent Nitroxide Probes

    SciTech Connect

    Popova, Anna; Kalai, Tamas; Hideg, Kalman; Qin, Peter Z.

    2009-09-15

    In site-directed spin labeling, a covalently attached nitroxide probe containing a chemically inert unpaired electron is utilized to obtain information on the local environment of the parent macromolecule. Studies presented here examine the feasibility of probing local DNA structural and dynamic features using a class of nitroxide probes that are linked to chemically substituted phosphorothioate positions at the DNA backbone. Two members of this family, designated as R5 and R5a, were attached to eight different sites of a dodecameric DNA duplex without severely perturbing the native B-form conformation. Measured X-band electron paramagnetic resonance (EPR) spectra, which report on nitroxide rotational motions, were found to vary depending on the location of the label (e.g., duplex center vs termini) and the surrounding DNA sequence. This indicates that R5 and R5a can provide information on the DNA local environment at the level of an individual nucleotide. As these probes can be attached to arbitrary nucleotides within a nucleic acid sequence, they may provide a means to “scan” a given DNA molecule in order to interrogate its local structural and dynamic features.

  17. Dynamic neural activity during stress signals resilient coping.

    PubMed

    Sinha, Rajita; Lacadie, Cheryl M; Constable, R Todd; Seo, Dongju

    2016-08-01

    Active coping underlies a healthy stress response, but neural processes supporting such resilient coping are not well-known. Using a brief, sustained exposure paradigm contrasting highly stressful, threatening, and violent stimuli versus nonaversive neutral visual stimuli in a functional magnetic resonance imaging (fMRI) study, we show significant subjective, physiologic, and endocrine increases and temporally related dynamically distinct patterns of neural activation in brain circuits underlying the stress response. First, stress-specific sustained increases in the amygdala, striatum, hypothalamus, midbrain, right insula, and right dorsolateral prefrontal cortex (DLPFC) regions supported the stress processing and reactivity circuit. Second, dynamic neural activation during stress versus neutral runs, showing early increases followed by later reduced activation in the ventrolateral prefrontal cortex (VLPFC), dorsal anterior cingulate cortex (dACC), left DLPFC, hippocampus, and left insula, suggested a stress adaptation response network. Finally, dynamic stress-specific mobilization of the ventromedial prefrontal cortex (VmPFC), marked by initial hypoactivity followed by increased VmPFC activation, pointed to the VmPFC as a key locus of the emotional and behavioral control network. Consistent with this finding, greater neural flexibility signals in the VmPFC during stress correlated with active coping ratings whereas lower dynamic activity in the VmPFC also predicted a higher level of maladaptive coping behaviors in real life, including binge alcohol intake, emotional eating, and frequency of arguments and fights. These findings demonstrate acute functional neuroplasticity during stress, with distinct and separable brain networks that underlie critical components of the stress response, and a specific role for VmPFC neuroflexibility in stress-resilient coping. PMID:27432990

  18. Early Site Permit Demonstration Program: Recommendations for communication activities and public participation in the Early Site Permit Demonstration Program

    SciTech Connect

    Not Available

    1993-01-27

    On October 24, 1992, President Bush signed into law the National Energy Policy Act of 1992. The bill is a sweeping, comprehensive overhaul of the Nation`s energy laws, the first in more than a decade. Among other provisions, the National Energy Policy Act reforms the licensing process for new nuclear power plants by adopting a new approach developed by the US Nuclear Regulatory Commission (NRC) in 1989, and upheld in court in 1992. The NRC 10 CFR Part 52 rule is a three-step process that guarantees public participation at each step. The steps are: early site permit approval; standard design certifications; and, combined construction/operating licenses for nuclear power reactors. Licensing reform increases an organization`s ability to respond to future baseload electricity generation needs with less financial risk for ratepayers and the organization. Costly delays can be avoided because design, safety and siting issues will be resolved before a company starts to build a plant. Specifically, early site permit approval allows for site suitability and acceptability issues to be addressed prior to an organization`s commitment to build a plant. Responsibility for site-specific activities, including communications and public participation, rests with those organizations selected to try out early site approval. This plan has been prepared to assist those companies (referred to as sponsoring organizations) in planning their communications and public involvement programs. It provides research findings, information and recommendations to be used by organizations as a resource and starting point in developing their own plans.

  19. Dynamics of telomerase activity in response to acute psychological stress

    PubMed Central

    Epel, Elissa S.; Lin, Jue; Dhabhar, Firdaus S.; Wolkowitz, Owen M.; Puterman, E; Karan, Lori; Blackburn, Elizabeth H.

    2010-01-01

    Telomerase activity plays an essential role in cel0l survival, by lengthening telomeres and promoting cell growth and longevity. It is now possible to quantify the low levels of telomerase activity in human leukocytes. Low basal telomerase activity has been related to chronic stress in people and to chronic glucocorticoid exposure in vitro. Here we test whether leukocyte telomerase activity changes under acute psychological stress. We exposed 44 elderly women, including 22 high stress dementia caregivers and 22 matched low stress controls, to a brief laboratory psychological stressor, while examining changes in telomerase activity of peripheral blood mononuclear cells (PBMC). At baseline, caregivers had lower telomerase activity levels than controls, but during stress telomerase activity increased similarly in both groups. Across the entire sample, subsequent telomerase activity increased by 18% one hour after the end of the stressor (p<0.01). The increase in telomerase activity was independent of changes in numbers or percentages of monocytes, lymphocytes, and specific T cell types, although we cannot fully rule out some potential contribution from immune cell redistribution in the change in telomerase activity. Telomerase activity increases were associated with greater cortisol increases in response to the stressor. Lastly, psychological response to the tasks (greater threat perception) was also related to greater telomerase activity increases in controls. These findings uncover novel relationships of dynamic telomerase activity with exposure to an acute stressor, and with two classic aspects of the stress response -- perceived psychological stress and neuroendocrine (cortisol) responses to the stressor. PMID:20018236

  20. Active Site Structure and Peroxidase Activity of Oxidatively Modified Cytochrome c Species in Complexes with Cardiolipin.

    PubMed

    Capdevila, Daiana A; Oviedo Rouco, Santiago; Tomasina, Florencia; Tortora, Verónica; Demicheli, Verónica; Radi, Rafael; Murgida, Daniel H

    2015-12-29

    We report a resonance Raman and UV-vis characterization of the active site structure of oxidatively modified forms of cytochrome c (Cyt-c) free in solution and in complexes with cardiolipin (CL). The studied post-translational modifications of Cyt-c include methionine sulfoxidation and tyrosine nitration, which lead to altered heme axial ligation and increased peroxidase activity with respect to those of the wild-type protein. In spite of the structural and activity differences between the protein variants free in solution, binding to CL liposomes induces in all cases the formation of a spectroscopically identical bis-His axial coordination conformer that more efficiently promotes lipid peroxidation. The spectroscopic results indicate that the bis-His form is in equilibrium with small amounts of high-spin species, thus suggesting a labile distal His ligand as the basis for the CL-induced increase in enzymatic activity observed for all protein variants. For Cyt-c nitrated at Tyr74 and sulfoxidized at Met80, the measured apparent binding affinities for CL are ∼4 times larger than for wild-type Cyt-c. On the basis of these results, we propose that these post-translational modifications may amplify the pro-apoptotic signal of Cyt-c under oxidative stress conditions at CL concentrations lower than for the unmodified protein.

  1. Identification of ice nucleation active sites on feldspar dust particles.

    PubMed

    Zolles, Tobias; Burkart, Julia; Häusler, Thomas; Pummer, Bernhard; Hitzenberger, Regina; Grothe, Hinrich

    2015-03-19

    Mineral dusts originating from Earth's crust are known to be important atmospheric ice nuclei. In agreement with earlier studies, feldspar was found as the most active of the tested natural mineral dusts. Here we investigated in closer detail the reasons for its activity and the difference in the activity of the different feldspars. Conclusions are drawn from scanning electron microscopy, X-ray powder diffraction, infrared spectroscopy, and oil-immersion freezing experiments. K-feldspar showed by far the highest ice nucleation activity. Finally, we give a potential explanation of this effect, finding alkali-metal ions having different hydration shells and thus an influence on the ice nucleation activity of feldspar surfaces. PMID:25584435

  2. Identification of Ice Nucleation Active Sites on Feldspar Dust Particles

    PubMed Central

    2015-01-01

    Mineral dusts originating from Earth’s crust are known to be important atmospheric ice nuclei. In agreement with earlier studies, feldspar was found as the most active of the tested natural mineral dusts. Here we investigated in closer detail the reasons for its activity and the difference in the activity of the different feldspars. Conclusions are drawn from scanning electron microscopy, X-ray powder diffraction, infrared spectroscopy, and oil-immersion freezing experiments. K-feldspar showed by far the highest ice nucleation activity. Finally, we give a potential explanation of this effect, finding alkali-metal ions having different hydration shells and thus an influence on the ice nucleation activity of feldspar surfaces. PMID:25584435

  3. Fault zone damage, nonlinear site response, and dynamic triggering associated with seismic waves

    NASA Astrophysics Data System (ADS)

    Wu, Chunquan

    My dissertation focuses primarily on the following three aspects associated with passing seismic waves in the field of earthquake seismology: temporal changes of fault zone properties, nonlinear site response, and dynamic triggering. I systematically analyze temporal changes of fault zone (FZ) site response along the Karadere-Duzce branch of the North Anatolian fault that ruptured during the 1999 Izmit and Duzce earthquake sequences. These results provide a bridge between the large-amplitude near-instantaneous changes and the lower-amplitude longer-duration variations observed in previous studies. The temporal changes measured from this high-resolution spectral ratio analysis also provide a refinement for the beginning of the longer more gradual process typically observed by analyzing repeating earthquakes. I use the same sliding-window spectral ratio technique to analyze temporal changes in site response associated with the strong ground motion of the Mw6.6 2004 Mid-Niigata earthquake sequence recorded by the borehole stations in Japanese Digital Strong-Motion Seismograph Network (KiK-Net). The results suggest that at a given site the input ground motion plays an important role in controlling both the coseismic change and postseismic recovery in site response. In a follow-up study, I apply the same sliding-window spectral ratio technique to surface and borehole strong motion records at 6 KiK-Net sites, and stack results associated with different earthquakes that produce similar PGAs. In some cases I observe a weak coseismic drop in the peak frequency when the PGA is as small as ˜20--30 Gal, and near instantaneous recovery after the passage of the direct S waves. The percentage of drop in the peak frequency starts to increase with increasing PGA values. A coseismic drop in the peak spectral ratio is also observed at 2 sites. When the PGA is larger than ˜60 Gal to more than 100 Gal, considerably stronger coseismic drops of the peak frequencies are observed

  4. Stereospecific suppression of active site mutants by methylphosphonate substituted substrates reveals the stereochemical course of site-specific DNA recombination.

    PubMed

    Rowley, Paul A; Kachroo, Aashiq H; Ma, Chien-Hui; Maciaszek, Anna D; Guga, Piotr; Jayaram, Makkuni

    2015-07-13

    Tyrosine site-specific recombinases, which promote one class of biologically important phosphoryl transfer reactions in DNA, exemplify active site mechanisms for stabilizing the phosphate transition state. A highly conserved arginine duo (Arg-I; Arg-II) of the recombinase active site plays a crucial role in this function. Cre and Flp recombinase mutants lacking either arginine can be rescued by compensatory charge neutralization of the scissile phosphate via methylphosphonate (MeP) modification. The chemical chirality of MeP, in conjunction with mutant recombinases, reveals the stereochemical contributions of Arg-I and Arg-II. The SP preference of the native reaction is specified primarily by Arg-I. MeP reaction supported by Arg-II is nearly bias-free or RP-biased, depending on the Arg-I substituent. Positional conservation of the arginines does not translate into strict functional conservation. Charge reversal by glutamic acid substitution at Arg-I or Arg-II has opposite effects on Cre and Flp in MeP reactions. In Flp, the base immediately 5' to the scissile MeP strongly influences the choice between the catalytic tyrosine and water as the nucleophile for strand scission, thus between productive recombination and futile hydrolysis. The recombinase active site embodies the evolutionary optimization of interactions that not only favor the normal reaction but also proscribe antithetical side reactions. PMID:25999343

  5. Stereospecific suppression of active site mutants by methylphosphonate substituted substrates reveals the stereochemical course of site-specific DNA recombination.

    PubMed

    Rowley, Paul A; Kachroo, Aashiq H; Ma, Chien-Hui; Maciaszek, Anna D; Guga, Piotr; Jayaram, Makkuni

    2015-07-13

    Tyrosine site-specific recombinases, which promote one class of biologically important phosphoryl transfer reactions in DNA, exemplify active site mechanisms for stabilizing the phosphate transition state. A highly conserved arginine duo (Arg-I; Arg-II) of the recombinase active site plays a crucial role in this function. Cre and Flp recombinase mutants lacking either arginine can be rescued by compensatory charge neutralization of the scissile phosphate via methylphosphonate (MeP) modification. The chemical chirality of MeP, in conjunction with mutant recombinases, reveals the stereochemical contributions of Arg-I and Arg-II. The SP preference of the native reaction is specified primarily by Arg-I. MeP reaction supported by Arg-II is nearly bias-free or RP-biased, depending on the Arg-I substituent. Positional conservation of the arginines does not translate into strict functional conservation. Charge reversal by glutamic acid substitution at Arg-I or Arg-II has opposite effects on Cre and Flp in MeP reactions. In Flp, the base immediately 5' to the scissile MeP strongly influences the choice between the catalytic tyrosine and water as the nucleophile for strand scission, thus between productive recombination and futile hydrolysis. The recombinase active site embodies the evolutionary optimization of interactions that not only favor the normal reaction but also proscribe antithetical side reactions.

  6. Active site densities, oxygen activation and adsorbed reactive oxygen in alcohol activation on npAu catalysts.

    PubMed

    Wang, Lu-Cun; Friend, C M; Fushimi, Rebecca; Madix, Robert J

    2016-07-01

    The activation of molecular O2 as well as the reactivity of adsorbed oxygen species is of central importance in aerobic selective oxidation chemistry on Au-based catalysts. Herein, we address the issue of O2 activation on unsupported nanoporous gold (npAu) catalysts by applying a transient pressure technique, a temporal analysis of products (TAP) reactor, to measure the saturation coverage of atomic oxygen, its collisional dissociation probability, the activation barrier for O2 dissociation, and the facility with which adsorbed O species activate methanol, the initial step in the catalytic cycle of esterification. The results from these experiments indicate that molecular O2 dissociation is associated with surface silver, that the density of reactive sites is quite low, that adsorbed oxygen atoms do not spill over from the sites of activation onto the surrounding surface, and that methanol reacts quite facilely with the adsorbed oxygen atoms. In addition, the O species from O2 dissociation exhibits reactivity for the selective oxidation of methanol but not for CO. The TAP experiments also revealed that the surface of the npAu catalyst is saturated with adsorbed O under steady state reaction conditions, at least for the pulse reaction. PMID:27376884

  7. Satellite Dynamic Damping via Active Force Control Augmentation

    NASA Astrophysics Data System (ADS)

    Varatharajoo, Renuganth

    2012-07-01

    An approach that incorporates the Active Force Control (AFC) technique into a conventional Proportional-Derivative (PD) controller is proposed for a satellite active dynamic damping towards a full attitude control. The AFC method has been established to facilitate a robust motion control of dynamical systems in the presence of disturbances, parametric uncertainties and changes that are commonly prevalent in the real-world environment. The usefulness of the method can be extended by introducing intelligent mechanisms to approximate the mass or inertia matrix of the dynamic system to trigger the compensation effect of the controller. AFC is a technique that relies on the appropriate estimation of the inertial or mass parameters of the dynamic system and the measurements of the acceleration and force signals induced by the system if practical implementation is ever considered. In AFC, it is shown that the system subjected to a number of disturbances remains stable and robust via the compensating action of the control strategy. We demonstrate that it is possible to design a spacecraft attitude feedback controller that will ensure the system dynamics set point remains unchanged even in the presence of the disturbances provided that the actual disturbances can be modeled effectively. In order to further facilitate this analysis, a combined energy and attitude control system (CEACS) is proposed as a model satellite attitude control actuator. All the governing equations are established and the proposed satellite attitude control architecture is made amenable to numerical treatments. The results show that the PD-AFC attitude damping performances are superiorly better than that of the solely PD type. It is also shown that the tunings of the AFC system gains are crucial to ensure a better attitude damping performance and this process is mandatory for AFC systems. Finally, the results demonstrate an important satellite dynamic damping enhancement capability using the AFC

  8. Comparison of Chlorpyrifos-Oxon and Paraoxon Acetylcholinesterase Inhibition Dynamics: Potential role of a peripheral binding site

    SciTech Connect

    Kousba, Ahmed A.; Sultatos, L G.; Poet, Torka S.; Timchalk, Chuck

    2004-08-02

    The primary mechanism of action for organophosphorus (OP) insecticides involves the inhibition of acetylcholinesterase (AChE) by oxygenated metabolites (oxons). This inhibition has been attributed to the phosphorylation of the serine hydroxyl group located in the active site of the AChE molecule. The rate of phosphorylation is described by the bimolecular inhibitory rate constant (ki), which has been utilized for quantification of OP inhibitory capacity. It has been previously proposed that a peripheral binding site exists on the AChE molecule, which when occupied, reduces the capacity of additional oxon molecules to phosphorylate the active site. The objective of the current study was to evaluate the interaction of chlorpyrifos oxon (CPO) and paraoxon (PO) with rat brain AChE using a modified Ellman assay in conjunction with a pharmacodynamic model to further assess the dynamics of AChE inhibition and the potential role of a peripheral binding site. The ki for AChE inhibition determined at oxon concentrations of 5 x 10{sup -4} 100 nM were 0.212 and 0.0216 nM-1h-1 for CPO and PO, respectively. The spontaneous reactivation rates of the inhibited AChE for CPO and PO were 0.087 and 0.078 h-1, respectively. In contrast, the ki estimated at a low oxon concentration (1 pM) were {approx} 1,000 and 10,000 -fold higher than those determined at high CPO and PO concentrations, respectively. At these low concentrations, the ki estimates were approximately similar for both CPO and PO (180 and 250 nM-1h-1, respectively). This implies that at low exposure concentrations, both oxons exhibited similar inhibitory potency in contrast to the marked difference exhibited at higher concentrations, which is consistent with the presence of a peripheral binding site on the AChE enzyme. These results support the potential importance of a secondary binding site associated with AChE kinetics, particularly at low environmentally relevant concentrations.

  9. Does Changing the Predicted Dynamics of a Phospholipase C Alter Activity and Membrane Binding?

    PubMed Central

    Cheng, Jiongjia; Karri, Sashank; Grauffel, Cédric; Wang, Fang; Reuter, Nathalie; Roberts, Mary F.; Wintrode, Patrick L.; Gershenson, Anne

    2013-01-01

    The enzymatic activity of secreted phosphatidylinositol-specific phospholipase C (PI-PLC) enzymes is associated with bacterial virulence. Although the PI-PLC active site has no obvious lid, molecular-dynamics simulations suggest that correlated loop motions may limit access to the active site, and two Pro residues, Pro245 and Pro254, are associated with these correlated motions. Whereas the region containing both Pro residues is quite variable among PI-PLCs, it shows high conservation in virulence-associated, secreted PI-PLCs that bind to the surface of cells. These regions of the protein are also associated with phosphatidylcholine binding, which enhances PI-PLC activity. In silico mutagenesis of Pro245 disrupts correlated motions between the two halves of Bacillus thuringiensis PI-PLC, and Pro245 variants show significantly reduced enzymatic activity in all assay systems. PC still enhanced activity, but not to the level of wild-type enzyme. Mutagenesis of Pro254 appears to stiffen the PI-PLC structure, but experimental mutations had minor effects on activity and membrane binding. With the exception of P245Y, reduced activity was not associated with reduced membrane affinity. This combination of simulations and experiments suggests that correlated motions between the two halves of PI-PLC may be more important for enzymatic activity than for vesicle binding. PMID:23332071

  10. Framework for coordination of activities in dynamic situations

    NASA Astrophysics Data System (ADS)

    Franke, Jörn; Charoy, François; El Khoury, Paul

    2013-02-01

    Recent disasters, such as Hurricane Katrina in 2005, have shown several issues for the coordination of human activities in these dynamic situations. Contemporary tools for the coordination used in the disaster response, such as e-mail, Whiteboards or phones, only allow for unstructured coordination, which can cause coordination problems. Hence, we discuss about the current information systems for coordinating the activities in a structured manner and identify their weaknesses in the context of a process modelling effort conducted together with experienced disaster managers. Afterwards, we propose a framework for coordination of activities in dynamic situations. The framework presented in this paper has been implemented as an extension to an open collaboration service. This shows how it can be used in the context of other tools required for disaster response management, such as maps, pictures or videos of the situation. The work described here is the foundation for enabling inter-organisational coordination of activities relevant in other domains, e.g. enterprise support processes, production processes or distributed software development projects. Furthermore, comments by disaster managers show that the concepts are relevant for their work. The expected impact is a more effective and efficient coordination of human activities in dynamic situations by structuring what needs to be coordinated.

  11. The active site of hen egg-white lysozyme: flexibility and chemical bonding

    SciTech Connect

    Held, Jeanette Smaalen, Sander van

    2014-04-01

    Chemical bonding at the active site of lysozyme is analyzed on the basis of a multipole model employing transferable multipole parameters from a database. Large B factors at low temperatures reflect frozen-in disorder, but therefore prevent a meaningful free refinement of multipole parameters. Chemical bonding at the active site of hen egg-white lysozyme (HEWL) is analyzed on the basis of Bader’s quantum theory of atoms in molecules [QTAIM; Bader (1994 ▶), Atoms in Molecules: A Quantum Theory. Oxford University Press] applied to electron-density maps derived from a multipole model. The observation is made that the atomic displacement parameters (ADPs) of HEWL at a temperature of 100 K are larger than ADPs in crystals of small biological molecules at 298 K. This feature shows that the ADPs in the cold crystals of HEWL reflect frozen-in disorder rather than thermal vibrations of the atoms. Directly generalizing the results of multipole studies on small-molecule crystals, the important consequence for electron-density analysis of protein crystals is that multipole parameters cannot be independently varied in a meaningful way in structure refinements. Instead, a multipole model for HEWL has been developed by refinement of atomic coordinates and ADPs against the X-ray diffraction data of Wang and coworkers [Wang et al. (2007), Acta Cryst. D63, 1254–1268], while multipole parameters were fixed to the values for transferable multipole parameters from the ELMAM2 database [Domagala et al. (2012), Acta Cryst. A68, 337–351] . Static and dynamic electron densities based on this multipole model are presented. Analysis of their topological properties according to the QTAIM shows that the covalent bonds possess similar properties to the covalent bonds of small molecules. Hydrogen bonds of intermediate strength are identified for the Glu35 and Asp52 residues, which are considered to be essential parts of the active site of HEWL. Furthermore, a series of weak C

  12. Defect dynamics and ordering in compressible active nematics

    NASA Astrophysics Data System (ADS)

    Mishra, Prashant; Srivastava, Pragya; Marchetti, M. Cristina

    Active nematics, such as suspensions of biopolymers activated by molecular motors or bacteria swimming in passive liquid crystals, exhibit complex self-sustained flow, excitability and defect generation. Activity renders the defect themselves self-propelled particles, capable of organizing in emergent ordered structures. We have developed a minimal model of compressible active nematics on a substrate. We eliminate the flow velocity in favor of the nematic order parameter via the balance of frictional dissipation and active driving to obtain a dynamical description entirely in terms of the nematic alignment order parameter. Activity renormalizes the bend and splay elastic constants rendering them anisotropic and driving them to zero or even negative, resulting in the appearance of modulated states and defective structures. Using linear stability analysis and numerics we organize the various regimes into a phase diagram and discuss the relation to experiments. This work was supported by NSF-DMR-1305184.

  13. Interplay activity-connectivity: Dynamics in patterned neuronal cultures

    NASA Astrophysics Data System (ADS)

    Tibau, E.; Bendiksen, Ch.; Teller, S.; Amigó, N.; Soriano, J.

    2013-01-01

    The ability of a neuronal tissue to efficiently process and transmit information depends on both the intrinsic dynamical properties of the neurons and the connectivity between them. One of the few experimental systems where one can vary the connectivity of a neuronal network in a control manner are neuronal cultures. Here we show that, by combining neuronal cultures with different pattering techniques, we can control and dictate the connectivity of neuronal networks. The emerging cultures are characterized by a rich spontaneous activity, but with some dynamical traits that can be ascribed to the underlying, engineered wiring architecture. Simple patterned cultures can be obtained by plating neurons onto predefined topographical molds, which guide neurons and connections through complex paths. In contrast to homogeneous cultures, characterized by an on/off behavior where all neurons fire in a short time window, patterned cultures show more complex spatio-temporal dynamics, and with varying propagation paths and velocities. Patterned cultures provide a valuable tool to understand not only the interplay activity-connectivity, but also aspects such as the emergence and maintenance of spontaneous activity, synchronization, or the presence of specific dynamic motifs.

  14. Evolution of Site-Selection Stabilizes Population Dynamics, Promotes Even Distribution of Individuals, and Occasionally Causes Evolutionary Suicide.

    PubMed

    Parvinen, Kalle; Brännström, Åke

    2016-08-01

    Species that compete for access to or use of sites, such as parasitic mites attaching to honey bees or apple maggots laying eggs in fruits, can potentially increase their fitness by carefully selecting sites at which they face little or no competition. Here, we systematically investigate the evolution of site-selection strategies among animals competing for discrete sites. By developing and analyzing a mechanistic and population-dynamical model of site selection in which searching individuals encounter sites sequentially and can choose to accept or continue to search based on how many conspecifics are already there, we give a complete characterization of the different site-selection strategies that can evolve. We find that evolution of site-selection stabilizes population dynamics, promotes even distribution of individuals among sites, and occasionally causes evolutionary suicide. We also discuss the broader implications of our findings and propose how they can be reconciled with an earlier study (Nonaka et al. in J Theor Biol 317:96-104, 2013) that reported selection toward ever higher levels of aggregation among sites as a consequence of site-selection. PMID:27647007

  15. Evolution of Site-Selection Stabilizes Population Dynamics, Promotes Even Distribution of Individuals, and Occasionally Causes Evolutionary Suicide.

    PubMed

    Parvinen, Kalle; Brännström, Åke

    2016-08-01

    Species that compete for access to or use of sites, such as parasitic mites attaching to honey bees or apple maggots laying eggs in fruits, can potentially increase their fitness by carefully selecting sites at which they face little or no competition. Here, we systematically investigate the evolution of site-selection strategies among animals competing for discrete sites. By developing and analyzing a mechanistic and population-dynamical model of site selection in which searching individuals encounter sites sequentially and can choose to accept or continue to search based on how many conspecifics are already there, we give a complete characterization of the different site-selection strategies that can evolve. We find that evolution of site-selection stabilizes population dynamics, promotes even distribution of individuals among sites, and occasionally causes evolutionary suicide. We also discuss the broader implications of our findings and propose how they can be reconciled with an earlier study (Nonaka et al. in J Theor Biol 317:96-104, 2013) that reported selection toward ever higher levels of aggregation among sites as a consequence of site-selection.

  16. Proteorhodopsin Activation Is Modulated by Dynamic Changes in Internal Hydration.

    PubMed

    Feng, Jun; Mertz, Blake

    2015-12-01

    Proteorhodopsin, a member of the microbial rhodopsin family, is a seven-transmembrane α-helical protein that functions as a light-driven proton pump. Understanding the proton-pumping mechanism of proteorhodopsin requires intimate knowledge of the proton transfer pathway via complex hydrogen-bonding networks formed by amino acid residues and internal water molecules. Here we conducted a series of microsecond time scale molecular dynamics simulations on both the dark state and the initial photoactivated state of blue proteorhodopsin to reveal the structural basis for proton transfer with respect to protein internal hydration. A complex series of dynamic hydrogen-bonding networks involving water molecules exists, facilitated by water channels and hydration sites within proteorhodopsin. High levels of hydration were discovered at each proton transfer site-the retinal binding pocket and proton uptake and release sites-underscoring the critical participation of water molecules in the proton-pumping mechanism. Water-bridged interactions and local water channels were also observed and can potentially mediate long-distance proton transfer between each site. The most significant phenomenon is after isomerization of retinal, an increase in water flux occurs that connects the proton release group, a conserved arginine residue, and the retinal binding pocket. Our results provide a detailed description of the internal hydration of the early photointermediates in the proteorhodopsin photocycle under alkaline pH conditions. These results lay the fundamental groundwork for understanding the intimate role that hydration plays in the structure-function relationship underlying the proteorhodopsin proton-pumping mechanism, as well as providing context for the relationship of hydration in proteorhodopsin to other microbial retinal proteins. PMID:26562497

  17. Possible active site of the sweet-tasting protein thaumatin.

    PubMed

    Slootstra, J W; De Geus, P; Haas, H; Verrips, C T; Meloen, R H

    1995-10-01

    Epitopes on thaumatin and monellin were studied using the PEPSCAN-technology. The antibodies used were raised against thaumatin. Only antibodies that, in an ELISA, both recognized thaumatin and monellin were used in the PEPSCAN-analyses. On thaumatin two major overlapping epitopes were identified. On monellin no epitopes could be identified. The identified epitope region on thaumatin shares structural features with various peptide and protein sweeteners. It contains an aspartame-like site which is formed by Asp21 and Phe80, tips of the two extruding loops KGDAALDAGGR19-29 and CKRFGRPP77-84, which are spatially positioned next to each other. Furthermore, sub-sequences of the KGDAALDAGGR19-29 loop are similar to peptide-sweeteners such as L-Asp-D-Ala-L-Ala-methyl ester and L-Asp-D-Ala-Gly-methyl ester. Since the aspartame-like Asp21-Phe80 site and the peptide-sweetener-like sequences are also not present in non-sweet thaumatin-like proteins it is postulated that the KGDAALDAGGR19-29- and CKRFGRPP77-84 loop contain important sweet-taste determinants. This region has previously not been implicated as a sweet-taste determinant of thaumatin.

  18. Characterizing and modeling the dynamics of activity and popularity.

    PubMed

    Zhang, Peng; Li, Menghui; Gao, Liang; Fan, Ying; Di, Zengru

    2014-01-01

    Social media, regarded as two-layer networks consisting of users and items, turn out to be the most important channels for access to massive information in the era of Web 2.0. The dynamics of human activity and item popularity is a crucial issue in social media networks. In this paper, by analyzing the growth of user activity and item popularity in four empirical social media networks, i.e., Amazon, Flickr, Delicious and Wikipedia, it is found that cross links between users and items are more likely to be created by active users and to be acquired by popular items, where user activity and item popularity are measured by the number of cross links associated with users and items. This indicates that users generally trace popular items, overall. However, it is found that the inactive users more severely trace popular items than the active users. Inspired by empirical analysis, we propose an evolving model for such networks, in which the evolution is driven only by two-step random walk. Numerical experiments verified that the model can qualitatively reproduce the distributions of user activity and item popularity observed in empirical networks. These results might shed light on the understandings of micro dynamics of activity and popularity in social media networks. PMID:24586586

  19. Characterizing and modeling the dynamics of activity and popularity.

    PubMed

    Zhang, Peng; Li, Menghui; Gao, Liang; Fan, Ying; Di, Zengru

    2014-01-01

    Social media, regarded as two-layer networks consisting of users and items, turn out to be the most important channels for access to massive information in the era of Web 2.0. The dynamics of human activity and item popularity is a crucial issue in social media networks. In this paper, by analyzing the growth of user activity and item popularity in four empirical social media networks, i.e., Amazon, Flickr, Delicious and Wikipedia, it is found that cross links between users and items are more likely to be created by active users and to be acquired by popular items, where user activity and item popularity are measured by the number of cross links associated with users and items. This indicates that users generally trace popular items, overall. However, it is found that the inactive users more severely trace popular items than the active users. Inspired by empirical analysis, we propose an evolving model for such networks, in which the evolution is driven only by two-step random walk. Numerical experiments verified that the model can qualitatively reproduce the distributions of user activity and item popularity observed in empirical networks. These results might shed light on the understandings of micro dynamics of activity and popularity in social media networks.

  20. Assessment of activation products in the Savannah River Site environment

    SciTech Connect

    Carlton, W.H.; Denham, M.

    1996-07-01

    This document assesses the impact of radioactive activation products released from SRS facilities since the first reactor became operational late in 1953. The isotopes reported here are those whose release resulted in the highest dose to people living near SRS: {sup 32}P, {sup 51}Cr, {sup 60}C, and {sup 65}Zn. Release pathways, emission control features, and annual releases to the aqueous and atmospheric environments are discussed. No single incident has resulted in a major acute release of activation products to the environment. The releases were the result of normal operations of the reactors and separations facilities. Releases declined over the years as better controls were established and production was reduced. The overall radiological impact of SRS activation product atmospheric releases from 1954 through 1994 on the offsite maximally exposed individual can be characterized by a total dose of 0.76 mrem. During the same period, such an individual received a total dose of 14,400 mrem from non-SRS sources of ionizing radiation present in the environment. SRS activation product aqueous releases between 1954 and 1994 resulted in a total dose of 54 mrem to the offsite maximally exposed individual. The impact of SRS activation product releases on offsite populations also has been evaluated.

  1. An active vision system for multitarget surveillance in dynamic environments.

    PubMed

    Bakhtari, Ardevan; Benhabib, Beno

    2007-02-01

    This paper presents a novel agent-based method for the dynamic coordinated selection and positioning of active-vision cameras for the simultaneous surveillance of multiple objects-of-interest as they travel through a cluttered environment with a-priori unknown trajectories. The proposed system dynamically adjusts not only the orientation but also the position of the cameras in order to maximize the system's performance by avoiding occlusions and acquiring images with preferred viewing angles. Sensor selection and positioning are accomplished through an agent-based approach. The proposed sensing-system reconfiguration strategy has been verified via simulations and implemented on an experimental prototype setup for automated facial recognition. Both simulations and experimental analyses have shown that the use of dynamic sensors along with an effective online dispatching strategy may tangibly improve the surveillance performance of a sensing system.

  2. Dynamical behaviors of a plate activated by an induction motor

    NASA Astrophysics Data System (ADS)

    Tcheutchoua Fossi, D. O.; Woafo, P.

    2010-08-01

    Dynamics and chaotification of a system consisting of an induction motor activating a mobile plate (with variable contents) fixed to a spring are studied. The dynamical model of the device is presented and the electromechanical equations are formulated. The oscillations of the plate are analyzed through variations of the following reliable control parameters: phase voltage supply of the motor, frequency of the external source and mass of the plate. The dynamics of the system near the fundamental resonance region presents jump phenomenon. Mapping of the control parameters planes in terms of types of motion reveals period- n motion, quasi-periodicity and chaos. Anti-control of chaos of the induction motor is also obtained using the field-oriented control associated to the time delay feedback control.

  3. Dynamics and Control of a Quadrotor with Active Geometric Morphing

    NASA Astrophysics Data System (ADS)

    Wallace, Dustin A.

    Quadrotors are manufactured in a wide variety of shapes, sizes, and performance levels to fulfill a multitude of roles. Robodub Inc. has patented a morphing quadrotor which will allow active reconfiguration between various shapes for performance optimization across a wider spectrum of roles. The dynamics of the system are studied and modeled using Newtonian Mechanics. Controls are developed and simulated using both Linear Quadratic and Numerical Nonlinear Optimal control for a symmetric simplificiation of the system dynamics. Various unique vehicle capabilities are investigated, including novel single-throttle flight control using symmetric geometric morphing, as well as recovery from motor loss by reconfiguring into a trirotor configuration. The system dynamics were found to be complex and highly nonlinear. All attempted control strategies resulted in controllability, suggesting further research into each may lead to multiple viable control strategies for a physical prototype.

  4. Quantifying the density and utilization of active sites in non-precious metal oxygen electroreduction catalysts

    PubMed Central

    Sahraie, Nastaran Ranjbar; Kramm, Ulrike I.; Steinberg, Julian; Zhang, Yuanjian; Thomas, Arne; Reier, Tobias; Paraknowitsch, Jens-Peter; Strasser, Peter

    2015-01-01

    Carbon materials doped with transition metal and nitrogen are highly active, non-precious metal catalysts for the electrochemical conversion of molecular oxygen in fuel cells, metal air batteries, and electrolytic processes. However, accurate measurement of their intrinsic turn-over frequency and active-site density based on metal centres in bulk and surface has remained difficult to date, which has hampered a more rational catalyst design. Here we report a successful quantification of bulk and surface-based active-site density and associated turn-over frequency values of mono- and bimetallic Fe/N-doped carbons using a combination of chemisorption, desorption and 57Fe Mössbauer spectroscopy techniques. Our general approach yields an experimental descriptor for the intrinsic activity and the active-site utilization, aiding in the catalyst development process and enabling a previously unachieved level of understanding of reactivity trends owing to a deconvolution of site density and intrinsic activity. PMID:26486465

  5. Divergent contributions of conserved active site residues to transcription by eukaryotic RNA polymerases I and II.

    PubMed

    Viktorovskaya, Olga V; Engel, Krysta L; French, Sarah L; Cui, Ping; Vandeventer, Paul J; Pavlovic, Emily M; Beyer, Ann L; Kaplan, Craig D; Schneider, David A

    2013-09-12

    Multisubunit RNA polymerases (msRNAPs) exhibit high sequence and structural homology, especially within their active sites, which is generally thought to result in msRNAP functional conservation. However, we show that mutations in the trigger loop (TL) in the largest subunit of RNA polymerase I (Pol I) yield phenotypes unexpected from studies of Pol II. For example, a well-characterized gain-of-function mutation in Pol II results in loss of function in Pol I (Pol II: rpb1- E1103G; Pol I: rpa190-E1224G). Studies of chimeric Pol II enzymes hosting Pol I or Pol III TLs suggest that consequences of mutations that alter TL dynamics are dictated by the greater enzymatic context and not solely the TL sequence. Although the rpa190-E1224G mutation diminishes polymerase activity, when combined with mutations that perturb Pol I catalysis, it enhances polymerase function, similar to the analogous Pol II mutation. These results suggest that Pol I and Pol II have different rate-limiting steps.

  6. Characterization of an Active Thermal Erosion Site, Caribou Creek, Alaska

    NASA Astrophysics Data System (ADS)

    Busey, R.; Bolton, W. R.; Cherry, J. E.; Hinzman, L. D.

    2013-12-01

    The goal of this project is to estimate volume loss of soil over time from this site, provide parameterizations on erodibility of ice rich permafrost and serve as a baseline for future landscape evolution simulations. Located in the zone of discontinuous permafrost, the interior region of Alaska (USA) is home to a large quantity of warm, unstable permafrost that is both high in ice content and has soil temperatures near the freezing point. Much of this permafrost maintains a frozen state despite the general warming air temperature trend in the region due to the presence of a thick insulating organic mat and a dense root network in the upper sub-surface of the soil column. At a rapidly evolving thermo-erosion site, located within the Caribou-Poker Creeks Research Watershed (part of the Bonanza Creek LTER) near Chatanika, Alaska (N65.140, W147.570), the protective organic layer and associated plants were disturbed by an adjacent traditional use trail and the shifting of a groundwater spring. These triggers have led to rapid geomorphological change on the landscape as the soil thaws and sediment is transported into the creek at the valley bottom. Since 2006 (approximately the time of initiation), the thermal erosion has grown to 170 meters length, 3 meters max depth, and 15 meters maximum width. This research combines several data sets: DGPS survey, imagery from an extremely low altitude pole-based remote sensing (3 to 5 meters above ground level), and imagery from an Unmanned Aerial System (UAS) at about 60m altitude.

  7. Molecular Basis for Enzymatic Sulfite Oxidation -- HOW THREE CONSERVED ACTIVE SITE RESIDUES SHAPE ENZYME ACTIVITY

    SciTech Connect

    Bailey, Susan; Rapson, Trevor; Johnson-Winters, Kayunta; Astashkin, Andrei; Enemark, John; Kappler, Ulrike

    2008-11-10

    Sulfite dehydrogenases (SDHs) catalyze the oxidation and detoxification of sulfite to sulfate, a reaction critical to all forms of life. Sulfite-oxidizing enzymes contain three conserved active site amino acids (Arg-55, His-57, and Tyr-236) that are crucial for catalytic competency. Here we have studied the kinetic and structural effects of two novel and one previously reported substitution (R55M, H57A, Y236F) in these residues on SDH catalysis. Both Arg-55 and His-57 were found to have key roles in substrate binding. An R55M substitution increased Km(sulfite)(app) by 2-3 orders of magnitude, whereas His-57 was required for maintaining a high substrate affinity at low pH when the imidazole ring is fully protonated. This effect may be mediated by interactions of His-57 with Arg-55 that stabilize the position of the Arg-55 side chain or, alternatively, may reflect changes in the protonation state of sulfite. Unlike what is seen for SDHWT and SDHY236F, the catalytic turnover rates of SDHR55M and SDHH57A are relatively insensitive to pH (~;;60 and 200 s-1, respectively). On the structural level, striking kinetic effects appeared to correlate with disorder (in SDHH57A and SDHY236F) or absence of Arg-55 (SDHR55M), suggesting that Arg-55 and the hydrogen bonding interactions it engages in are crucial for substrate binding and catalysis. The structure of SDHR55M has sulfate bound at the active site, a fact that coincides with a significant increase in the inhibitory effect of sulfate in SDHR55M. Thus, Arg-55 also appears to be involved in enabling discrimination between the substrate and product in SDH.

  8. A rapid and direct method for the determination of active site accessibility in proteins based on ESI-MS and active site titrations.

    PubMed

    O'Farrell, Norah; Kreiner, Michaela; Moore, Barry D; Parker, Marie-Claire

    2006-11-01

    We have developed an electrospray ionisation mass spectrometry (ESI-MS) technique that can be applied to rapidly determine the number of intact active sites in proteins. The methodology relies on inhibiting the protein with an active-site irreversible inhibitor and then using ESI-MS to determine the extent of inhibition. We have applied this methodology to a test system: a serine protease, subtilisin Carlsberg, and monitored the extent of inhibition by phenylmethylsulfonyl fluoride (PMSF), an irreversible serine hydrolase inhibitor as a function of the changes in immobilisation and hydration conditions. Two types of enzyme preparation were investigated, lyophilised enzymes and protein-coated microcrystals (PCMC).

  9. Cyanide does more to inhibit heme enzymes, than merely serving as an active-site ligand.

    PubMed

    Parashar, Abhinav; Venkatachalam, Avanthika; Gideon, Daniel Andrew; Manoj, Kelath Murali

    2014-12-12

    The toxicity of cyanide is hitherto attributed to its ability to bind to heme proteins' active site and thereby inhibit their activity. It is shown herein that the long-held interpretation is inadequate to explain several observations in heme-enzyme reaction systems. Generation of cyanide-based diffusible radicals in heme-enzyme reaction milieu could shunt electron transfers (by non-active site processes), and thus be detrimental to the efficiency of oxidative outcomes.

  10. Marine Biology Field Trip Sites. Ocean Related Curriculum Activities.

    ERIC Educational Resources Information Center

    Pauls, John

    The ocean affects all of our lives. Therefore, awareness of and information about the interconnections between humans and oceans are prerequisites to making sound decisions for the future. Project ORCA (Ocean Related Curriculum Activities) has developed interdisciplinary curriculum materials designed to meet the needs of students and teachers…

  11. ATPase active-site electrostatic interactions control the global conformation of the 100 kDa SecA translocase.

    PubMed

    Kim, Dorothy M; Zheng, Haiyan; Huang, Yuanpeng J; Montelione, Gaetano T; Hunt, John F

    2013-02-27

    SecA is an intensively studied mechanoenzyme that uses ATP hydrolysis to drive processive extrusion of secreted proteins through a protein-conducting channel in the cytoplasmic membrane of eubacteria. The ATPase motor of SecA is strongly homologous to that in DEAD-box RNA helicases. It remains unclear how local chemical events in its ATPase active site control the overall conformation of an ~100 kDa multidomain enzyme and drive protein transport. In this paper, we use biophysical methods to establish that a single electrostatic charge in the ATPase active site controls the global conformation of SecA. The enzyme undergoes an ATP-modulated endothermic conformational transition (ECT) believed to involve similar structural mechanics to the protein transport reaction. We have characterized the effects of an isosteric glutamate-to-glutamine mutation in the catalytic base, a mutation which mimics the immediate electrostatic consequences of ATP hydrolysis in the active site. Calorimetric studies demonstrate that this mutation facilitates the ECT in Escherichia coli SecA and triggers it completely in Bacillus subtilis SecA. Consistent with the substantial increase in entropy observed in the course of the ECT, hydrogen-deuterium exchange mass spectrometry demonstrates that it increases protein backbone dynamics in domain-domain interfaces at remote locations from the ATPase active site. The catalytic glutamate is one of ~250 charged amino acids in SecA, and yet neutralization of its side chain charge is sufficient to trigger a global order-disorder transition in this 100 kDa enzyme. The intricate network of structural interactions mediating this effect couples local electrostatic changes during ATP hydrolysis to global conformational and dynamic changes in SecA. This network forms the foundation of the allosteric mechanochemistry that efficiently harnesses the chemical energy stored in ATP to drive complex mechanical processes. PMID:23167435

  12. Dynamics of a membrane interacting with an active wall.

    PubMed

    Yasuda, Kento; Komura, Shigeyuki; Okamoto, Ryuichi

    2016-05-01

    Active motions of a biological membrane can be induced by nonthermal fluctuations that occur in the outer environment of the membrane. We discuss the dynamics of a membrane interacting hydrodynamically with an active wall that exerts random velocities on the ambient fluid. Solving the hydrodynamic equations of a bound membrane, we first derive a dynamic equation for the membrane fluctuation amplitude in the presence of different types of walls. Membrane two-point correlation functions are calculated for three different cases: (i) a static wall, (ii) an active wall, and (iii) an active wall with an intrinsic time scale. We focus on the mean squared displacement (MSD) of a tagged membrane describing the Brownian motion of a membrane segment. For the static wall case, there are two asymptotic regimes of MSD (∼t^{2/3} and ∼t^{1/3}) when the hydrodynamic decay rate changes monotonically. In the case of an active wall, the MSD grows linearly in time (∼t) in the early stage, which is unusual for a membrane segment. This linear-growth region of the MSD is further extended when the active wall has a finite intrinsic time scale. PMID:27300924

  13. Crosstalk between circadian rhythmicity, mitochondrial dynamics and macrophage bactericidal activity

    PubMed Central

    Oliva-Ramírez, Jacqueline; Moreno-Altamirano, María Maximina B; Pineda-Olvera, Benjamín; Cauich-Sánchez, Patricia; Sánchez-García, F Javier

    2014-01-01

    Biological functions show rhythmic fluctuations with 24-hr periodicity regulated by circadian proteins encoded by the so-called ‘clock’ genes. The absence or deregulation of circadian proteins in mice leads to metabolic disorders and in vitro models have shown that the synthesis of pro-inflammatory cytokines by macrophages follows a circadian rhythm so showing a link between circadian rhythmicity, metabolism and immunity. Recent evidence reveals that mitochondrial shape, position and size, collectively referred to as mitochondrial dynamics, are related to both cell metabolism and immune function. However, studies addressing the simultaneous crosstalk between circadian rhythm, mitochondrial dynamics and cell immune function are scarce. Here, by using an in vitro model of synchronized murine peritoneal macrophages, we present evidence that the mitochondrial dynamics and the mitochondrial membrane potential (Δψm) follow a circadian rhythmic pattern. In addition, it is shown that the fusion of mitochondria along with high Δψm, indicative of high mitochondrial activity, precede the highest phagocytic and bactericidal activity of macrophages on Salmonella typhimurium. Taken together, our results suggest a timely coordination between circadian rhythmicity, mitochondrial dynamics, and the bactericidal capacity of macrophages. PMID:24903615

  14. Active site proton delivery and the lyase activity of human CYP17A1

    SciTech Connect

    Khatri, Yogan; Gregory, Michael C.; Grinkova, Yelena V.; Denisov, Ilia G.; Sligar, Stephen G.

    2014-01-03

    equivalents and protons are funneled into non-productive pathways. This is similar to previous work with other P450 catalyzed hydroxylation. However, catalysis of carbon–carbon bond scission by the T306A mutant was largely unimpeded by disruption of the CYP17A1 acid-alcohol pair. The unique response of CYP17A1 lyase activity to mutation of Thr306 is consistent with a reactive intermediate formed independently of proton delivery in the active site, and supports involvement of a nucleophilic peroxo-anion rather than the traditional Compound I in catalysis.

  15. Time and space in the middle paleolithic: Spatial structure and occupation dynamics of seven open-air sites.

    PubMed

    Clark, Amy E

    2016-05-01

    The spatial structure of archeological sites can help reconstruct the settlement dynamics of hunter-gatherers by providing information on the number and length of occupations. This study seeks to access this information through a comparison of seven sites. These sites are open-air and were all excavated over large spatial areas, up to 2,000 m(2) , and are therefore ideal for spatial analysis, which was done using two complementary methods, lithic refitting and density zones. Both methods were assessed statistically using confidence intervals. The statistically significant results from each site were then compiled to evaluate trends that occur across the seven sites. These results were used to assess the "spatial consistency" of each assemblage and, through that, the number and duration of occupations. This study demonstrates that spatial analysis can be a powerful tool in research on occupation dynamics and can help disentangle the many occupations that often make up an archeological assemblage.

  16. A "Kane's Dynamics" Model for the Active Rack Isolation System

    NASA Technical Reports Server (NTRS)

    Hampton, R. D.; Beech, G. S.; Rao, N. N. S.; Rupert, J. K.; Kim, Y. K.

    2001-01-01

    Many microgravity space science experiments require vibratory acceleration levels unachievable without active isolation. The Boeing Corporation's Active Rack Isolation System (ARIS) employs a novel combination of magnetic actuation and mechanical linkages to address these isolation requirements on the International Space Station (ISS). ARIS provides isolation at the rack (International Standard Payload Rack (ISPR)) level. Effective model-based vibration isolation requires: (1) an appropriate isolation device, (2) an adequate dynamic (i.e., mathematical) model of that isolator, and (3) a suitable, corresponding controller. ARIS provides the ISS response to the first requirement. This paper presents one response to the second, in a state space framework intended to facilitate an optimal-controls approach to the third. The authors use "Kane's Dynamics" to develop a state-space, analytical (algebraic) set of linearized equations of motion for ARIS.

  17. A "Kanes's Dynamics" Model for the Active Rack Isolation System

    NASA Technical Reports Server (NTRS)

    Hampton, R. David; Beech, Geoffrey

    1999-01-01

    Many microgravity space-science experiments require vibratory acceleration levels unachievable without active isolation. The Boeing Corporation's Active Rack Isolation System (ARIS) employs a novel combination of magnetic actuation and mechanical linkages, to address these isolation requirements on the International Space Station (ISS). ARIS provides isolation at the rack (international Standard Payload Rack, or ISPR) level. Effective model-based vibration isolation requires (1) an appropriate isolation device, (2) an adequate dynamic (i.e., mathematical) model of that isolator, and (3) a suitable, corresponding controller. ARIS provides the ISS response to the first requirement. This paper presents one response to the second, in a state-space framework intended to facilitate an optimal-controls approach to the third. The authors use "Kane's Dynamics" to develop an state-space, analytical (algebraic) set of linearized equations of motion for ARIS.

  18. Dynamic active constraints for hyper-redundant flexible robots.

    PubMed

    Kwok, Ka-Wai; Mylonas, George P; Sun, Loi Wah; Lerotic, Mirna; Clark, James; Athanasiou, Thanos; Darzi, Ara; Yang, Guang-Zhong

    2009-01-01

    In robot-assisted procedures, the surgeon's ability can be enhanced by navigation guidance through the use of virtual fixtures or active constraints. This paper presents a real-time modeling scheme for dynamic active constraints with fast and simple mesh adaptation under cardiac deformation and changes in anatomic structure. A smooth tubular pathway is constructed which provides assistance for a flexible hyper-redundant robot to circumnavigate the heart with the aim of undertaking bilateral pulmonary vein isolation as part of a modified maze procedure for the treatment of debilitating arrhythmia and atrial fibrillation. In contrast to existing approaches, the method incorporates detailed geometrical constraints with explicit manipulation margins of the forbidden region for an entire articulated surgical instrument, rather than just the end-effector itself. Detailed experimental validation is conducted to demonstrate the speed and accuracy of the instrument navigation with and without the use of the proposed dynamic constraints.

  19. Identification of inhibitors against the potential ligandable sites in the active cholera toxin.

    PubMed

    Gangopadhyay, Aditi; Datta, Abhijit

    2015-04-01

    The active cholera toxin responsible for the massive loss of water and ions in cholera patients via its ADP ribosylation activity is a heterodimer of the A1 subunit of the bacterial holotoxin and the human cytosolic ARF6 (ADP Ribosylation Factor 6). The active toxin is a potential target for the design of inhibitors against cholera. In this study we identified the potential ligandable sites of the active cholera toxin which can serve as binding sites for drug-like molecules. By employing an energy-based approach to identify ligand binding sites, and comparison with the results of computational solvent mapping, we identified two potential ligandable sites in the active toxin which can be targeted during structure-based drug design against cholera. Based on the probe affinities of the identified ligandable regions, docking-based virtual screening was employed to identify probable inhibitors against these sites. Several indole-based alkaloids and phosphates showed strong interactions to the important residues of the ligandable region at the A1 active site. On the other hand, 26 top scoring hits were identified against the ligandable region at the A1 ARF6 interface which showed strong hydrogen bonding interactions, including guanidines, phosphates, Leucopterin and Aristolochic acid VIa. This study has important implications in the application of hybrid structure-based and ligand-based methods against the identified ligandable sites using the identified inhibitors as reference ligands, for drug design against the active cholera toxin.

  20. Encroachment of Human Activity on Sea Turtle Nesting Sites

    NASA Astrophysics Data System (ADS)

    Ziskin, D.; Aubrecht, C.; Elvidge, C.; Tuttle, B.; Baugh, K.; Ghosh, T.

    2008-12-01

    The encroachment of anthropogenic lighting on sea turtle nesting sites poses a serious threat to the survival of these animals [Nicholas, 2001]. This danger is quantified by combining two established data sets. The first is the Nighttime Lights data produced by the NOAA National Geophysical Data Center [Elvidge et al., 1997]. The second is the Marine Turtle Database produced by the World Conservation Monitoring Centre (WCMC). The technique used to quantify the threat of encroachment is an adaptation of the method described in Aubrecht et al. [2008], which analyzes the stress on coral reef systems by proximity to nighttime lights near the shore. Nighttime lights near beaches have both a direct impact on turtle reproductive success since they disorient hatchlings when they mistake land-based lights for the sky-lit surf [Lorne and Salmon, 2007] and the lights are also a proxy for other anthropogenic threats. The identification of turtle nesting sites with high rates of encroachment will hopefully steer conservation efforts to mitigate their effects [Witherington, 1999]. Aubrecht, C, CD Elvidge, T Longcore, C Rich, J Safran, A Strong, M Eakin, KE Baugh, BT Tuttle, AT Howard, EH Erwin, 2008, A global inventory of coral reef stressors based on satellite observed nighttime lights, Geocarto International, London, England: Taylor and Francis. In press. Elvidge, CD, KE Baugh, EA Kihn, HW Kroehl, ER Davis, 1997, Mapping City Lights with Nighttime Data from the DMSP Operational Linescan System, Photogrammatic Engineering and Remote Sensing, 63:6, pp. 727-734. Lorne, JK, M Salmon, 2007, Effects of exposure to artificial lighting on orientation of hatchling sea turtles on the beach and in the ocean, Endangered Species Research, Vol. 3: 23-30. Nicholas, M, 2001, Light Pollution and Marine Turtle Hatchlings: The Straw that Breaks the Camel's Back?, George Wright Forum, 18:4, p77-82. Witherington, BE, 1999, Reducing Threats To Nesting Habitat, Research and Management Techniques for

  1. Dynamics of two-site Fermi-Hubbard and Bose-Hubbard systems

    SciTech Connect

    Ziegler, K.

    2010-03-15

    This paper analyzes dynamical properties of small Fermi-Hubbard and Bose-Hubbard systems, focusing on the structure of the underlying Hilbert space. We evaluate time-dependent quantities such as the return probability to the initial state and the spin imbalance of spin-1/2 fermions. For the symmetric two-site Fermi-Hubbard model we find that the spin imbalance and the return probability are controlled by two and three frequencies, respectively. The spin imbalance and the return probability are identical for the asymmetric Falicov-Kimball limit and controlled by only one frequency. In general, the transition probabilities between the initial state and the energy eigenstates depend strongly on the particle-particle interaction. This is discussed for 'self-trapping' of spinless bosons in a double-well potential. We observe that the available Hilbert space is reduced significantly by strong interaction.

  2. Seasonal Carbon Dynamics on Selected Fen Peatland Sites in NE-Germany

    NASA Astrophysics Data System (ADS)

    Giebels, Michael; Beyer, Madlen; Augustin, Jürgen; Minke, Merten; Juszczak, Radoszlav; Serba, Tomasz

    2010-05-01

    In Germany more than 99 % of fens have lost their carbon and nutrient sink function due to heavy drainage and agricultural land use especially during the last decades and thus resulted in compression and heavy peat loss (CHARMAN 2002; JOOSTEN & CLARKE 2002; SUCCOW & JOOSTEN 2001; AUGUSTIN et al. 1996; KUNTZE 1993). Therefore fen peatlands play an important part (4-5 %) in the national anthropogenic trace gas budget. But only a small part of drained and agricultural used fens in NE Germany can be restored. Knowledge of the influence of land use to trace gas exchange is important for mitigation of the climate impact of the anthropogenic peatland use. We study carbon exchanges of several fen peatland use areas between soil and atmosphere at different sites in NE-Germany. Our research covers peatlands of supposed strongly climate forcing land use (cornfield and intensive pasture) and of probably less forcing, alternative types (meadow and extensive pasture) as well as rewetted (formerly drained) areas and near-natural sites like a low-degraded fen and a wetted alder woodland. We measured trace gas fluxes with manual and automatic chambers in periodic routines since spring 2007. The used chamber technique bases on DROESLER (2005). In total we now do research at 22 sites situated in 5 different locations covering agricultural, varying states of rewetted and near-natural treatments. We present results of at least 2 years of measurements and show significant differences in their annual carbon balances depending on the genesis of the observed sites and the seasonal dynamics. Crosswise comparison of different site treatments combined with the seasonal environmental observations give good hints for the identification of main flux driving parameters. That is that a reduced intensity in land use as a supposed mitigating treatment did not show the expected effect, though a normal meadow treatment surprisingly resulted in the lowest CO2 balances in both years. For implementing a

  3. Coarsening dynamics of binary liquids with active rotation.

    PubMed

    Sabrina, Syeda; Spellings, Matthew; Glotzer, Sharon C; Bishop, Kyle J M

    2015-11-21

    Active matter comprised of many self-driven units can exhibit emergent collective behaviors such as pattern formation and phase separation in both biological (e.g., mussel beds) and synthetic (e.g., colloidal swimmers) systems. While these behaviors are increasingly well understood for ensembles of linearly self-propelled "particles", less is known about the collective behaviors of active rotating particles where energy input at the particle level gives rise to rotational particle motion. A recent simulation study revealed that active rotation can induce phase separation in mixtures of counter-rotating particles in 2D. In contrast to that of linearly self-propelled particles, the phase separation of counter-rotating fluids is accompanied by steady convective flows that originate at the fluid-fluid interface. Here, we investigate the influence of these flows on the coarsening dynamics of actively rotating binary liquids using a phenomenological, hydrodynamic model that combines a Cahn-Hilliard equation for the fluid composition with a Navier-Stokes equation for the fluid velocity. The effect of active rotation is introduced though an additional force within the Navier-Stokes equations that arises due to gradients in the concentrations of clockwise and counter-clockwise rotating particles. Depending on the strength of active rotation and that of frictional interactions with the stationary surroundings, we observe and explain new dynamical behaviors such as "active coarsening" via self-generated flows as well as the emergence of self-propelled "vortex doublets". We confirm that many of the qualitative behaviors identified by the continuum model can also be found in discrete, particle-based simulations of actively rotating liquids. Our results highlight further opportunities for achieving complex dissipative structures in active materials subject to distributed actuation. PMID:26345231

  4. Turbidite Paleoseismology: Site Selection, Physiography, Sediment Supply, Current Dynamics and Temporal Considerations as Applied in Cascadia and Elsewhere

    NASA Astrophysics Data System (ADS)

    Goldfinger, C.; Hamilton, T. S.; Beeson, J.; Galer, S.; Nelson, C. H.; Morey, A. E.; Udrekh, U.

    2014-12-01

    Turbidite paleoseismology requires careful consideration of site context, temporal interval of interest, sediment supply, and the flow dynamics before interpretations can be drawn. These factors are predicated on precise navigation so that the context of the core within modern bathymetric, sub-bottom, sidescan, and backscatter data are known. In Cascadia, numerous channel systems exist and cover a range of time intervals since the Early Pleistocene. During high stands, many of these systems are relict, with limited terrigenous sediment supply. Holocene paleoseismic records may depend on recycled materials from failure of local slopes to supply channels, slope basins, or fans. Local failures may serve to supply sediment at any point along a canyon system under expected shaking levels of ~ 1.0 g with or without recent sediment recharge. Recharge by active terrigenous sedimentation is apparently not required in Cascadia or Sumatra, where site locations, without this recharge possibility have excellent records correlable to other paleoseismic sites. By comparison to Pleistocene fan-building currents, Holocene currents are weak, rendering most areas of fan systems inactive. Core and backscatter data show the Astoria and Nitinat Fans have little Holocene activity outside the main channels. Pleistocene channels are crosscut by active Holocene incisions and levees, restricting their role as depocenters. In the main channels, the most recent currents are largely confined closely within their levees. Recent proposals for alternate Holocene pathways in Cascadia attempt to integrate data from inactive fans, pose implausible pathways over the top of the growing accretionary wedge, or use other inactive channels. Resolution of observations is also critical and simple visual core logging is inadequate when compared to modern CT data. Thus for Holocene paleoseismology, cores must be collected from within main channels or near enough to local slopes (1-2 km) to receive

  5. Ab initio molecular dynamics studies on HIV-1 reverse transcriptase triphosphate binding site: implications for nucleoside-analog drug resistance.

    PubMed

    Alber, F; Carloni, P

    2000-12-01

    Quantum-chemical methods are used to shed light on the functional role of residues involved in the resistance of HIV-1 reverse transcriptase against nucleoside-analog drugs. Ab initio molecular dynamics simulations are carried out for models representing the adduct between the triphosphate substrate and the nucleoside binding site. The triphosphate is considered either deprotonated or protonated at the gamma-position. Although the protonated form already experiences large rearrangements in the ps time scale, the fully deprotonated state exhibits a previously unrecognized low-barrier hydrogen bond between Lys65 and gamma-phosphate. Absence of this interaction in Lys65-->Arg HIV-1 RT might play a prominent role in the resistance of this mutant for nucleoside analogs (Gu Z et al., 1994b, Antimicrob Agents Chemother 38:275-281; Zhang D et al., 1994, Antimicrob Agents Chemother 38:282-287). Water molecules present in the active site, not detected in the X-ray structure, form a complex H-bond network. Among these waters, one may be crucial for substrate recognition as it bridges Gln151 and Arg72 with the beta-phosphate. Absence of this stabilizing interaction in Gln151-->Met HIV-1 RT mutant may be a key factor for the known drug resistance of this mutant toward dideoxy-type drugs and AZT (Shirasaka T et al., 1995, Proc Natl Acad Sci USA 92:2398-2402: Iversen AK et al., 1996, J Virol 70:1086-1090).

  6. Dynamics of a deformable active particle under shear flow.

    PubMed

    Tarama, Mitsusuke; Menzel, Andreas M; ten Hagen, Borge; Wittkowski, Raphael; Ohta, Takao; Löwen, Hartmut

    2013-09-14

    The motion of a deformable active particle in linear shear flow is explored theoretically. Based on symmetry considerations, we propose coupled nonlinear dynamical equations for the particle position, velocity, deformation, and rotation. In our model, both, passive rotations induced by the shear flow as well as active spinning motions, are taken into account. Our equations reduce to known models in the two limits of vanishing shear flow and vanishing particle deformability. For varied shear rate and particle propulsion speed, we solve the equations numerically in two spatial dimensions and obtain a manifold of different dynamical modes including active straight motion, periodic motions, motions on undulated cycloids, winding motions, as well as quasi-periodic and chaotic motions induced at high shear rates. The types of motion are distinguished by different characteristics in the real-space trajectories and in the dynamical behavior of the particle orientation and its deformation. Our predictions can be verified in experiments on self-propelled droplets exposed to a linear shear flow.

  7. Global segregation of cortical activity and metastable dynamics

    PubMed Central

    Stratton, Peter; Wiles, Janet

    2015-01-01

    Cortical activity exhibits persistent metastable dynamics. Assemblies of neurons transiently couple (integrate) and decouple (segregate) at multiple spatiotemporal scales; both integration and segregation are required to support metastability. Integration of distant brain regions can be achieved through long range excitatory projections, but the mechanism supporting long range segregation is not clear. We argue that the thalamocortical matrix connections, which project diffusely from the thalamus to the cortex and have long been thought to support cortical gain control, play an equally-important role in cortical segregation. We present a computational model of the diffuse thalamocortical loop, called the competitive cross-coupling (CXC) spiking network. Simulations of the model show how different levels of tonic input from the brainstem to the thalamus could control dynamical complexity in the cortex, directing transitions between sleep, wakefulness and high attention or vigilance. The model also explains how mutually-exclusive activity could arise across large portions of the cortex, such as between the default-mode and task-positive networks. It is robust to noise but does not require noise to autonomously generate metastability. We conclude that the long range segregation observed in brain activity and required for global metastable dynamics could be provided by the thalamocortical matrix, and is strongly modulated by brainstem input to the thalamus. PMID:26379514

  8. Global segregation of cortical activity and metastable dynamics.

    PubMed

    Stratton, Peter; Wiles, Janet

    2015-01-01

    Cortical activity exhibits persistent metastable dynamics. Assemblies of neurons transiently couple (integrate) and decouple (segregate) at multiple spatiotemporal scales; both integration and segregation are required to support metastability. Integration of distant brain regions can be achieved through long range excitatory projections, but the mechanism supporting long range segregation is not clear. We argue that the thalamocortical matrix connections, which project diffusely from the thalamus to the cortex and have long been thought to support cortical gain control, play an equally-important role in cortical segregation. We present a computational model of the diffuse thalamocortical loop, called the competitive cross-coupling (CXC) spiking network. Simulations of the model show how different levels of tonic input from the brainstem to the thalamus could control dynamical complexity in the cortex, directing transitions between sleep, wakefulness and high attention or vigilance. The model also explains how mutually-exclusive activity could arise across large portions of the cortex, such as between the default-mode and task-positive networks. It is robust to noise but does not require noise to autonomously generate metastability. We conclude that the long range segregation observed in brain activity and required for global metastable dynamics could be provided by the thalamocortical matrix, and is strongly modulated by brainstem input to the thalamus.

  9. Temporal dynamics of a homeostatic pathway controlling neural network activity

    PubMed Central

    Bateup, Helen S.; Denefrio, Cassandra L.; Johnson, Caroline A.; Saulnier, Jessica L.; Sabatini, Bernardo L.

    2013-01-01

    Neurons use a variety of mechanisms to homeostatically regulate neural network activity in order to maintain firing in a bounded range. One such process involves the bi-directional modulation of excitatory synaptic drive in response to chronic changes in network activity. Down-scaling of excitatory synapses in response to high activity requires Arc-dependent endocytosis of glutamate receptors. However, the temporal dynamics and signaling pathways regulating Arc during homeostatic plasticity are not well understood. Here we determine the relative contribution of transcriptional and translational control in the regulation of Arc, the signaling pathways responsible for the activity-dependent production of Arc, and the time course of these signaling events as they relate to the homeostatic adjustment of network activity in hippocampal neurons. We find that an ERK1/2-dependent transcriptional pathway active within 1–2 h of up-regulated network activity induces Arc leading to a restoration of network spiking rates within 12 h. Under basal and low activity conditions, specialized mechanisms are in place to rapidly degrade Arc mRNA and protein such that they have half-lives of less than 1 h. In addition, we find that while mTOR signaling is regulated by network activity on a similar time scale, mTOR-dependent translational control is not a major regulator of Arc production or degradation suggesting that the signaling pathways underlying homeostatic plasticity are distinct from those mediating synapse-specific forms of synaptic depression. PMID:24065881

  10. Active gels: dynamics of patterning and self-organization

    NASA Astrophysics Data System (ADS)

    Backouche, F.; Haviv, L.; Groswasser, D.; Bernheim-Groswasser, A.

    2006-12-01

    The actin cytoskeleton is an active gel which constantly remodels during cellular processes such as motility and division. Myosin II molecular motors are involved in this active remodeling process and therefore control the dynamic self-organization of cytoskeletal structures. Due to the complexity of in vivo systems, it is hard to investigate the role of myosin II in the reorganization process which determines the resulting cytoskeletal structures. Here we use an in vitro model system to show that myosin II actively reorganizes actin into a variety of mesoscopic patterns, but only in the presence of bundling proteins. We find that the nature of the reorganization process is complex, exhibiting patterns and dynamical phenomena not predicted by current theoretical models and not observed in corresponding passive systems (excluding motors). This system generates active networks, asters and even rings depending on motor and bundling protein concentrations. Furthermore, the motors generate the formation of the patterns, but above a critical concentration they can also disassemble them and even totally prevent the polymerization and bundling of actin filaments. These results may suggest that tuning the assembly and disassembly of cytoskeletal structures can be obtained by tuning the local myosin II concentration/activity.

  11. Dynamics and control of substrate inhibition in activated sludge

    SciTech Connect

    Allsop, P.J.; Moo-Young, M.; Sullivan, G.R. )

    1990-01-01

    The activated sludge wastewater treatment process predominantly used in the chemical and steel industries was reviewed to determine the dynamics and control of activated sludge systems treating inhibitory wastes. While this process has the capability to degrade a variety of toxic or inhibitory wastes, the underlying mechanisms are not clear. A variety of issues exist requiring further study: (1) the role of various microorganisms in waste removal and system stability, (2) the mechanisms of inhibitory action at both the level of the primary consumer and at the level of the whole process, (3) the suitability of phenol as a model inhibitory substrate, (4) the appropriateness of using pure culture, CSTR results obtained at relatively high specific growth rates to predict the response of activated sludge systems, (5) the rationalization of microbiological predictions for oligotrophic systems with observations in activated sludge systems, and (6) the development of appropriate monitoring tools for detecting process instabilities. 265 refs., 8 figs., 2 tabs.

  12. Exciton dynamics in a site-controlled quantum dot coupled to a photonic crystal cavity

    SciTech Connect

    Jarlov, C. Lyasota, A.; Ferrier, L.; Gallo, P.; Dwir, B.; Rudra, A.; Kapon, E.

    2015-11-09

    Exciton and cavity mode (CM) dynamics in site-controlled pyramidal quantum dots (QDs), integrated with linear photonic crystal membrane cavities, are investigated for a range of temperatures and photo-excitation power levels. The absence of spurious multi-excitonic effects, normally observed in similar structures based on self-assembled QDs, permits the observation of effects intrinsic to two-level systems embedded in a solid state matrix and interacting with optical cavity modes. The coupled exciton and CM dynamics follow the same trend, indicating that the CM is fed only by the exciton transition. The Purcell reduction of the QD and CM decay times is reproduced well by a theoretical model that includes exciton linewidth broadening and temperature dependent non-radiative processes, from which we extract a Purcell factor of 17 ± 5. For excitation powers above QD saturation, we show the influence of quantum wire barrier states at short delay time, and demonstrate the absence of multiexcitonic background emission.

  13. Bayesian inversions of a dynamic vegetation model in four European grassland sites

    NASA Astrophysics Data System (ADS)

    Minet, J.; Laloy, E.; Tychon, B.; François, L.

    2015-01-01

    Eddy covariance data from four European grassland sites are used to probabilistically invert the CARAIB dynamic vegetation model (DVM) with ten unknown parameters, using the DREAM(ZS) Markov chain Monte Carlo (MCMC) sampler. We compare model inversions considering both homoscedastic and heteroscedastic eddy covariance residual errors, with variances either fixed a~priori or jointly inferred with the model parameters. Agreements between measured and simulated data during calibration are comparable with previous studies, with root-mean-square error (RMSE) of simulated daily gross primary productivity (GPP), ecosystem respiration (RECO) and evapotranspiration (ET) ranging from 1.73 to 2.19 g C m-2 day-1, 1.04 to 1.56 g C m-2 day-1, and 0.50 to 1.28 mm day-1, respectively. In validation, mismatches between measured and simulated data are larger, but still with Nash-Sutcliffe efficiency scores above 0.5 for three out of the four sites. Although measurement errors associated with eddy covariance data are known to be heteroscedastic, we showed that assuming a classical linear heteroscedastic model of the residual errors in the inversion do not fully remove heteroscedasticity. Since the employed heteroscedastic error model allows for larger deviations between simulated and measured data as the magnitude of the measured data increases, this error model expectedly lead to poorer data fitting compared to inversions considering a constant variance of the residual errors. Furthermore, sampling the residual error variances along with model parameters results in overall similar model parameter posterior distributions as those obtained by fixing these variances beforehand, while slightly improving model performance. Despite the fact that the calibrated model is generally capable of fitting the data within measurement errors, systematic bias in the model simulations are observed. These are likely due to model inadequacies such as shortcomings in the photosynthesis modelling

  14. Rosetta lander Philae: Flight Dynamics analyses for landing site selection and post-landing operations

    NASA Astrophysics Data System (ADS)

    Jurado, Eric; Martin, Thierry; Canalias, Elisabet; Blazquez, Alejandro; Garmier, Romain; Ceolin, Thierry; Gaudon, Philippe; Delmas, Cedric; Biele, Jens; Ulamec, Stephan; Remetean, Emile; Torres, Alex; Laurent-Varin, Julien; Dolives, Benoit; Herique, Alain; Rogez, Yves; Kofman, Wlodek; Jorda, Laurent; Zakharov, Vladimir; Crifo, Jean-François; Rodionov, Alexander; Heinish, P.; Vincent, Jean-Baptiste

    2016-08-01

    On the 12th of November 2014, The Rosetta Lander Philae became the first spacecraft to softly land on a comet nucleus. Due to the double failure of the cold gas hold-down thruster and the anchoring harpoons that should have fixed Philae to the surface, it spent approximately two hours bouncing over the comet surface to finally come at rest one km away from its target site. Nevertheless it was operated during the 57 h of its First Science Sequence. The FSS, performed with the two batteries, should have been followed by the Long Term Science Sequence but Philae was in a place not well illuminated and fell into hibernation. Yet, thanks to reducing distance to the Sun and to seasonal effect, it woke up at end of April and on 13th of June it contacted Rosetta again. To achieve this successful landing, an intense preparation work had been carried out mainly between August and November 2014 to select the targeted landing site and define the final landing trajectory. After the landing, the data collected during on-comet operations have been used to assess the final position and orientation of Philae, and to prepare the wake-up. This paper addresses the Flight Dynamics studies done in the scope of this landing preparation from Lander side, in close cooperation with the team at ESA, responsible for Rosetta, as well as for the reconstruction of the bouncing trajectory and orientation of the Lander after touchdown.

  15. Site and bond-specific dynamics of reactions at the gas-liquid interface.

    PubMed

    Tesa-Serrate, Maria A; King, Kerry L; Paterson, Grant; Costen, Matthew L; McKendrick, Kenneth G

    2014-01-01

    The dynamics of the interfacial reactions of O((3)P) with the hydrocarbon liquids squalane (C30H62, 2,6,10,15,19,23-hexamethyltetracosane) and squalene (C30H50, trans-2,6,10,15,19,23-hexamethyltetracosa-2,6,10,14,18,22-hexaene) have been studied experimentally. Laser-induced fluorescence (LIF) was used to detect the nascent gas-phase OH products. The O((3)P) atoms are acutely sensitive to the chemical differences of the squalane and squalene surfaces. The larger exothermicity of abstraction from allylic C-H sites in squalene is reflected in markedly hotter OH rotational and vibrational distributions. There is a more modest increase in translational energy release. A larger fraction of the available energy is deposited in the liquid for squalene than for squalane, consistent with a more extensive geometry change on formation of the allylic radical co-product. Although the dominant reaction mechanism is direct, impulsive scattering, there is some evidence for OH being accommodated at both liquid surfaces, resulting in thermalised translation and rotational distributions. Despite the H-abstraction reaction being strongly favoured energetically for squalene, the yield of OH is substantially lower than for squalane. This is very likely due to competitive addition of O((3)P) to the unsaturated sites in squalene, implying that double bonds are extensively exposed at the liquid surface.

  16. Subcellular metal imaging identifies dynamic sites of Cu accumulation in Chlamydomonas

    SciTech Connect

    Hong-Hermesdorf, Anne; Miethke, Marcus; Gallaher, Sean D.; Kropat, Janette; Dodani, Sheel C.; Chan, Jefferson; Barupala, Dulmini; Domaille, Dylan W.; Shirasaki, Dyna I.; Loo, Joseph A.; Weber, Peter K.; Pett-Ridge, Jennifer; Stemmler, Timothy L.; Chang, Christopher J.; Merchant, Sabeeha S.

    2014-10-26

    Here we identified a Cu-accumulating structure with a dynamic role in intracellular Cu homeostasis. During Zn limitation, Chlamydomonas reinhardtii hyperaccumulates Cu, a process dependent on the nutritional Cu sensor CRR1, but it is functionally Cu deficient. Visualization of intracellular Cu revealed major Cu accumulation sites coincident with electron-dense structures that stained positive for low pH and polyphosphate, suggesting that they are lysosome-related organelles. Nano-secondary ion MS showed colocalization of Ca and Cu, and X-ray absorption spectroscopy was consistent with Cu+ accumulation in an ordered structure. Zn resupply restored Cu homeostasis concomitant with reduced abundance of these structures. Cu isotope labeling demonstrated that sequestered Cu+ became bioavailable for the synthesis of plastocyanin, and transcriptome profiling indicated that mobilized Cu became visible to CRR1. Cu trafficking to intracellular accumulation sites may be a strategy for preventing protein mismetallation during Zn deficiency and enabling efficient cuproprotein metallation or remetallation upon Zn resupply.

  17. Subcellular metal imaging identifies dynamic sites of Cu accumulation in Chlamydomonas

    DOE PAGES

    Hong-Hermesdorf, Anne; Miethke, Marcus; Gallaher, Sean D.; Kropat, Janette; Dodani, Sheel C.; Chan, Jefferson; Barupala, Dulmini; Domaille, Dylan W.; Shirasaki, Dyna I.; Loo, Joseph A.; et al

    2014-10-26

    Here we identified a Cu-accumulating structure with a dynamic role in intracellular Cu homeostasis. During Zn limitation, Chlamydomonas reinhardtii hyperaccumulates Cu, a process dependent on the nutritional Cu sensor CRR1, but it is functionally Cu deficient. Visualization of intracellular Cu revealed major Cu accumulation sites coincident with electron-dense structures that stained positive for low pH and polyphosphate, suggesting that they are lysosome-related organelles. Nano-secondary ion MS showed colocalization of Ca and Cu, and X-ray absorption spectroscopy was consistent with Cu+ accumulation in an ordered structure. Zn resupply restored Cu homeostasis concomitant with reduced abundance of these structures. Cu isotope labelingmore » demonstrated that sequestered Cu+ became bioavailable for the synthesis of plastocyanin, and transcriptome profiling indicated that mobilized Cu became visible to CRR1. Cu trafficking to intracellular accumulation sites may be a strategy for preventing protein mismetallation during Zn deficiency and enabling efficient cuproprotein metallation or remetallation upon Zn resupply.« less

  18. Preferential sites for InAsP/InP quantum wire nucleation using molecular dynamics

    NASA Astrophysics Data System (ADS)

    Nuñez-Moraleda, Bernardo; Pizarro, Joaquin; Guerrero, Elisa; Guerrero-Lebrero, Maria P.; Yáñez, Andres; Molina, Sergio Ignacio; Galindo, Pedro Luis

    2014-11-01

    In this paper, stress fields at the surface of the capping layer of self-assembled InAsP quantum wires grown on an InP (001) substrate have been determined from atomistic models using molecular dynamics and Stillinger-Weber potentials. To carry out these calculations, the quantum wire compositional distribution was extracted from previous works, where the As and P distributions were determined by electron energy loss spectroscopy and high-resolution aberration-corrected Z-contrast imaging. Preferential sites for the nucleation of wires on the surface of the capping layer were studied and compared with (i) previous simulations using finite element analysis to solve anisotropic elastic theory equations and (ii) experimentally measured locations of stacked wires. Preferential nucleation sites of stacked wires were determined by the maximum stress location at the MD model surface in good agreement with experimental results and those derived from finite element analysis. This indicates that MD simulations based on empirical potentials provide a suitable and flexible tool to study strain dependent atom processes.

  19. Modelling Transmission of Vector-Borne Pathogens Shows Complex Dynamics When Vector Feeding Sites Are Limited

    PubMed Central

    Kershenbaum, Arik; Stone, Lewi; Ostfeld, Richard S.; Blaustein, Leon

    2012-01-01

    The relationship between species richness and the prevalence of vector-borne disease has been widely studied with a range of outcomes. Increasing the number of host species for a pathogen may decrease infection prevalence (dilution effect), increase it (amplification), or have no effect. We derive a general model, and a specific implementation, which show that when the number of vector feeding sites on each host is limiting, the effects on pathogen dynamics of host population size are more complex than previously thought. The model examines vector-borne disease in the presence of different host species that are either competent or incompetent (i.e. that cannot transmit the pathogen to vectors) as reservoirs for the pathogen. With a single host species present, the basic reproduction ratio R0 is a non-monotonic function of the population size of host individuals (H), i.e. a value exists that maximises R0. Surprisingly, if a reduction in host population size may actually increase R0. Extending this model to a two-host species system, incompetent individuals from the second host species can alter the value of which may reverse the effect on pathogen prevalence of host population reduction. We argue that when vector-feeding sites on hosts are limiting, the net effect of increasing host diversity might not be correctly predicted using simple frequency-dependent epidemiological models. PMID:22590597

  20. Sub-cellular metal imaging identifies dynamic sites of Cu accumulation in Chlamydomonas

    PubMed Central

    Hong-Hermesdorf, Anne; Miethke, Marcus; Gallaher, Sean D; Kropat, Janette; Dodani, Sheel C; Chan, Jefferson; Barupala, Dulmini; Domaille, Dylan W; Shirasaki, Dyna I; Loo, Joseph A; Weber, Peter K; Pett-Ridge, Jennifer; Stemmler, Timothy L; Chang, Christopher J; Merchant, Sabeeha S

    2014-01-01

    We identified a Cu accumulating structure with a dynamic role in intracellular Cu homeostasis. During Zn limitation, Chlamydomonas reinhardtii hyperaccumulated Cu, dependent on the nutritional Cu sensor CRR1, but was functionally Cu-deficient. Visualization of intracellular Cu revealed major Cu accumulation sites coincident with electron-dense structures that stained positive for low pH and polyphosphate, suggesting that they are lysosome-related organelles. NanoSIMS showed colocalization of Ca and Cu, and X-ray absorption spectroscopy (XAS) was consistent with Cu+ accumulation in an ordered structure. Zn resupply restored Cu homeostasis concomitant with reduced abundance of these structures. Cu isotope labeling demonstrated that sequestered Cu+ became bio-available for the synthesis of plastocyanin, and transcriptome profiling indicated that mobilized Cu became visible to CRR1. Cu trafficking to intracellular accumulation sites may be a strategy for preventing protein mis-metallation during Zn deficiency and enabling efficient cuproprotein (re)-metallation upon Zn resupply. PMID:25344811

  1. Subcellular metal imaging identifies dynamic sites of Cu accumulation in Chlamydomonas.

    PubMed

    Hong-Hermesdorf, Anne; Miethke, Marcus; Gallaher, Sean D; Kropat, Janette; Dodani, Sheel C; Chan, Jefferson; Barupala, Dulmini; Domaille, Dylan W; Shirasaki, Dyna I; Loo, Joseph A; Weber, Peter K; Pett-Ridge, Jennifer; Stemmler, Timothy L; Chang, Christopher J; Merchant, Sabeeha S

    2014-12-01

    We identified a Cu-accumulating structure with a dynamic role in intracellular Cu homeostasis. During Zn limitation, Chlamydomonas reinhardtii hyperaccumulates Cu, a process dependent on the nutritional Cu sensor CRR1, but it is functionally Cu deficient. Visualization of intracellular Cu revealed major Cu accumulation sites coincident with electron-dense structures that stained positive for low pH and polyphosphate, suggesting that they are lysosome-related organelles. Nano-secondary ion MS showed colocalization of Ca and Cu, and X-ray absorption spectroscopy was consistent with Cu(+) accumulation in an ordered structure. Zn resupply restored Cu homeostasis concomitant with reduced abundance of these structures. Cu isotope labeling demonstrated that sequestered Cu(+) became bioavailable for the synthesis of plastocyanin, and transcriptome profiling indicated that mobilized Cu became visible to CRR1. Cu trafficking to intracellular accumulation sites may be a strategy for preventing protein mismetallation during Zn deficiency and enabling efficient cuproprotein metallation or remetallation upon Zn resupply. PMID:25344811

  2. Spectroscopic definition of the copper active sites in mordenite: selective methane oxidation.

    PubMed

    Vanelderen, Pieter; Snyder, Benjamin E R; Tsai, Ming-Li; Hadt, Ryan G; Vancauwenbergh, Julie; Coussens, Olivier; Schoonheydt, Robert A; Sels, Bert F; Solomon, Edward I

    2015-05-20

    Two distinct [Cu-O-Cu](2+) sites with methane monooxygenase activity are identified in the zeolite Cu-MOR, emphasizing that this Cu-O-Cu active site geometry, having a ∠Cu-O-Cu ∼140°, is particularly formed and stabilized in zeolite topologies. Whereas in ZSM-5 a similar [Cu-O-Cu](2+) active site is located in the intersection of the two 10 membered rings, Cu-MOR provides two distinct local structures, situated in the 8 membered ring windows of the side pockets. Despite their structural similarity, as ascertained by electronic absorption and resonance Raman spectroscopy, the two Cu-O-Cu active sites in Cu-MOR clearly show different kinetic behaviors in selective methane oxidation. This difference in reactivity is too large to be ascribed to subtle differences in the ground states of the Cu-O-Cu sites, indicating the zeolite lattice tunes their reactivity through second-sphere effects. The MOR lattice is therefore functionally analogous to the active site pocket of a metalloenzyme, demonstrating that both the active site and its framework environment contribute to and direct reactivity in transition metal ion-zeolites.

  3. School Pharmacist/School Environmental Hygienic Activities at School Site.

    PubMed

    Muramatsu, Akiyoshi

    2016-01-01

    The "School Health and Safety Act" was enforced in April 2009 in Japan, and "school environmental health standards" were established by the Minister of Education, Culture, Sports, Science and Technology. In Article 24 of the Enforcement Regulations, the duties of the school pharmacist have been clarified; school pharmacists have charged with promoting health activities in schools and carrying out complete and regular checks based on the "school environmental health standards" in order to protect the health of students and staff. In supported of this, the school pharmacist group of Japan Pharmaceutical Association has created and distributed digital video discs (DVDs) on "check methods of school environmental health standards" as support material. We use the DVD to ensure the basic issues that school pharmacists deal with, such as objectives, criteria, and methods for each item to be checked, advice, and post-measures. We conduct various workshops and classes, and set up Q&A committees so that inquiries from members are answered with the help of such activities. In addition, school pharmacists try to improve the knowledge of the school staff on environmental hygiene during their in-service training. They also conduct "drug abuse prevention classes" at school and seek to improve knowledge and recognition of drugs, including "dangerous drugs". PMID:27252053

  4. Ligand binding and proton exchange dynamics in site-specific mutants of human myoglobin

    SciTech Connect

    Lambright, D.G.

    1992-01-01

    Site specific mutagenesis was used to make substitutions of four residues in the distal heme pocket of human myoglobin: Val68, His64, Lys45, and Asp60. Strongly diffracting crystals of the conservative mutation K45R in the met aquo form were grown in the trigonal space group P3[sub 2]21 and the X-ray crystal structure determined at 1.6 [angstrom] resolution. The overall structure is similar to that of sperm whale met aquo myoglobin. Several of the mutant proteins were characterized by 2-D NMR spectroscopy. The NMR data suggest the structural changes are localized to the region of the mutation. The dynamics of ligand binding to myoglobin mutants were studied by transient absorption spectroscopy following photolysis of the CO complexes. Transient absorption kinetics and spectra on the ns to ms timescale were measured in aqueous solution from 280 K to 310 K and in 75% glycerol: water from 250 K to 310 K. Two significant basis spectra were obtained from singular value decomposition of the matrix of time dependent spectra. The information was used to obtain approximations for the extent of ligand rebinding and the kinetics of conformational relaxation. Except for K45R, substitutions at Lys45 or Asp60 produce changes in the kinetics for ligand rebinding. Replacement of Lys45 with Arg increases the rate of ligand rebinding from the protein matrix by a factor of 2, but does not alter the rates for ligand escape or entry into the protein or the dynamics of the conformational relaxation. Substitutions at His64 and Val68 influence the kinetics of ligand rebinding and the dynamics of conformational relaxation. The results do not support the hypothesis that ligand migration between the heme pocket and solvent is determined solely by fluctuations of Arg45 and His64 between open and closed conformations of the heme pocket but can be rationalized if ligand diffusion through the protein matrix involves multiple competing pathways.

  5. Dynamic Kinetics of Nitrogen Cycle in Groundwater-Surface Water Interaction Zone at Hanford Site

    NASA Astrophysics Data System (ADS)

    Liu, Y.; Liu, C.; Liu, Y.; Xu, F.; Yan, A.; Shi, L.; Zachara, J. M.; Gao, Y.; Qian, W.; Nelson, W.; Fredrickson, J.; Zhong, L.; Thompson, C.

    2015-12-01

    Nitrogen cycle carried out by microbes is an important geobiological process that has global implications for carbon and nitrogen cycling and climate change. This presentation describes a study of nitrogen cycle in groundwater-surface water interaction zone (GSIZ) at the US Department of Energy's Hanford Site. Groundwater at Hanford sites has long been documented with nitrate contamination. Nearby Columbia River stage changes of up to 3 m every day because of daily discharge fluctuation from upstream Priest Rapids Dam; resulting an exchange of groundwater and surface water in a short time period. Yet, nitrogen cycle in the GSIZ at Hanford Site remains unclear. Column studies have been used to identify nitrogen metabolism pathways and investigate kinetics of nitrogen cycle in groundwater saturated zone, surface water saturated zone, and GSIZ. Functional gene and protein abundances were determined by qPCR and PRISM-SRM (high-pressure, high-resolution separations coupled with intelligent selection and multiplexing for sensitive selected reaction monitoring) to identify key enzymatic reactions and metabolic pathways of nitrogen cycle. The results showed that dissimilatory nitrate reduction to ammonium (DNRA) competed with denitrification under anaerobic conditions, reducing 30% of NO3- to NH4+, a cation strongly retained on the sediments. As dissolved oxygen intruded the anaerobic zone with river water, NH4+ was oxidized to NO3-, increasing the mobility of NO3-. Multiplicative Monod models were established to describe nitrogen cycle in columns fed with O2 depleted synthetic groundwater and O2 saturated synthetic river water, respectively. The two models were then coupled to predict the dynamic kinetics of nitrogen cycle in GSIZ.

  6. Bayesian inversions of a dynamic vegetation model at four European grassland sites

    NASA Astrophysics Data System (ADS)

    Minet, J.; Laloy, E.; Tychon, B.; Francois, L.

    2015-05-01

    Eddy covariance data from four European grassland sites are used to probabilistically invert the CARAIB (CARbon Assimilation In the Biosphere) dynamic vegetation model (DVM) with 10 unknown parameters, using the DREAM(ZS) (DiffeRential Evolution Adaptive Metropolis) Markov chain Monte Carlo (MCMC) sampler. We focus on comparing model inversions, considering both homoscedastic and heteroscedastic eddy covariance residual errors, with variances either fixed a priori or jointly inferred together with the model parameters. Agreements between measured and simulated data during calibration are comparable with previous studies, with root mean square errors (RMSEs) of simulated daily gross primary productivity (GPP), ecosystem respiration (RECO) and evapotranspiration (ET) ranging from 1.73 to 2.19, 1.04 to 1.56 g C m-2 day-1 and 0.50 to 1.28 mm day-1, respectively. For the calibration period, using a homoscedastic eddy covariance residual error model resulted in a better agreement between measured and modelled data than using a heteroscedastic residual error model. However, a model validation experiment showed that CARAIB models calibrated considering heteroscedastic residual errors perform better. Posterior parameter distributions derived from using a heteroscedastic model of the residuals thus appear to be more robust. This is the case even though the classical linear heteroscedastic error model assumed herein did not fully remove heteroscedasticity of the GPP residuals. Despite the fact that the calibrated model is generally capable of fitting the data within measurement errors, systematic bias in the model simulations are observed. These are likely due to model inadequacies such as shortcomings in the photosynthesis modelling. Besides the residual error treatment, differences between model parameter posterior distributions among the four grassland sites are also investigated. It is shown that the marginal distributions of the specific leaf area and characteristic

  7. [In vitro evaluation of antacid activity in gastric acid secretion in static and dynamic systems].

    PubMed

    Vatier, J; Malikova-Sekera, E; Vitre, M T; Mignon, M

    1992-01-01

    A valid in vitro evaluation of antacid capacity should consider: 1) the intragastric pH-range; 2) the antacid mechanism; 3) the dependence of antacid activity from intraluminal flux variations; 4) the interaction between proteins and antacids. Pharmacologically, a static method allows 1) to quantify H+ binding sites at different pH-end points of the titration: pH 3.0, 2.0 and 1.0 and 2) to characterize the antacid mechanism, neutralizing activity and/or buffering capacity. In dynamic conditions, using the "artificial stomach-duodenum" model the antacid-induced resistance to acidification was measured, the antacid mechanisms were characterized in regard to intraluminal gastroduodenal flux variations and the incidence of antacid activity on duodenal pH was evaluated. These procedures were applied to antacid evaluation of proteins, as natural antacids, and of drugs containing aluminium salts alone or combined with magnesium salts. Pharmacologically, antacid drugs exhibited a greater amount of H+ binding sites when titration end-point was pH 1.0 than pH 3.0 corresponding to the development of neutralizing activity and/or buffering capacity. In dynamic conditions, the drugs, like proteins, induced a potent resistance to acidification related to gastric emptying fluxes. Antacid effect was supported by neutralizing activity and/or by buffering capacity. It was prolonged by removal of H+ ions since lagtimes for recovering initial pH were longer than antacid total emptying, the dilution of intragastric content by H+ impoverished secretory flux contributing thus to prevent gastric acidification. At duodenal site, proteins and aluminium-containing antacids induced the same duodenal pH as controls, without antacids, while magnesium-containing antacids increased it. PMID:1412150

  8. An Accessory Agonist Binding Site Promotes Activation of α4β2* Nicotinic Acetylcholine Receptors.

    PubMed

    Wang, Jingyi; Kuryatov, Alexander; Sriram, Aarati; Jin, Zhuang; Kamenecka, Theodore M; Kenny, Paul J; Lindstrom, Jon

    2015-05-29

    Neuronal nicotinic acetylcholine receptors containing α4, β2, and sometimes other subunits (α4β2* nAChRs) regulate addictive and other behavioral effects of nicotine. These nAChRs exist in several stoichiometries, typically with two high affinity acetylcholine (ACh) binding sites at the interface of α4 and β2 subunits and a fifth accessory subunit. A third low affinity ACh binding site is formed when this accessory subunit is α4 but not if it is β2. Agonists selective for the accessory ACh site, such as 3-[3-(3-pyridyl)-1,2,4-oxadiazol-5-yl]benzonitrile (NS9283), cannot alone activate a nAChR but can facilitate more efficient activation in combination with agonists at the canonical α4β2 sites. We therefore suggest categorizing agonists according to their site selectivity. NS9283 binds to the accessory ACh binding site; thus it is termed an accessory site-selective agonist. We expressed (α4β2)2 concatamers in Xenopus oocytes with free accessory subunits to obtain defined nAChR stoichiometries and α4/accessory subunit interfaces. We show that α2, α3, α4, and α6 accessory subunits can form binding sites for ACh and NS9283 at interfaces with α4 subunits, but β2 and β4 accessory subunits cannot. To permit selective blockage of the accessory site, α4 threonine 126 located on the minus side of α4 that contributes to the accessory site, but not the α4β2 sites, was mutated to cysteine. Alkylation of this cysteine with a thioreactive reagent blocked activity of ACh and NS9283 at the accessory site. Accessory agonist binding sites are promising drug targets.

  9. Structural and Dynamic Requirements for Optimal Activity of the Essential Bacterial Enzyme Dihydrodipicolinate Synthase

    PubMed Central

    Reboul, C. F.; Porebski, B. T.; Griffin, M. D. W.; Dobson, R. C. J.; Perugini, M. A.; Gerrard, J. A.; Buckle, A. M.

    2012-01-01

    Dihydrodipicolinate synthase (DHDPS) is an essential enzyme involved in the lysine biosynthesis pathway. DHDPS from E. coli is a homotetramer consisting of a ‘dimer of dimers’, with the catalytic residues found at the tight-dimer interface. Crystallographic and biophysical evidence suggest that the dimers associate to stabilise the active site configuration, and mutation of a central dimer-dimer interface residue destabilises the tetramer, thus increasing the flexibility and reducing catalytic efficiency and substrate specificity. This has led to the hypothesis that the tetramer evolved to optimise the dynamics within the tight-dimer. In order to gain insights into DHDPS flexibility and its relationship to quaternary structure and function, we performed comparative Molecular Dynamics simulation studies of native tetrameric and dimeric forms of DHDPS from E. coli and also the native dimeric form from methicillin-resistant Staphylococcus aureus (MRSA). These reveal a striking contrast between the dynamics of tetrameric and dimeric forms. Whereas the E. coli DHDPS tetramer is relatively rigid, both the E. coli and MRSA DHDPS dimers display high flexibility, resulting in monomer reorientation within the dimer and increased flexibility at the tight-dimer interface. The mutant E. coli DHDPS dimer exhibits disorder within its active site with deformation of critical catalytic residues and removal of key hydrogen bonds that render it inactive, whereas the similarly flexible MRSA DHDPS dimer maintains its catalytic geometry and is thus fully functional. Our data support the hypothesis that in both bacterial species optimal activity is achieved by fine tuning protein dynamics in different ways: E. coli DHDPS buttresses together two dimers, whereas MRSA dampens the motion using an extended tight-dimer interface. PMID:22685390

  10. How Intrinsic Molecular Dynamics Control Intramolecular Communication in Signal Transducers and Activators of Transcription Factor STAT5

    PubMed Central

    Langenfeld, Florent; Guarracino, Yann; Arock, Michel; Trouvé, Alain; Tchertanov, Luba

    2015-01-01

    Signal Transducer and Activator of Transcription STAT5 is a key mediator of cell proliferation, differentiation and survival. While STAT5 activity is tightly regulated in normal cells, its constitutive activation directly contributes to oncogenesis and is associated with a broad range of hematological and solid tumor cancers. Therefore the development of compounds able to modulate pathogenic activation of this protein is a very challenging endeavor. A crucial step of drug design is the understanding of the protein conformational features and the definition of putative binding site(s) for such modulators. Currently, there is no structural data available for human STAT5 and our study is the first footprint towards the description of structure and dynamics of this protein. We investigated structural and dynamical features of the two STAT5 isoforms, STAT5a and STAT5b, taken into account their phosphorylation status. The study was based on the exploration of molecular dynamics simulations by different analytical methods. Despite the overall folding similarity of STAT5 proteins, the MD conformations display specific structural and dynamical features for each protein, indicating first, sequence-encoded structural properties and second, phosphorylation-induced effects which contribute to local and long-distance structural rearrangements interpreted as allosteric event. Further examination of the dynamical coupling between distant sites provides evidence for alternative profiles of the communication pathways inside and between the STAT5 domains. These results add a new insight to the understanding of the crucial role of intrinsic molecular dynamics in mediating intramolecular signaling in STAT5. Two pockets, localized in close proximity to the phosphotyrosine-binding site and adjacent to the channel for communication pathways across STAT5, may constitute valid targets to develop inhibitors able to modulate the function-related communication properties of this signaling

  11. Isolated metal active site concentration and stability control catalytic CO2 reduction selectivity.

    PubMed

    Matsubu, John C; Yang, Vanessa N; Christopher, Phillip

    2015-03-01

    CO2 reduction by H2 on heterogeneous catalysts is an important class of reactions that has been studied for decades. However, atomic scale details of structure-function relationships are still poorly understood. Particularly, it has been suggested that metal particle size plays a unique role in controlling the stability of CO2 hydrogenation catalysts and the distribution of active sites, which dictates reactivity and selectivity. These studies often have not considered the possible role of isolated metal active sites in the observed dependences. Here, we utilize probe molecule diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) with known site-specific extinction coefficients to quantify the fraction of Rh sites residing as atomically dispersed isolated sites (Rhiso), as well as Rh sites on the surface of Rh nanoparticles (RhNP) for a series of TiO2 supported Rh catalysts. Strong correlations were observed between the catalytic reverse water gas shift turn over frequency (TOF) and the fraction of Rhiso sites and between catalytic methanation TOF and the fraction of RhNP sites. Furthermore, it was observed that reaction condition-induced disintegration of Rh nanoparticles, forming Rhiso active sites, controls the changing reactivity with time on stream. This work demonstrates that isolated atoms and nanoparticles of the same metal on the same support can exhibit uniquely different catalytic selectivity in competing parallel reaction pathways and that disintegration of nanoparticles under reaction conditions can play a significant role in controlling stability.

  12. Isolated metal active site concentration and stability control catalytic CO2 reduction selectivity.

    PubMed

    Matsubu, John C; Yang, Vanessa N; Christopher, Phillip

    2015-03-01

    CO2 reduction by H2 on heterogeneous catalysts is an important class of reactions that has been studied for decades. However, atomic scale details of structure-function relationships are still poorly understood. Particularly, it has been suggested that metal particle size plays a unique role in controlling the stability of CO2 hydrogenation catalysts and the distribution of active sites, which dictates reactivity and selectivity. These studies often have not considered the possible role of isolated metal active sites in the observed dependences. Here, we utilize probe molecule diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) with known site-specific extinction coefficients to quantify the fraction of Rh sites residing as atomically dispersed isolated sites (Rhiso), as well as Rh sites on the surface of Rh nanoparticles (RhNP) for a series of TiO2 supported Rh catalysts. Strong correlations were observed between the catalytic reverse water gas shift turn over frequency (TOF) and the fraction of Rhiso sites and between catalytic methanation TOF and the fraction of RhNP sites. Furthermore, it was observed that reaction condition-induced disintegration of Rh nanoparticles, forming Rhiso active sites, controls the changing reactivity with time on stream. This work demonstrates that isolated atoms and nanoparticles of the same metal on the same support can exhibit uniquely different catalytic selectivity in competing parallel reaction pathways and that disintegration of nanoparticles under reaction conditions can play a significant role in controlling stability. PMID:25671686

  13. Site specific rationale for technical impracticability of active groundwater restoration at a former manufactured gas plant site

    SciTech Connect

    Logan, C.M.; Walden, R.H.; MacFarlane, I.D.

    1995-12-31

    The National Contingency Plan (40 CFR Part 300 ) requires that remedial strategies must, at minimum, protect human health and the environment and meet applicable and relevant or appropriate requirements (ARARs). Where groundwater is impacted, maximum contaminant levels (MCLs) and maximum contaminant level goals (MCLGs) set under the Safe Drinking Water Act are often used as ARARs, whether or not the aquifer is a reasonably anticipated future source of drinking water. The US Environmental Protection Agency now recognizes the difficulty of groundwater restoration at sites where dense nonaqueous phase liquids are present, particularly in certain complex hydrogeological settings (EPA 1993). However, demonstration of impracticability generally does not occur until active remediation (e.g., pump and treat) has been shown to be ineffective. A case study of a former manufactured gas plant (MGP) is used to demonstrate how physical and chemical properties of the aquifer and coal tar, the major waste product from MGP sites, influence the feasibility of active restoration. Field characterization investigations, laboratory studies, and groundwater modeling are integrated into a demonstration following EPA guidelines. Laboratory studies included microbiological characterization and natural biodegradation and suggest that intrinsic bioremediation is occurring at this site. This work will be useful as EPA continues to develop presumptive remedies for cleanup under Superfund.

  14. The calculation of surface orbital energies for specific types of active sites on dispersed metal catalysts

    SciTech Connect

    Augustine, R.L.; Lahanas, K.M.; Cole, F.

    1992-11-01

    An angular overlap calculation has been used to determine the s, p, and d orbital energy levels of the different types of surface sites present on dispersed metal catalysts. These data can permit a Frontier Molecular Orbital treatment of specific site activities as long as the surface orbital availability for overlap with adsorbed substrates is considered along with its energy value and symmetry.

  15. The calculation of surface orbital energies for specific types of active sites on dispersed metal catalysts

    SciTech Connect

    Augustine, R.L.; Lahanas, K.M.; Cole, F.

    1992-01-01

    An angular overlap calculation has been used to determine the s, p, and d orbital energy levels of the different types of surface sites present on dispersed metal catalysts. These data can permit a Frontier Molecular Orbital treatment of specific site activities as long as the surface orbital availability for overlap with adsorbed substrates is considered along with its energy value and symmetry.

  16. Extending the Diffuse Layer Model of Surface Acidity Behavior: III. Estimating Bound Site Activity Coefficients

    EPA Science Inventory

    Although detailed thermodynamic analyses of the 2-pK diffuse layer surface complexation model generally specify bound site activity coefficients for the purpose of accounting for those non-ideal excess free energies contributing to bound site electrochemical potentials, in applic...

  17. Activity of site-specific endonucleases on complexes of plasmid DNA with multiwalled carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Egorova, V. P.; Krylova, H. V.; Lipnevich, I. V.; Veligura, A. A.; Shulitsky, B. G.; Asayonok, A. A.; Vaskovtsev, E. V.

    2016-08-01

    We have synthesized and investigated structural and functional properties of plasmid DNA complexes with multi-walled carbon nanotubes (MWCNTs) for detection of changes in structural state of the plasmid DNA at its recognition by site-specific endonuclease. It has been also established that the site-specific endonuclease is functionally active on the surface of MWCNTs.

  18. 77 FR 5560 - Commercial Wind Lease Issuance and Site Assessment Activities on the Atlantic Outer Continental...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-03

    ... project proposals on those leases) in identified Wind Energy Areas (WEAs) on the OCS offshore New Jersey... Bureau of Ocean Energy Management Commercial Wind Lease Issuance and Site Assessment Activities on the... site assessment plans (SAPs) on those leases. BOEM may issue one or more commercial wind energy...

  19. The balance of flexibility and rigidity in the active site residues of hen egg white lysozyme

    NASA Astrophysics Data System (ADS)

    Qi, Jian-Xun; Jiang, Fan

    2011-05-01

    The crystallographic temperature factors (B factor) of individual atoms contain important information about the thermal motion of the atoms in a macromolecule. Previously the theory of flexibility of active site has been established based on the observation that the enzyme activity is sensitive to low concentration denaturing agents. It has been found that the loss of enzyme activity occurs well before the disruption of the three-dimensional structural scaffold of the enzyme. To test the theory of conformational flexibility of enzyme active site, crystal structures were perturbed by soaking in low concentration guanidine hydrochloride solutions. It was found that many lysozyme crystals tested could still diffract until the concentration of guanidine hydrochloride reached 3 M. It was also found that the B factors averaged over individually collected data sets were more accurate. Thus it suggested that accurate measurement of crystal temperature factors could be achieved for medium-high or even medium resolution crystals by averaging over multiple data sets. Furthermore, we found that the correctly predicted active sites included not only the more flexible residues, but also some more rigid residues. Both the flexible and the rigid residues in the active site played an important role in forming the active site residue network, covering the majority of the substrate binding residues. Therefore, this experimental prediction method may be useful for characterizing the binding site and the function of a protein, such as drug targeting.

  20. Automatic generation of active coordinates for quantum dynamics calculations: Application to the dynamics of benzene photochemistry

    SciTech Connect

    Lasorne, Benjamin; Sicilia, Fabrizio; Bearpark, Michael J.; Robb, Michael A.; Worth, Graham A.; Blancafort, Lluis

    2008-03-28

    A new practical method to generate a subspace of active coordinates for quantum dynamics calculations is presented. These reduced coordinates are obtained as the normal modes of an analytical quadratic representation of the energy difference between excited and ground states within the complete active space self-consistent field method. At the Franck-Condon point, the largest negative eigenvalues of this Hessian correspond to the photoactive modes: those that reduce the energy difference and lead to the conical intersection; eigenvalues close to 0 correspond to bath modes, while modes with large positive eigenvalues are photoinactive vibrations, which increase the energy difference. The efficacy of quantum dynamics run in the subspace of the photoactive modes is illustrated with the photochemistry of benzene, where theoretical simulations are designed to assist optimal control experiments.

  1. Dynamic changes during acid-induced activation of influenza hemagglutinin

    PubMed Central

    Garcia, Natalie K.; Guttman, Miklos; Ebner, Jamie L.; Lee, Kelly K.

    2015-01-01

    SUMMARY Influenza hemagglutinin (HA) mediates virus attachment to host cells and fusion of the viral and endosomal membranes during entry. While high-resolution structures are available for the pre-fusion HA ectodomain and the post-fusion HA2 subunit, the sequence of conformational changes during HA activation has eluded structural characterization. Here we apply hydrogen-deuterium exchange with mass spectrometry to examine changes in structural dynamics of the HA ectodomain at various stages of activation, as well as to compare the soluble ectodomain with intact HA on virions. At pH conditions approaching activation (pH 6.0–5.5) HA exhibits increased dynamics at the fusion peptide and neighboring regions, while the interface between receptor-binding subunits (HA1) becomes stabilized. In contrast to many activation models, these data suggest that HA responds to endosomal acidification by releasing the fusion peptide prior to HA1 uncaging and the spring-loaded refolding of HA2. This staged process may facilitate efficient HA-mediated fusion. PMID:25773144

  2. Chemical modification studies on arginine kinase: essential cysteine and arginine residues at the active site.

    PubMed

    Zhu, Wen-Jing; Li, Miao; Wang, Xiao-Yun

    2007-12-01

    Chemical modification was used to elucidate the essential amino acids in the catalytic activity of arginine kinase (AK) from Migratoria manilensis. Among six cysteine (Cys) residues only one Cys residue was determined to be essential in the active site by Tsou's method. Furthermore, the AK modified by DTNB can be fully reactivated by dithiothreitol (DTT) in a monophasic kinetic course. At the same time, this reactivation can be slowed down in the presence of ATP, suggesting that the essential Cys is located near the ATP binding site. The ionizing groups at the AK active site were studied and the standard dissociation enthalpy (DeltaH degrees ) was 12.38kcal/mol, showing that the dissociation group may be the guanidino of arginine (Arg). Using the specific chemical modifier phenylglyoxal (PG) demonstrated that only one Arg, located near the ATP binding site, is essential for the activity of AK. PMID:17765964

  3. 1993 annual report of hazardous waste activities for the Oak Ridge K-25 site

    SciTech Connect

    Not Available

    1994-02-01

    This report is a detailed listing of all of the Hazardous Waste activities occurring at Martin Marietta`s K-25 site. Contained herein are hazardous waste notification forms, waste stream reports, generator fee forms and various TSDR reports.

  4. Multi-Dimensional Dynamics of Human Electromagnetic Brain Activity

    PubMed Central

    Kida, Tetsuo; Tanaka, Emi; Kakigi, Ryusuke

    2016-01-01

    Magnetoencephalography (MEG) and electroencephalography (EEG) are invaluable neuroscientific tools for unveiling human neural dynamics in three dimensions (space, time, and frequency), which are associated with a wide variety of perceptions, cognition, and actions. MEG/EEG also provides different categories of neuronal indices including activity magnitude, connectivity, and network properties along the three dimensions. In the last 20 years, interest has increased in inter-regional connectivity and complex network properties assessed by various sophisticated scientific analyses. We herein review the definition, computation, short history, and pros and cons of connectivity and complex network (graph-theory) analyses applied to MEG/EEG signals. We briefly describe recent developments in source reconstruction algorithms essential for source-space connectivity and network analyses. Furthermore, we discuss a relatively novel approach used in MEG/EEG studies to examine the complex dynamics represented by human brain activity. The correct and effective use of these neuronal metrics provides a new insight into the multi-dimensional dynamics of the neural representations of various functions in the complex human brain. PMID:26834608

  5. Multi-Dimensional Dynamics of Human Electromagnetic Brain Activity.

    PubMed

    Kida, Tetsuo; Tanaka, Emi; Kakigi, Ryusuke

    2015-01-01

    Magnetoencephalography (MEG) and electroencephalography (EEG) are invaluable neuroscientific tools for unveiling human neural dynamics in three dimensions (space, time, and frequency), which are associated with a wide variety of perceptions, cognition, and actions. MEG/EEG also provides different categories of neuronal indices including activity magnitude, connectivity, and network properties along the three dimensions. In the last 20 years, interest has increased in inter-regional connectivity and complex network properties assessed by various sophisticated scientific analyses. We herein review the definition, computation, short history, and pros and cons of connectivity and complex network (graph-theory) analyses applied to MEG/EEG signals. We briefly describe recent developments in source reconstruction algorithms essential for source-space connectivity and network analyses. Furthermore, we discuss a relatively novel approach used in MEG/EEG studies to examine the complex dynamics represented by human brain activity. The correct and effective use of these neuronal metrics provides a new insight into the multi-dimensional dynamics of the neural representations of various functions in the complex human brain. PMID:26834608

  6. Dynamic Control of Synchronous Activity in Networks of Spiking Neurons

    PubMed Central

    Hutt, Axel; Mierau, Andreas; Lefebvre, Jérémie

    2016-01-01

    Oscillatory brain activity is believed to play a central role in neural coding. Accumulating evidence shows that features of these oscillations are highly dynamic: power, frequency and phase fluctuate alongside changes in behavior and task demands. The role and mechanism supporting this variability is however poorly understood. We here analyze a network of recurrently connected spiking neurons with time delay displaying stable synchronous dynamics. Using mean-field and stability analyses, we investigate the influence of dynamic inputs on the frequency of firing rate oscillations. We show that afferent noise, mimicking inputs to the neurons, causes smoothing of the system’s response function, displacing equilibria and altering the stability of oscillatory states. Our analysis further shows that these noise-induced changes cause a shift of the peak frequency of synchronous oscillations that scales with input intensity, leading the network towards critical states. We lastly discuss the extension of these principles to periodic stimulation, in which externally applied driving signals can trigger analogous phenomena. Our results reveal one possible mechanism involved in shaping oscillatory activity in the brain and associated control principles. PMID:27669018

  7. Divergent contributions of conserved active site residues to transcription by eukaryotic RNA polymerases I and II

    PubMed Central

    Viktorovskaya, Olga V.; Engel, Krysta L.; French, Sarah L.; Cui, Ping; Vandeventer, Paul J.; Pavlovic, Emily M.; Beyer, Ann L.; Kaplan, Craig D.; Schneider, David A.

    2013-01-01

    SUMMARY Multisubunit RNA polymerases (msRNAPs) exhibit high sequence and structural homology, especially within their active sites, which is generally thought to result in msRNAP functional conservation. However, we show that mutations in the trigger loop (TL) in the largest subunit of RNA polymerase I (Pol I) yield phenotypes unexpected from studies of Pol II. For example, a well-characterized gain-of-function mutation in Pol II results in loss-of-function in Pol I [Pol II: rpb1- E1103G; Pol I: rpa190-E1224G]. Studies of chimeric Pol II enzymes hosting Pol I or Pol III TLs suggest that consequences of mutations that alter TL dynamics are dictated by the greater enzymatic context and not solely the TL sequence. Although the rpa190-E1224G mutation diminishes polymerase function, when combined with mutations that perturb Pol I catalysis, it enhances polymerase function, similar to the analogous Pol II mutation. These results suggest that Pol I and Pol II have different rate-limiting steps. PMID:23994471

  8. The transient catalytically competent coenzyme allocation into the active site of Anabaena ferredoxin NADP+ -reductase.

    PubMed

    Peregrina, José Ramón; Lans, Isaías; Medina, Milagros

    2012-01-01

    Ferredoxin-NADP(+) reductase (FNR) catalyses the electron transfer from ferredoxin to NADP(+) via its flavin FAD cofactor. A molecular dynamics theoretical approach is applied here to visualise the transient catalytically competent interaction of Anabaena FNR with its coenzyme, NADP(+). The particular role of some of the residues identified as key in binding and accommodating the 2'P-AMP moiety of the coenzyme is confirmed in molecular terms. Simulations also indicate that the architecture of the active site precisely contributes to the orientation of the N5 of the FAD isoalloxazine ring and the C4 of the coenzyme nicotinamide ring in the conformation of the catalytically competent hydride transfer complex and, therefore, contributes to the efficiency of the process. In particular, the side chain of the C-terminal Y303 in Anabaena FNR appears key to providing the optimum geometry by reducing the stacking probability between the isoalloxazine and nicotinamide rings, thus providing the required co-linearity and distance among the N5 of the flavin cofactor, the C4 of the coenzyme nicotinamide and the hydride that has to be transferred between them. All these factors are highly related to the reaction efficiency, mechanism and reversibility of the process.

  9. Visual Experience Modulates Spatio-Temporal Dynamics of Circuit Activation

    PubMed Central

    Wang, Lang; Fontanini, Alfredo; Maffei, Arianna

    2011-01-01

    Persistent reduction in sensory drive in early development results in multiple plastic changes of different cortical synapses. How these experience-dependent modifications affect the spatio-temporal dynamics of signal propagation in neocortical circuits is poorly understood. Here we demonstrate that brief visual deprivation significantly affects the propagation of electrical signals in the primary visual cortex. The spatio-temporal spread of circuit activation upon direct stimulation of its input layer (Layer 4) is reduced, as is the activation of L2/3 – the main recipient of the output from L4. Our data suggest that the decrease in spatio-temporal activation of L2/3 depends on reduced L4 output, and is not intrinsically generated within L2/3. The data shown here suggest that changes in the synaptic components of the visual cortical circuit result not only in alteration of local integration of excitatory and inhibitory inputs, but also in a significant decrease in overall circuit activation. Furthermore, our data indicate a differential effect of visual deprivation on L4 and L2/3, suggesting that while feedforward activation of L2/3 is reduced, its activation by long range, within layer inputs is unaltered. Thus, brief visual deprivation induces experience-dependent circuit re-organization by modulating not only circuit excitability, but also the spatio-temporal patterns of cortical activation within and between layers. PMID:21743804

  10. Anisotropic Covalency Contributions to Superexchange Pathways in Type One Copper Active Sites

    PubMed Central

    2015-01-01

    Type one (T1) Cu sites deliver electrons to catalytic Cu active sites: the mononuclear type two (T2) Cu site in nitrite reductases (NiRs) and the trinuclear Cu cluster in the multicopper oxidases (MCOs). The T1 Cu and the remote catalytic sites are connected via a Cys-His intramolecular electron-transfer (ET) bridge, which contains two potential ET pathways: P1 through the protein backbone and P2 through the H-bond between the Cys and the His. The high covalency of the T1 Cu–S(Cys) bond is shown here to activate the T1 Cu site for hole superexchange via occupied valence orbitals of the bridge. This covalency-activated electronic coupling (HDA) facilitates long-range ET through both pathways. These pathways can be selectively activated depending on the geometric and electronic structure of the T1 Cu site and thus the anisotropic covalency of the T1 Cu–S(Cys) bond. In NiRs, blue (π-type) T1 sites utilize P1 and green (σ-type) T1 sites utilize P2, with P2 being more efficient. Comparing the MCOs to NiRs, the second-sphere environment changes the conformation of the Cys-His pathway, which selectively activates HDA for superexchange by blue π sites for efficient turnover in catalysis. These studies show that a given protein bridge, here Cys-His, provides different superexchange pathways and electronic couplings depending on the anisotropic covalencies of the donor and acceptor metal sites. PMID:25310460

  11. Superfluid-like dynamics in active vortex fluids

    NASA Astrophysics Data System (ADS)

    Slomka, Jonasz; Dunkel, Jorn

    Active biological fluids exhibit rich non-equilibrium dynamics and share striking similarities with quantum fluids, from vortex formation and magnetic ordering to superfluid-like behavior. Building on universality ideas, we have recently proposed a generalization of the Navier-Stokes equations that captures qualitatively the active bulk flow structures observed in bacterial suspensions. Here, we present new numerical simulations that explicitly account for boundary and shear effects. The theory successfully reproduces recent experimental observations of bacterial suspensions, including a superfluid-like regime of nearly vanishing shear viscosity. Our simulations further predict a geometry-induced 'quantization' of viscosity and the existence of excited states capable of performing mechanical work. It is plausible that these results generalize to a broad a class of fluids that are subject to an active scale selection mechanism.

  12. CARER: Efficient Dynamic Sensing for Continuous Activity Monitoring

    PubMed Central

    Au, Lawrence K.; Bui, Alex A.T.; Batalin, Maxim A.; Xu, Xiaoyu; Kaiser, William J.

    2016-01-01

    Advancement in wireless health sensor systems has triggered rapidly expanding research in continuous activity monitoring for chronic disease management or promotion and assessment of physical rehabilitation. Wireless motion sensing is increasingly important in treatments where remote collection of sensor measurements can provide an in-field objective evaluation of physical activity patterns. The well-known challenge of limited operating lifetime of energy-constrained wireless health sensor systems continues to present a primary limitation for these applications. This paper introduces CARER, a software system that supports a novel algorithm that exploits knowledge of context and dynamically schedules sensor measurement episodes within an energy consumption budget while ensuring classification accuracy. The sensor selection algorithm in the CARER system is based on Partially Observable Markov Decision Process (POMDP). The parameters for the POMDP algorithm can be obtained through standard maximum likelihood estimation. Sensor data are also collected from multiple locations of the subjects body, providing estimation of an individual's daily activity patterns. PMID:22254783

  13. Vertebral shape: automatic measurement with dynamically sequenced active appearance models.

    PubMed

    Roberts, M G; Cootes, T F; Adams, J E

    2005-01-01

    The shape and appearance of vertebrae on lateral dual x-ray absorptiometry (DXA) scans were statistically modelled. The spine was modelled by a sequence of overlapping triplets of vertebrae, using Active Appearance Models (AAMs). To automate vertebral morphometry, the sequence of trained models was matched to previously unseen scans. The dataset includes a significant number of pathologies. A new dynamic ordering algorithm was assessed for the model fitting sequence, using the best quality of fit achieved by multiple sub-model candidates. The accuracy of the search was improved by dynamically imposing the best quality candidate first. The results confirm the feasibility of substantially automating vertebral morphometry measurements even with fractures or noisy images.

  14. Probing impact of active site residue mutations on stability and activity of Neisseria polysaccharea amylosucrase.

    PubMed

    Daudé, David; Topham, Christopher M; Remaud-Siméon, Magali; André, Isabelle

    2013-12-01

    The amylosucrase from Neisseria polysaccharea is a transglucosidase from the GH13 family of glycoside-hydrolases that naturally catalyzes the synthesis of α-glucans from the widely available donor sucrose. Interestingly, natural molecular evolution has modeled a dense hydrogen bond network at subsite -1 responsible for the specific recognition of sucrose and conversely, it has loosened interactions at the subsite +1 creating a highly promiscuous subsite +1. The residues forming these subsites are considered to be likely involved in the activity as well as the overall stability of the enzyme. To assess their role, a structure-based approach was followed to reshape the subsite -1. A strategy based on stability change predictions, using the FoldX algorithm, was considered to identify the best candidates for site-directed mutagenesis and guide the construction of a small targeted library. A miniaturized purification protocol was developed and both mutant stability and substrate promiscuity were explored. A range of 8 °C between extreme melting temperature values was observed and some variants were able to synthesize series of oligosaccharides with distributions differing from that of the parental enzyme. The crucial role of subsite -1 was thus highlighted and the biocatalysts generated can now be considered as starting points for further engineering purposes.

  15. 'Unconventional' coordination chemistry by metal chelating fragments in a metalloprotein active site.

    PubMed

    Martin, David P; Blachly, Patrick G; Marts, Amy R; Woodruff, Tessa M; de Oliveira, César A F; McCammon, J Andrew; Tierney, David L; Cohen, Seth M

    2014-04-01

    The binding of three closely related chelators: 5-hydroxy-2-methyl-4H-pyran-4-thione (allothiomaltol, ATM), 3-hydroxy-2-methyl-4H-pyran-4-thione (thiomaltol, TM), and 3-hydroxy-4H-pyran-4-thione (thiopyromeconic acid, TPMA) to the active site of human carbonic anhydrase II (hCAII) has been investigated. Two of these ligands display a monodentate mode of coordination to the active site Zn(2+) ion in hCAII that is not recapitulated in model complexes of the enzyme active site. This unprecedented binding mode in the hCAII-thiomaltol complex has been characterized by both X-ray crystallography and X-ray spectroscopy. In addition, the steric restrictions of the active site force the ligands into a 'flattened' mode of coordination compared with inorganic model complexes. This change in geometry has been shown by density functional computations to significantly decrease the strength of the metal-ligand binding. Collectively, these data demonstrate that the mode of binding by small metal-binding groups can be significantly influenced by the protein active site. Diminishing the strength of the metal-ligand bond results in unconventional modes of metal coordination not found in typical coordination compounds or even carefully engineered active site models, and understanding these effects is critical to the rational design of inhibitors that target clinically relevant metalloproteins.

  16. Site-directed mutagenesis and high-resolution NMR spectroscopy of the active site of porphobilinogen deaminase

    SciTech Connect

    Scott, A.I.; Roessner, C.A.; Stolowich, N.J.; Karuso, P.; Williams, H.J.; Grant, S.K.; Gonzalez, M.D.; Hoshino, T. )

    1988-10-18

    The active site of porphobilinogen (PBG){sup 1} deaminase from Escherichia coli has been found to contain an unusual dipyrromethane derived from four molecules of 5-aminolevulinic acid (ALA) covalently linked to Cys-242, one of the two cysteine residues conserved in E. coli and human deaminase. By use of a hemA{sup {minus}} strain of E. coli the enzyme was enriched from (5-{sup 13}C)ALA and examined by {sup 1}H-detected multiple quantum coherence spectroscopy, which revealed all of the salient features of a dipyrromethane composed of two PBG units linked heat to tail and terminating in a CH{sub 2}-S bond to a cysteine residue. Site-specific mutagenesis of Cys-99 and Cys-242, respectively, has shown that substitution of Ser for Cys-99 does not affect the enzymatic activity, whereas substitution of Ser for Cys-242 removes essentially all of the catalytic activity as measured by the conversion of the substrate PBG to uro'gen I. The NMR spectrum of the covalent complex of deaminase with the suicide inhibitor 2-bromo-(2,11-{sup 13}C{sub 2})PBG reveals that the aminomethyl terminus of the inhibitor reacts with the enzyme's cofactor at the {alpha}-free pyrrole. NMR spectroscopy of the ES{sub 2} complex confirmed a PBG-derived head-to-tail dipyrromethane attached to the {alpha}-free pyrrole position of the enzyme. A mechanistic rationale for deaminase is presented.

  17. Nuclear waste: Status of DOE`s nuclear waste site characterization activities

    SciTech Connect

    1987-12-31

    Three potential nuclear waste repository sites have been selected to carry out characterization activities-the detailed geological testing to determine the suitability of each site as a repository. The sites are Hanford in south-central Washington State, Yucca Mountain in southern Nevada, and Deaf Smith in the Texas Panhandle. Two key issues affecting the total program are the estimations of the site characterization completion data and costs and DOE`s relationship with the Nuclear Regulatory Commission which has been limited and its relations with affected states and Indian tribes which continue to be difficult.

  18. XAFS Study of the Photo-Active Site of Mo/MCM-41

    NASA Astrophysics Data System (ADS)

    Miyamoto, Daisuke; Ichikuni, Nobuyuki; Shimazu, Shogo

    2007-02-01

    An Mo/MCM-41 catalyst was prepared and used for study of propene and 1-butene photo-metathesis reactions. XAFS analysis revealed that hydrogen reduction leads to a decreased role for the Mo=O site. The Mo-O site plays an important role for the olefin photo-metathesis reaction on the H2 reduced Mo/MCM-41. From EXAFS analysis, the active site of photo-metathesis reaction is the Mo=O part for oxidized Mo/MCM-41, whereas it is the Mo-O site for reduced Mo/MCM-41.

  19. Binding stability of peptides derived from 1ALA residue and 7GLY residues to sites near active center of fluctuating papain

    NASA Astrophysics Data System (ADS)

    Nishiyama, Katsuhiko

    2012-05-01

    We investigated the binding stability of peptides derived from 1ALA residue and 7GLY residues to sites near active center of fluctuating papain via molecular dynamics and docking simulations. Replacing GLY residue in 8GLY with ALA residue had a positive effect on binding stability to the sites in some cases although the replacing had a negative effect on it in other cases. Furthermore the replacing had a negative effect on the chance of binding to the sites. Residue in peptide should be replaced on the basis of systematic exploration of its position.

  20. Energetics of Ortho-7 (Oxime Drug) Translocation through the Active-Site Gorge of Tabun Conjugated Acetylcholinesterase

    PubMed Central

    Sinha, Vivek; Ganguly, Bishwajit; Bandyopadhyay, Tusar

    2012-01-01

    Oxime drugs translocate through the 20 Å active-site gorge of acetylcholinesterase in order to liberate the enzyme from organophosphorus compounds’ (such as tabun) conjugation. Here we report bidirectional steered molecular dynamics simulations of oxime drug (Ortho-7) translocation through the gorge of tabun intoxicated enzyme, in which time dependent external forces accelerate the translocation event. The simulations reveal the participation of drug-enzyme hydrogen bonding, hydrophobic interactions and water bridges between them. Employing nonequilibrium theorems that recovers the free energy from irreversible work done, we reconstruct potential of mean force along the translocation pathway such that the desired quantity represents an unperturbed system. The potential locates the binding sites and barriers for the drug to translocate inside the gorge. Configurational entropic contribution of the protein-drug binding entity and the role of solvent translational mobility in the binding energetics is further assessed. PMID:22808117

  1. Transcriptional activation through ETS domain binding sites in the cytochrome c oxidase subunit IV gene

    SciTech Connect

    Virbasius, J.V.; Scarpulla, R.C. )

    1991-11-01

    A mutational analysis of the rat cytochrome c oxidase subunit IV (RCO4) promoter region revealed the presence of a major control element consisting of a tandemly repeated pair of binding sites for a nuclear factor from HeLa cells. This factor was designated NRF-2 (nuclear respiratory factor 2) because a functional recognition site was also found in the human ATP synthase {beta}-subunit gene. Deletion or site-directed point mutations of the NRF-2 binding sites in the RCO4 promoter resulted in substantial loss of transcriptional activity, and synthetic oligomers of the NRF-2 binding sites from both genes stimulated a heterologous promoter when cloned in cis. NRF-2 binding a transcriptional activation required a purine-rich core sequence, GGAA. This motif is characteristic of the recognition site for a family of activators referred to as ETS domain proteins because of the similarity within their DNA-binding domains to the ets-1 proto-oncogene product. NRF-2 recognized an authentic Ets-1 site within the Moloney murine sarcoma virus long terminal repeat, and this site was able to compete for NRF-2 binding to the RCO4 promoter sequence. However, in contrast to Ets-1, which appears to be exclusive to lymphoid tissues, NRF-2 has the broad tissue distribution expected of a regulator of respiratory chain expression.

  2. Ligand uptake in Mycobacterium tuberculosis truncated hemoglobins is controlled by both internal tunnels and active site water molecules

    PubMed Central

    Davidge, Kelly S; Singh, Sandip; Bowman, Lesley AH; Tinajero-Trejo, Mariana; Carballal, Sebastián; Radi, Rafael; Poole, Robert K; Dikshit, Kanak; Estrin, Dario A; Marti, Marcelo A; Boechi, Leonardo

    2015-01-01

    Mycobacterium tuberculosis, the causative agent of human tuberculosis, has two proteins belonging to the truncated hemoglobin (trHb) family. Mt-trHbN presents well-defined internal hydrophobic tunnels that allow O 2 and •NO to migrate easily from the solvent to the active site, whereas Mt-trHbO possesses tunnels that are partially blocked by a few bulky residues, particularly a tryptophan at position G8. Differential ligand migration rates allow Mt-trHbN to detoxify •NO, a crucial step for pathogen survival once under attack by the immune system, much more efficiently than Mt-trHbO. In order to investigate the differences between these proteins, we performed experimental kinetic measurements, •NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W. These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site. We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site. PMID:26478812

  3. Ligand uptake in Mycobacterium tuberculosis truncated hemoglobins is controlled by both internal tunnels and active site water molecules.

    PubMed

    Boron, Ignacio; Bustamante, Juan Pablo; Davidge, Kelly S; Singh, Sandip; Bowman, Lesley Ah; Tinajero-Trejo, Mariana; Carballal, Sebastián; Radi, Rafael; Poole, Robert K; Dikshit, Kanak; Estrin, Dario A; Marti, Marcelo A; Boechi, Leonardo

    2015-01-01

    Mycobacterium tuberculosis, the causative agent of human tuberculosis, has two proteins belonging to the truncated hemoglobin (trHb) family. Mt-trHbN presents well-defined internal hydrophobic tunnels that allow O 2 and (•)NO to migrate easily from the solvent to the active site, whereas Mt-trHbO possesses tunnels that are partially blocked by a few bulky residues, particularly a tryptophan at position G8. Differential ligand migration rates allow Mt-trHbN to detoxify (•)NO, a crucial step for pathogen survival once under attack by the immune system, much more efficiently than Mt-trHbO. In order to investigate the differences between these proteins, we performed experimental kinetic measurements, (•)NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W. These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site. We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site.

  4. Activity of radon (222Rn) in the lower atmospheric surface layer of a typical rural site in south India

    NASA Astrophysics Data System (ADS)

    Kumar, K. Charan; Prasad, T. Rajendra; Ratnam, M. Venkat; Nagaraja, Kamsali

    2016-09-01

    Analysis of one year measurements of in situ radon (222Rn) and its progenies along with surface air temperature, relative humidity and pressure near to the Earth's surface has been carried out for the first time at the National Atmospheric Research Laboratory (NARL, 13.5∘N and 79.2∘E) located in a rural site in Gadanki, south India. The dataset was analysed to understand the behaviour of radon in relation to the surface air temperature and relative humidity at a rural site. It was observed that over a period of the 24 hours in a day, the activity of radon and its progenies reaches a peak in the morning hours followed by a remarkable decrease in the afternoon hours. Relatively, a higher concentration of radon was observed at NARL during fair weather days, and this can be attributed to the presence of rocky hills and dense vegetation surrounding the site. The high negative correlation between surface air temperature and activity of radon (R = - 0.70, on an annual scale) suggests that dynamical removal of radon due to increased vertical mixing is one of the most important controlling processes of the radon accumulation in the atmospheric surface layer. The annual averaged activity of radon was found to be 12.01±0.66 Bq m-3 and 4.25±0.18 Bq m-3 for its progenies, in the study period.

  5. A Hydrophobic Pocket in the Active Site of Glycolytic Aldolase Mediates Interactions with Wiskott-Aldrich Syndrome Protein

    SciTech Connect

    St-Jean,M.; Izard, T.; Sygusch, J.

    2007-01-01

    Aldolase plays essential catalytic roles in glycolysis and gluconeogenesis. However, aldolase is a highly abundant protein that is remarkably promiscuous in its interactions with other cellular proteins. In particular, aldolase binds to highly acidic amino acid sequences, including the C-terminus of the Wiskott-Aldrich syndrome protein, an actin nucleation promoting factor. Here we report the crystal structure of tetrameric rabbit muscle aldolase in complex with a C-terminal peptide of Wiskott-Aldrich syndrome protein. Aldolase recognizes a short, 4-residue DEWD motif (residues 498-501), which adopts a loose hairpin turn that folds about the central aromatic residue, enabling its tryptophan side chain to fit into a hydrophobic pocket in the active site of aldolase. The flanking acidic residues in this binding motif provide further interactions with conserved aldolase active site residues, Arg-42 and Arg-303, aligning their side chains and forming the sides of the hydrophobic pocket. The binding of Wiskott-Aldrich syndrome protein to aldolase precludes intramolecular interactions of its C-terminus with its active site, and is competitive with substrate as well as with binding by actin and cortactin. Finally, based on this structure a novel naphthol phosphate-based inhibitor of aldolase was identified and its structure in complex with aldolase demonstrated mimicry of the Wiskott-Aldrich syndrome protein-aldolase interaction. The data support a model whereby aldolase exists in distinct forms that regulate glycolysis or actin dynamics.

  6. Quantitative, directional measurement of electric field heterogeneity in the active site of ketosteroid isomerase.

    PubMed

    Fafarman, Aaron T; Sigala, Paul A; Schwans, Jason P; Fenn, Timothy D; Herschlag, Daniel; Boxer, Steven G

    2012-02-01

    Understanding the electrostatic forces and features within highly heterogeneous, anisotropic, and chemically complex enzyme active sites and their connection to biological catalysis remains a longstanding challenge, in part due to the paucity of incisive experimental probes of electrostatic properties within proteins. To quantitatively assess the landscape of electrostatic fields at discrete locations and orientations within an enzyme active site, we have incorporated site-specific thiocyanate vibrational probes into multiple positions within bacterial ketosteroid isomerase. A battery of X-ray crystallographic, vibrational Stark spectroscopy, and NMR studies revealed electrostatic field heterogeneity of 8 MV/cm between active site probe locations and widely differing sensitivities of discrete probes to common electrostatic perturbations from mutation, ligand binding, and pH changes. Electrostatic calculations based on active site ionization states assigned by literature precedent and computational pK(a) prediction were unable to quantitatively account for the observed vibrational band shifts. However, electrostatic models of the D40N mutant gave qualitative agreement with the observed vibrational effects when an unusual ionization of an active site tyrosine with a pK(a) near 7 was included. UV-absorbance and (13)C NMR experiments confirmed the presence of a tyrosinate in the active site, in agreement with electrostatic models. This work provides the most direct measure of the heterogeneous and anisotropic nature of the electrostatic environment within an enzyme active site, and these measurements provide incisive benchmarks for further developing accurate computational models and a foundation for future tests of electrostatics in enzymatic catalysis.

  7. Prokaryotic dynamics and heterotrophic metabolism in a deep convection site of Eastern Mediterranean Sea (the Southern Adriatic Pit)

    NASA Astrophysics Data System (ADS)

    Azzaro, M.; La Ferla, R.; Maimone, G.; Monticelli, L. S.; Zaccone, R.; Civitarese, G.

    2012-08-01

    We report on investigations of prokaryotic abundance, biomass, extracellular enzymatic activity, prokaryotic heterotrophic production and respiration in the full water column (˜1200 m) of a deep convection site (the Southern Adriatic Pit), carried out on six cruises in 2006-2008. Prokaryotic abundance (PA) varied vertically and temporally and ranged from 1.2 to 20.4×105 cell ml-1. Cell volumes, generally increased with depth; the lowest mean cell volume was observed in a period with no active convective process (Feb-07) and the highest in a period of stratification (Jun-08) following the convection process occurred in Feb-08. Prokaryotic biomass decreased with the depth and was related with both seasonal cycles of organic matter and hydrological processes. The picophytoplankton ranged in the upper layer (UL) from 0.089 to 10.71×104 cell ml-1. Cells were also recorded till 500 m depth in Feb-08 and this finding could be linked to water convection occurred in the Southern Adriatic Pit in that month. In UL the variations of enzymatic activities as well as leucine-aminopeptidase/ß-glucosidase ratio showed a seasonal trend probably linked to the productive processes of the photic layer. An inverse relation between alkaline phosphatase activity (APA) and phosphate concentrations was found (APA=0.0003PO4-1.7714, R2=0.333, P<0.05). Generally cell-specific enzymatic activities increased with depth as did cell-specific carbon dioxide production rates, while cell-specific prokaryotic heterotrophic production had an opposite trend. High values of prokaryotic growth efficiency registered in the deep layers in Nov-06 reflected a supply of preformed C transported within the deep water masses. Overall, in 2007 when no convective phenomenon was observed, the variability of prokaryotic metabolism was governed by the seasonal cycle of the organic matter, while in Nov-06 and Jun-08 the dynamics of deep water ventilation influenced the trend along the water column of many microbial

  8. The dynamic effect of muscle activation on knee stiffness.

    PubMed

    Ludvig, Daniel; Perreault, Eric J

    2014-01-01

    Adapting limb mechanics in a task and environment dependent manner is one component of human motor control. Joint mechanics have been extensively studied under static postural conditions, but less so under time-varying movement conditions. The limited studies that have investigated joint mechanics during movement, have found a drop in joint stiffness during movement, however the source of this decrease in stiffness remains unknown. Here in this paper we investigate whether time-varying muscle activation, which occurs during volitional movement, can lead to the drop in stiffness seen during movement. We found that under time-varying isometric conditions stiffness dropped when subjects transitioned from extension to flexion and vice-versa, a phenomenon that could not be explained by simply superimposing extension and flexion contractions. These findings suggest that dynamics of muscle activation may be responsible for the complex pattern of stiffness changes seen during simple movements. Furthermore, these results imply that EMG-based estimates of stiffness, which work well for steady-state postural conditions, will need to be augmented to account for the highly non-linear relationship between muscle activation and stiffness before they can also be used to estimate stiffness during dynamic contractions.

  9. An undergraduate laboratory activity on molecular dynamics simulations.

    PubMed

    Spitznagel, Benjamin; Pritchett, Paige R; Messina, Troy C; Goadrich, Mark; Rodriguez, Juan

    2016-01-01

    Vision and Change [AAAS, 2011] outlines a blueprint for modernizing biology education by addressing conceptual understanding of key concepts, such as the relationship between structure and function. The document also highlights skills necessary for student success in 21st century Biology, such as the use of modeling and simulation. Here we describe a laboratory activity that allows students to investigate the dynamic nature of protein structure and function through the use of a modeling technique known as molecular dynamics (MD). The activity takes place over two lab periods that are 3 hr each. The first lab period unpacks the basic approach behind MD simulations, beginning with the kinematic equations that all bioscience students learn in an introductory physics course. During this period students are taught rudimentary programming skills in Python while guided through simple modeling exercises that lead up to the simulation of the motion of a single atom. In the second lab period students extend concepts learned in the first period to develop skills in the use of expert MD software. Here students simulate and analyze changes in protein conformation resulting from temperature change, solvation, and phosphorylation. The article will describe how these activities can be carried out using free software packages, including Abalone and VMD/NAMD.

  10. Dynamical activities of primary somatosensory cortices studied by magnetoencephalography

    NASA Astrophysics Data System (ADS)

    Kishida, Kuniharu

    2009-11-01

    A blind identification method of transfer functions in feedback systems is introduced for examination of dynamical activities of cortices by magnetoencephalography study. Somatosensory activities are examined in 5 Hz periodical median nerve stimulus. In the present paper, we will try two careful preprocessing procedures for the identification method to obtain impulse responses between primary somatosensory cortices. Time series data of the somatosensory evoked field are obtained by using a blind source separation of the T/k type (fractional) decorrelation method. Time series data of current dipoles of primary somatosensory cortices are transformed from the time series data of the somatosensory evoked field by the inverse problem. Fluctuations of current dipoles of them are obtained after elimination of deterministic periodical evoked waveforms. An identification method based on feedback system theory is used for estimation of transfer functions in a feedback model from obtained fluctuations of currents dipoles of primary somatosensory cortices. Dynamical activities between them are presented by Bode diagrams of transfer functions and their impulse responses: the time delay of about 30 ms via corpus callosum is found in the impulse response of identified transfer function.

  11. An undergraduate laboratory activity on molecular dynamics simulations.

    PubMed

    Spitznagel, Benjamin; Pritchett, Paige R; Messina, Troy C; Goadrich, Mark; Rodriguez, Juan

    2016-01-01

    Vision and Change [AAAS, 2011] outlines a blueprint for modernizing biology education by addressing conceptual understanding of key concepts, such as the relationship between structure and function. The document also highlights skills necessary for student success in 21st century Biology, such as the use of modeling and simulation. Here we describe a laboratory activity that allows students to investigate the dynamic nature of protein structure and function through the use of a modeling technique known as molecular dynamics (MD). The activity takes place over two lab periods that are 3 hr each. The first lab period unpacks the basic approach behind MD simulations, beginning with the kinematic equations that all bioscience students learn in an introductory physics course. During this period students are taught rudimentary programming skills in Python while guided through simple modeling exercises that lead up to the simulation of the motion of a single atom. In the second lab period students extend concepts learned in the first period to develop skills in the use of expert MD software. Here students simulate and analyze changes in protein conformation resulting from temperature change, solvation, and phosphorylation. The article will describe how these activities can be carried out using free software packages, including Abalone and VMD/NAMD. PMID:26751047

  12. On the Dynamics of the Spontaneous Activity in Neuronal Networks

    PubMed Central

    Bonifazi, Paolo; Ruaro, Maria Elisabetta; Torre, Vincent

    2007-01-01

    Most neuronal networks, even in the absence of external stimuli, produce spontaneous bursts of spikes separated by periods of reduced activity. The origin and functional role of these neuronal events are still unclear. The present work shows that the spontaneous activity of two very different networks, intact leech ganglia and dissociated cultures of rat hippocampal neurons, share several features. Indeed, in both networks: i) the inter-spike intervals distribution of the spontaneous firing of single neurons is either regular or periodic or bursting, with the fraction of bursting neurons depending on the network activity; ii) bursts of spontaneous spikes have the same broad distributions of size and duration; iii) the degree of correlated activity increases with the bin width, and the power spectrum of the network firing rate has a 1/f behavior at low frequencies, indicating the existence of long-range temporal correlations; iv) the activity of excitatory synaptic pathways mediated by NMDA receptors is necessary for the onset of the long-range correlations and for the presence of large bursts; v) blockage of inhibitory synaptic pathways mediated by GABAA receptors causes instead an increase in the correlation among neurons and leads to a burst distribution composed only of very small and very large bursts. These results suggest that the spontaneous electrical activity in neuronal networks with different architectures and functions can have very similar properties and common dynamics. PMID:17502919

  13. Multivariable Dynamic Ankle Mechanical Impedance With Active Muscles

    PubMed Central

    Lee, Hyunglae; Krebs, Hermano Igo; Hogan, Neville

    2015-01-01

    Multivariable dynamic ankle mechanical impedance in two coupled degrees-of-freedom (DOFs) was quantified when muscles were active. Measurements were performed at five different target activation levels of tibialis anterior and soleus, from 10% to 30% of maximum voluntary contraction (MVC) with increments of 5% MVC. Interestingly, several ankle behaviors characterized in our previous study of the relaxed ankle were observed with muscles active: ankle mechanical impedance in joint coordinates showed responses largely consistent with a second-order system consisting of inertia, viscosity, and stiffness; stiffness was greater in the sagittal plane than in the frontal plane at all activation conditions for all subjects; and the coupling between dorsiflexion–plantarflexion and inversion–eversion was small—the two DOF measurements were well explained by a strictly diagonal impedance matrix. In general, ankle stiffness increased linearly with muscle activation in all directions in the 2-D space formed by the sagittal and frontal planes, but more in the sagittal than in the frontal plane, resulting in an accentuated “peanut shape.” This characterization of young healthy subjects’ ankle mechanical impedance with active muscles will serve as a baseline to investigate pathophysiological ankle behaviors of biomechanically and/or neurologically impaired patients. PMID:25203497

  14. Sequences flanking the core-binding site modulate glucocorticoid receptor structure and activity.

    PubMed

    Schöne, Stefanie; Jurk, Marcel; Helabad, Mahdi Bagherpoor; Dror, Iris; Lebars, Isabelle; Kieffer, Bruno; Imhof, Petra; Rohs, Remo; Vingron, Martin; Thomas-Chollier, Morgane; Meijsing, Sebastiaan H

    2016-09-01

    The glucocorticoid receptor (GR) binds as a homodimer to genomic response elements, which have particular sequence and shape characteristics. Here we show that the nucleotides directly flanking the core-binding site, differ depending on the strength of GR-dependent activation of nearby genes. Our study indicates that these flanking nucleotides change the three-dimensional structure of the DNA-binding site, the DNA-binding domain of GR and the quaternary structure of the dimeric complex. Functional studies in a defined genomic context show that sequence-induced changes in GR activity cannot be explained by differences in GR occupancy. Rather, mutating the dimerization interface mitigates DNA-induced changes in both activity and structure, arguing for a role of DNA-induced structural changes in modulating GR activity. Together, our study shows that DNA sequence identity of genomic binding sites modulates GR activity downstream of binding, which may play a role in achieving regulatory specificity towards individual target genes.

  15. An overlapping kinase and phosphatase docking site regulates activity of the retinoblastoma protein.

    PubMed

    Hirschi, Alexander; Cecchini, Matthew; Steinhardt, Rachel C; Schamber, Michael R; Dick, Frederick A; Rubin, Seth M

    2010-09-01

    The phosphorylation state and corresponding activity of the retinoblastoma tumor suppressor protein (Rb) are modulated by a balance of kinase and phosphatase activities. Here we characterize the association of Rb with the catalytic subunit of protein phosphatase 1 (PP1c). A crystal structure identifies an enzyme docking site in the Rb C-terminal domain that is required for efficient PP1c activity toward Rb. The phosphatase docking site overlaps with the known docking site for cyclin-dependent kinase (Cdk), and PP1 competition with Cdk-cyclins for Rb binding is sufficient to retain Rb activity and block cell-cycle advancement. These results provide the first detailed molecular insights into Rb activation and establish a novel mechanism for Rb regulation in which kinase and phosphatase compete for substrate docking. PMID:20694007

  16. Computer modeling of large asteroid impacts into continental and oceanic sites: Atmospheric, cratering, and ejecta dynamics

    NASA Technical Reports Server (NTRS)

    Roddy, D. J.; Schuster, S. H.; Rosenblatt, M.; Grant, L. B.; Hassig, P. J.; Kreyenhagen, K. N.

    1988-01-01

    Numerous impact cratering events have occurred on the Earth during the last several billion years that have seriously affected our planet and its atmosphere. The largest cratering events, which were caused by asteroids and comets with kinetic energies equivalent to tens of millions of megatons of TNT, have distributed substantial quantities of terrestrial and extraterrestrial material over much or all of the Earth. In order to study a large-scale impact event in detail, computer simulations were completed that model the passage of a 10 km-diameter asteroid through the Earth's atmosphere and the subsequent cratering and ejecta dynamics associated with impact of the asteroid into two different targets, i.e., an oceanic site and a continental site. The calcuations were designed to broadly represent giant impact events that have occurred on the Earth since its formation and specifically represent an impact cratering event proposed to have occurred at the end of Cretaceous time. Calculation of the passage of the asteroid through a U.S. Standard Atmosphere showed development of a strong bow shock that expanded radially outward. Behind the shock front was a region of highly shock compressed and intensely heated air. Behind the asteroid, rapid expansion of this shocked air created a large region of very low density that also expanded away from the impact area. Calculations of the cratering events in both the continental and oceanic targets were carried to 120 s. Despite geologic differences, impacts in both targets developed comparable dynamic flow fields, and by approx. 29 s similar-sized transient craters approx. 39 km deep and approx. 62 km across had formed. For all practical purposes, the atmosphere was nearly completely removed from the impact area for tens of seconds, i.e., air pressures were less than fractions of a bar out to ranges of over 50 km. Consequently, much of the asteroid and target materials were ejected upward into a near vacuum. Effects of secondary

  17. Molecular dynamics study to identify the reactive sites of a liquid squalane surface.

    PubMed

    Köhler, Sven P K; Reed, Stewart K; Westacott, Robin E; McKendrick, Kenneth G

    2006-06-22

    Molecular dynamics simulations of liquid squalane, C30H62, were performed, focusing in particular on the liquid-vacuum interface. These theoretical studies were aimed at identifying potentially reactive sites on the surface, knowledge of which is important for a number of inelastic and reactive scattering experiments. A united atom force field (Martin, M. G.; Siepmann, J. I. J. Phys. Chem. B 1999, 103, 4508-4517) was used, and the simulations were analyzed with respect to their interfacial properties. A modest but clearly identifiable preference for methyl groups to protrude into the vacuum has been found at lower temperatures. This effect decreases when going to higher temperatures. Additional simulations tracking the flight paths of projectiles directed at a number of randomly chosen surfaces extracted from the molecular dynamics simulations were performed. The geometrical parameters for these calculations were chosen to imitate a typical abstraction reaction, such as the reaction between ground-state oxygen atoms and hydrocarbons. Despite the preference for methyl groups to protrude further into the vacuum, Monte Carlo tracking simulations suggest, on geometric grounds, that primary and secondary hydrogen atoms are roughly equally likely to react with incoming gas-phase atoms. These geometric simulations also indicate that a substantial fraction of the scattered products is likely to undergo at least one secondary collision with hydrocarbon side chains. These results help to interpret the outcome of previous measurements of the internal and external energy distribution of the gas-phase OH products of the interfacial reaction between oxygen atoms and liquid squalane.

  18. Dynamic regulation of Polycomb group activity during plant development.

    PubMed

    Bemer, Marian; Grossniklaus, Ueli

    2012-11-01

    Polycomb group (PcG) complexes play important roles in phase transitions and cell fate determination in plants and animals, by epigenetically repressing sets of genes that promote either proliferation or differentiation. The continuous differentiation of new organs in plants, such as leaves or flowers, requires a highly dynamic PcG function, which can be induced, modulated, or repressed when necessary. In this review, we discuss the recent advance in understanding PcG function in plants and focus on the diverse molecular mechanisms that have been described to regulate and counteract PcG activity in Arabidopsis.

  19. Dynamic regulation of Polycomb group activity during plant development.

    PubMed

    Bemer, Marian; Grossniklaus, Ueli

    2012-11-01

    Polycomb group (PcG) complexes play important roles in phase transitions and cell fate determination in plants and animals, by epigenetically repressing sets of genes that promote either proliferation or differentiation. The continuous differentiation of new organs in plants, such as leaves or flowers, requires a highly dynamic PcG function, which can be induced, modulated, or repressed when necessary. In this review, we discuss the recent advance in understanding PcG function in plants and focus on the diverse molecular mechanisms that have been described to regulate and counteract PcG activity in Arabidopsis. PMID:22999383

  20. Structural and Kinetic Analyses of Macrophage Migration Inhibitory Factor Active Site Interactions

    SciTech Connect

    Crichlow, G.; Lubetsky, J; Leng, L; Bucala, R; Lolis, E

    2009-01-01

    Macrophage migration inhibitory factor (MIF) is a secreted protein expressed in numerous cell types that counters the antiinflammatory effects of glucocorticoids and has been implicated in sepsis, cancer, and certain autoimmune diseases. Interestingly, the structure of MIF contains a catalytic site resembling the tautomerase/isomerase sites of microbial enzymes. While bona fide physiological substrates remain unknown, model substrates have been identified. Selected compounds that bind in the tautomerase active site also inhibit biological functions of MIF. It had previously been shown that the acetaminophen metabolite, N-acetyl-p-benzoquinone imine (NAPQI), covalently binds to the active site of MIF. In this study, kinetic data indicate that NAPQI inhibits MIF both covalently and noncovalently. The structure of MIF cocrystallized with NAPQI reveals that the NAPQI has undergone a chemical alteration forming an acetaminophen dimer (bi-APAP) and binds noncovalently to MIF at the mouth of the active site. We also find that the commonly used protease inhibitor, phenylmethylsulfonyl fluoride (PMSF), forms a covalent complex with MIF and inhibits the tautomerase activity. Crystallographic analysis reveals the formation of a stable, novel covalent bond for PMSF between the catalytic nitrogen of the N-terminal proline and the sulfur of PMSF with complete, well-defined electron density in all three active sites of the MIF homotrimer. Conclusions are drawn from the structures of these two MIF-inhibitor complexes regarding the design of novel compounds that may provide more potent reversible and irreversible inhibition of MIF.

  1. Modeling colony site dynamics: a case study of gull-billed terns (Sterna nilotica) in coastal Virginia

    USGS Publications Warehouse

    Erwin, R.M.; Nichols, J.D.; Eyler, T.B.; Stotts, D.B.; Truitt, B.R.

    1998-01-01

    We developed a Markov process model for colony site dynamics of Gull-billed Terns (Sterna nilotica) in coastal Virginia. We used the model and data on colony site occupation from 1993 to 1996 to estimate model parameters. Each year, we monitored the breeding numbers of Gull-billed Terns and their frequent colony associates, Common Terns (Sterna hirundo) and Black Skimmers (Rynchops niger) at colony sites along about 80 km of the barrier island region of Virginia. We also monitored flooding events and renesting. We developed the model for colony survival, extinction, and recolonization at potential colony sites over the four-year period. We then used data on annual site occupation by Gull-billed Terns to estimate model parameters and test between different structures reflecting competing hypotheses. Results revealed a dynamic system, but provided no evidence that the dynamics were Markovian , i.e. the probability of occupancy of a site in one year was not influenced by whether it had been occupied the previous year. Nor did the colony-level reproductive success the previous season seem to affect the probability of site occupancy. Site survival and recolonization rates were similar, and the overall annual probability of a site being occupied over the course of the four-year period was estimated to be 0.59 Of the total of 25 sites that were used during the four-year period, 16 were used in only one or two years while only three were used all four years.. Flooding and renesting were frequent in both habitat types in all years. The frequent flooding of nests on shellpiles argues for more effective management; augmentation with shell and sand to increase elevations as little as 20 cm could have reduced flooding at a number of sites. The low colony-site fidelity we demonstrate suggests that an effective management approach is to provide a large number of alternative sand and/or shellpile sites that the terns may use. Sites not used one year may still be used in subsequent

  2. Flexibility Correlation between Active Site Regions Is Conserved across Four AmpC β-Lactamase Enzymes

    PubMed Central

    Brown, Jenna R.; Livesay, Dennis R.

    2015-01-01

    β-lactamases are bacterial enzymes that confer resistance to β-lactam antibiotics, such as penicillins and cephalosporins. There are four classes of β-lactamase enzymes, each with characteristic sequence and structure properties. Enzymes from class A are the most common and have been well characterized across the family; however, less is known about how physicochemical properties vary across the C and D families. In this report, we compare the dynamical properties of four AmpC (class C) β-lactamases using our distance constraint model (DCM). The DCM reliably predicts thermodynamic and mechanical properties in an integrated way. As a consequence, quantitative stability/flexibility relationships (QSFR) can be determined and compared across the whole family. The DCM calculates a large number of QSFR metrics. Perhaps the most useful is the flexibility index (FI), which quantifies flexibility along the enzyme backbone. As typically observed in other systems, FI is well conserved across the four AmpC enzymes. Cooperativity correlation (CC), which quantifies intramolecular couplings within structure, is rarely conserved across protein families; however, it is in AmpC. In particular, the bulk of each structure is composed of a large rigid cluster, punctuated by three flexibly correlated regions located at the active site. These regions include several catalytic residues and the Ω-loop. This evolutionary conservation combined with active their site location strongly suggests that these coupled dynamical modes are important for proper functioning of the enzyme. PMID:26018804

  3. Adsorption dynamics of trichlorofluoromethane in activated carbon fiber beds.

    PubMed

    Zhang, Xiaoping; Zhao, Xin; Hu, Jiaqi; Wei, Chaohai; Bi, Hsiaotao T

    2011-02-28

    Adsorption on carbon fixed-beds is considered as an inexpensive and highly effective way for controlling chlorofluorocarbons (CFCs) emissions. In the present work, a dynamic model under constant-pattern wave conditions has been developed to predict the breakthrough behavior of trichlorofluoromethane (CFC-11) adsorption in a fixed bed packed with activated carbon fibers (ACFs). The adsorption of CFC-11 vapor onto viscose-based ACFs was performed in a fixed bed at different test conditions. The results showed that, in a deep bed (>120 mm), the analytical model based on the external mass transfer with the Langmuir isotherm could describe the adsorption dynamics well. The model parameters, the characteristic breakthrough time and the film mass-transfer coefficients are related to such operating parameters as the superficial gas velocity, feed concentration and bed height. It was found from the breakthrough dynamics that the mass transfer from the fluid phase to the fiber surface dominated the CFC-11 adsorption onto ACFs in fixed beds.

  4. Adsorption dynamics of trichlorofluoromethane in activated carbon fiber beds.

    PubMed

    Zhang, Xiaoping; Zhao, Xin; Hu, Jiaqi; Wei, Chaohai; Bi, Hsiaotao T

    2011-02-28

    Adsorption on carbon fixed-beds is considered as an inexpensive and highly effective way for controlling chlorofluorocarbons (CFCs) emissions. In the present work, a dynamic model under constant-pattern wave conditions has been developed to predict the breakthrough behavior of trichlorofluoromethane (CFC-11) adsorption in a fixed bed packed with activated carbon fibers (ACFs). The adsorption of CFC-11 vapor onto viscose-based ACFs was performed in a fixed bed at different test conditions. The results showed that, in a deep bed (>120 mm), the analytical model based on the external mass transfer with the Langmuir isotherm could describe the adsorption dynamics well. The model parameters, the characteristic breakthrough time and the film mass-transfer coefficients are related to such operating parameters as the superficial gas velocity, feed concentration and bed height. It was found from the breakthrough dynamics that the mass transfer from the fluid phase to the fiber surface dominated the CFC-11 adsorption onto ACFs in fixed beds. PMID:21216098

  5. Mapping the conformational transition in Src activation by cumulating the information from multiple molecular dynamics trajectories

    PubMed Central

    Yang, Sichun; Banavali, Nilesh K.; Roux, Benoît

    2009-01-01

    The Src-family kinases are allosteric enzymes that play a key role in the regulation of cell growth and proliferation. In response to cellular signals, they undergo large conformational changes to switch between distinct inactive and active states. A computational strategy for characterizing the conformational transition pathway is presented to bridge the inactive and active states of the catalytic domain of Hck. The information from a large number (78) of independent all-atom molecular dynamics trajectories with explicit solvent is combined together to assemble a connectivity map of the conformational transition. Two intermediate states along the activation pathways are identified, and their structural features are characterized. A coarse free-energy landscape is built in terms of the collective motions corresponding to the opening of the activation loop (A-loop) and the rotation of the αC helix. This landscape shows that the protein can adopt a multitude of conformations in which the A-loop is partially open, while the αC helix remains in the orientation characteristic of the inactive conformation. The complete transition leading to the active conformation requires a concerted movement involving further opening of the A-loop, the relative alignment of N-lobe and C-lobe, and the rotation of the αC helix needed to recruit the residues necessary for catalysis in the active site. The analysis leads to a dynamic view of the full-length kinase activation, whereby transitions of the catalytic domain to intermediate configurations with a partially open A-loop are permitted, even while the SH2-SH3 clamp remains fully engaged. These transitions would render Y416 available for the transphosphorylation event that ultimately locks down the active state. The results provide a broad framework for picturing the conformational transitions leading to kinase activation. PMID:19225111

  6. Broken Detailed Balance of Filament Dynamics in Active Networks

    NASA Astrophysics Data System (ADS)

    Gladrow, J.; Fakhri, N.; MacKintosh, F. C.; Schmidt, C. F.; Broedersz, C. P.

    2016-06-01

    Myosin motor proteins drive vigorous steady-state fluctuations in the actin cytoskeleton of cells. Endogenous embedded semiflexible filaments such as microtubules, or added filaments such as single-walled carbon nanotubes are used as novel tools to noninvasively track equilibrium and nonequilibrium fluctuations in such biopolymer networks. Here, we analytically calculate shape fluctuations of semiflexible probe filaments in a viscoelastic environment, driven out of equilibrium by motor activity. Transverse bending fluctuations of the probe filaments can be decomposed into dynamic normal modes. We find that these modes no longer evolve independently under nonequilibrium driving. This effective mode coupling results in nonzero circulatory currents in a conformational phase space, reflecting a violation of detailed balance. We present predictions for the characteristic frequencies associated with these currents and investigate how the temporal signatures of motor activity determine mode correlations, which we find to be consistent with recent experiments on microtubules embedded in cytoskeletal networks.

  7. A nonlinear model of the phasic dynamics of muscle activation

    NASA Technical Reports Server (NTRS)

    Hannaford, Blake

    1990-01-01

    A phasic excitation-activation (PEXA) model is presented of the process of motoneuron excitation and the resultant activation and force development of a motor unit. The model input is an amount of depolarizing current (as when injected with an intracellular electrode), and the model output is muscle force. The model includes dynamics and nonlinearities similar to phenomena discovered experimentally by others: the firing rate response of motoneurons to steps of depolarizing current and the catch-like enhancement of force produced by overlapping motor neuron action potentials. The parameter values used in this model are derived from experimentally measured data and are expressed in physical units. Model predictions extend to published data beyond those used in generating the model parameter values.

  8. Using Carbohydrate Interaction Assays to Reveal Novel Binding Sites in Carbohydrate Active Enzymes.

    PubMed

    Cockburn, Darrell; Wilkens, Casper; Dilokpimol, Adiphol; Nakai, Hiroyuki; Lewińska, Anna; Abou Hachem, Maher; Svensson, Birte

    2016-01-01

    Carbohydrate active enzymes often contain auxiliary binding sites located either on independent domains termed carbohydrate binding modules (CBMs) or as so-called surface binding sites (SBSs) on the catalytic module at a certain distance from the active site. The SBSs are usually critical for the activity of their cognate enzyme, though they are not readily detected in the sequence of a protein, but normally require a crystal structure of a complex for their identification. A variety of methods, including affinity electrophoresis (AE), insoluble polysaccharide pulldown (IPP) and surface plasmon resonance (SPR) have been used to study auxiliary binding sites. These techniques are complementary as AE allows monitoring of binding to soluble polysaccharides, IPP to insoluble polysaccharides and SPR to oligosaccharides. Here we show that these methods are useful not only for analyzing known binding sites, but also for identifying new ones, even without structural data available. We further verify the chosen assays discriminate between known SBS/CBM containing enzymes and negative controls. Altogether 35 enzymes are screened for the presence of SBSs or CBMs and several novel binding sites are identified, including the first SBS ever reported in a cellulase. This work demonstrates that combinations of these methods can be used as a part of routine enzyme characterization to identify new binding sites and advance the study of SBSs and CBMs, allowing them to be detected in the absence of structural data. PMID:27504624

  9. Using Carbohydrate Interaction Assays to Reveal Novel Binding Sites in Carbohydrate Active Enzymes

    PubMed Central

    Wilkens, Casper; Dilokpimol, Adiphol; Nakai, Hiroyuki; Lewińska, Anna; Abou Hachem, Maher; Svensson, Birte

    2016-01-01

    Carbohydrate active enzymes often contain auxiliary binding sites located either on independent domains termed carbohydrate binding modules (CBMs) or as so-called surface binding sites (SBSs) on the catalytic module at a certain distance from the active site. The SBSs are usually critical for the activity of their cognate enzyme, though they are not readily detected in the sequence of a protein, but normally require a crystal structure of a complex for their identification. A variety of methods, including affinity electrophoresis (AE), insoluble polysaccharide pulldown (IPP) and surface plasmon resonance (SPR) have been used to study auxiliary binding sites. These techniques are complementary as AE allows monitoring of binding to soluble polysaccharides, IPP to insoluble polysaccharides and SPR to oligosaccharides. Here we show that these methods are useful not only for analyzing known binding sites, but also for identifying new ones, even without structural data available. We further verify the chosen assays discriminate between known SBS/CBM containing enzymes and negative controls. Altogether 35 enzymes are screened for the presence of SBSs or CBMs and several novel binding sites are identified, including the first SBS ever reported in a cellulase. This work demonstrates that combinations of these methods can be used as a part of routine enzyme characterization to identify new binding sites and advance the study of SBSs and CBMs, allowing them to be detected in the absence of structural data. PMID:27504624

  10. Static and dynamic activity of warm receptors in Boa constrictor.

    PubMed

    Hensel, H

    1975-01-01

    Afferent impulses from multi- and single-fiber preparations of the trigeminal nerve in Boa constrictor were recorded during exactly controlled thermal stimulation of the receptive field in the labial region. At constant temperatures in the range between 18 and 37 degrees C, multi-fiber preparations showed a continuous discharge with a maximum around 30 degrees C. Dynamic warming caused a high increase of the discharge, whereas dynamic cooling led to a complete inhibition. No cold-sensitive fivers have been found. Mechanical stimulation elicited large spikes from specific mechanoreceptors. Single-fiber preparations from labial warm receptors did not respond to mechanical stimulation. Their discharge was always regular at constant temperatures. The average frequency of a warm receptor population was zero at about 18 degrees C, reached a maximum of 13 sec-1 at 30 degrees C and fell again to zero at 37 degrees C. In addition, a few warm receptors increased their static discharge with temperature up to 36 degrees C, the highest frequency being 38 sec-1. Stepwise warming by delta T = + 5 degrees C caused a marked overshoot in frequency, after which the discharge usually fell to a minimum and then rose again to a new static level. Stepwise cooling by delta T = MINUS 5 DEGREES C led to a transient inhibition of activity followed by an increase until the new static level was reached. In the first group of warm receptors the height of the dynamic overshoot varied with the adapting temperature, the largest average overshoot of 160 sec-1 occurring at an adapting temperature of 30 degrees C. These receptors have their static maximum as well as their highest dynamic sensitivity in the temperature range of the natural tropical habitat of Boidae.

  11. 40 CFR 61.154 - Standard for active waste disposal sites.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... (20″×14″) upright format signs specified in 29 CFR 1910.145(d)(4) and this paragraph; and (iii... 40 Protection of Environment 8 2011-07-01 2011-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an...

  12. 40 CFR 61.154 - Standard for active waste disposal sites.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... (20″×14″) upright format signs specified in 29 CFR 1910.145(d)(4) and this paragraph; and (iii... 40 Protection of Environment 8 2010-07-01 2010-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an...

  13. 40 CFR 61.154 - Standard for active waste disposal sites.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... (20″×14″) upright format signs specified in 29 CFR 1910.145(d)(4) and this paragraph; and (iii... 40 Protection of Environment 9 2013-07-01 2013-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an...

  14. 77 FR 39508 - Commercial Wind Lease Issuance and Site Assessment Activities on the Atlantic Outer Continental...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-03

    ... specific project proposals on those leases) in an identified Wind Energy Area (WEA) on the OCS offshore... Bureau of Ocean Energy Management Commercial Wind Lease Issuance and Site Assessment Activities on the... Activities on the Atlantic OCS Offshore RI and MA'' to: Program Manager, Office of Renewable Energy...

  15. Effects of resource activities upon repository siting and waste containment with reference to bedded salt

    SciTech Connect

    Ashby, J.; Rowe, J.

    1980-02-01

    The primary consideration for the suitability of a nuclear waste repository site is the overall ability of the repository to safely contain radioactive waste. This report is a discussion of the past, present, and future effects of resource activities on waste containment. Past and present resource activities which provide release pathways (i.e., leaky boreholes, adjacent mines) will receive initial evaluation during the early stages of any repository site study. However, other resource activities which may have subtle effects on containment (e.g., long-term pumping causing increased groundwater gradients, invasion of saline water causing lower retardation) and all potential future resource activities must also be considered during the site evaluation process. Resource activities will affect both the siting and the designing of repositories. Ideally, sites should be located in areas of low resource activity and low potential for future activity, and repository design should seek to eliminate or minimize the adverse effects of any resource activity. Buffer zones should be created to provide areas in which resource activities that might adversely affect containment can be restricted or curtailed. This could mean removing large areas of land from resource development. The impact of these frozen assets should be assessed in terms of their economic value and of their effect upon resource reserves. This step could require a major effort in data acquisition and analysis followed by extensive numerical modeling of regional fluid flow and mass transport. Numerical models should be used to assess the effects of resource activity upon containment and should include the cumulative effects of different resource activities. Analysis by other methods is probably not possible except for relatively simple cases.

  16. Computational approaches to the determination of active site structures and reaction mechanisms in heterogeneous catalysts.

    PubMed

    Catlow, C R A; French, S A; Sokol, A A; Thomas, J M

    2005-04-15

    We apply quantum chemical methods to the study of active site structures and reaction mechanisms in mesoporous silica and metal oxide catalysts. Our approach is based on the use of both molecular cluster and embedded cluster (QM/MM) techniques, where the active site and molecular complex are described using density functional theory (DFT) and the embedding matrix simulated by shell model potentials. We consider three case studies: alkene epoxidation over the microporous TS-1 catalyst; methanol synthesis on ZnO and Cu/ZnO and C-H bond activation over Li-doped MgO.

  17. Computational approaches to the determination of active site structures and reaction mechanisms in heterogeneous catalysts.

    PubMed

    Catlow, C R A; French, S A; Sokol, A A; Thomas, J M

    2005-04-15

    We apply quantum chemical methods to the study of active site structures and reaction mechanisms in mesoporous silica and metal oxide catalysts. Our approach is based on the use of both molecular cluster and embedded cluster (QM/MM) techniques, where the active site and molecular complex are described using density functional theory (DFT) and the embedding matrix simulated by shell model potentials. We consider three case studies: alkene epoxidation over the microporous TS-1 catalyst; methanol synthesis on ZnO and Cu/ZnO and C-H bond activation over Li-doped MgO. PMID:15901543

  18. Dynamics of Active Separation Control at High Reynolds Numbers

    NASA Technical Reports Server (NTRS)

    Pack, LaTunia G.; Seifert, Avi

    2000-01-01

    A series of active flow control experiments were recently conducted at high Reynolds numbers on a generic separated configuration. The model simulates the upper surface of a 20% thick Glauert-Goldschmied type airfoil at zero angle of attack. The flow is fully turbulent since the tunnel sidewall boundary layer flows over the model. The main motivation for the experiments is to generate a comprehensive data base for validation of unsteady numerical simulation as a first step in the development of a CFD design tool, without which it would not be possible to effectively utilize the great potential of unsteady flow control. This paper focuses on the dynamics of several key features of the baseline as well as the controlled flow. It was found that the thickness of the upstream boundary layer has a negligible effect on the flow dynamics. It is speculated that separation is caused mainly by the highly convex surface while viscous effects are less important. The two-dimensional separated flow contains unsteady waves centered on a reduced frequency of 0.8, while in the three dimensional separated flow, frequencies around a reduced frequency of 0.3 and 1 are active. Several scenarios of resonant wave interaction take place at the separated shear-layer and in the pressure recovery region. The unstable reduced frequency bands for periodic excitation are centered on 1.5 and 5, but these reduced frequencies are based on the length of the baseline bubble that shortens due to the excitation. The conventional swept wing-scaling works well for the coherent wave features. Reproduction of these dynamic effects by a numerical simulation would provide benchmark validation.

  19. Dynamic positioning system based on active disturbance rejection technology

    NASA Astrophysics Data System (ADS)

    Lei, Zhengling; Guo, Chen; Fan, Yunsheng

    2015-08-01

    A dynamically positioned vessel, by the International Maritime Organization (IMO) and the certifying class societies (DNV, ABS, LR, etc.), is defined as a vessel that maintains its position and heading (fixed location or pre-determined track) exclusively by means of active thrusters. The development of control technology promotes the upgrading of dynamic positioning (DP) systems. Today there are two different DP systems solutions available on the market: DP system based on PID regulator and that based on model-based control. Both systems have limited disturbance rejection capability due to their design principle. In this paper, a new DP system solution is proposed based on Active Disturbance Rejection Control (ADRC) technology. This technology is composed of Tracking-Differentiator (TD), Extended State Observer (ESO) and Nonlinear Feedback Combination. On one hand, both TD and ESO can act as filters and can be used in place of conventional filters; on the other hand, the total disturbance of the system can be estimated and compensated by ESO, which therefore enhances the system's disturbance rejection capability. This technology's advantages over other methods lie in two aspects: 1) This method itself can not only achieve control objectives but also filter noisy measurements without other specialized filters; 2) This method offers a new useful approach to suppress the ocean disturbance. The simulation results demonstrate the effectiveness of the proposed method.

  20. Molecular Dynamics of "Fuzzy" Transcriptional Activator-Coactivator Interactions

    PubMed Central

    Scholes, Natalie S.; Weinzierl, Robert O. J.

    2016-01-01

    Transcriptional activation domains (ADs) are generally thought to be intrinsically unstructured, but capable of adopting limited secondary structure upon interaction with a coactivator surface. The indeterminate nature of this interface made it hitherto difficult to study structure/function relationships of such contacts. Here we used atomistic accelerated molecular dynamics (aMD) simulations to study the conformational changes of the GCN4 AD and variants thereof, either free in solution, or bound to the GAL11 coactivator surface. We show that the AD-coactivator interactions are highly dynamic while obeying distinct rules. The data provide insights into the constant and variable aspects of orientation of ADs relative to the coactivator, changes in secondary structure and energetic contributions stabilizing the various conformers at different time points. We also demonstrate that a prediction of α-helical propensity correlates directly with the experimentally measured transactivation potential of a large set of mutagenized ADs. The link between α-helical propensity and the stimulatory activity of ADs has fundamental practical and theoretical implications concerning the recruitment of ADs to coactivators. PMID:27175900

  1. Dynamic Stimuli And Active Processing In Human Visual Perception

    NASA Astrophysics Data System (ADS)

    Haber, Ralph N.

    1990-03-01

    Theories of visual perception traditionally have considered a static retinal image to be the starting point for processing; and has considered processing both to be passive and a literal translation of that frozen, two dimensional, pictorial image. This paper considers five problem areas in the analysis of human visually guided locomotion, in which the traditional approach is contrasted to newer ones that utilize dynamic definitions of stimulation, and an active perceiver: (1) differentiation between object motion and self motion, and among the various kinds of self motion (e.g., eyes only, head only, whole body, and their combinations); (2) the sources and contents of visual information that guide movement; (3) the acquisition and performance of perceptual motor skills; (4) the nature of spatial representations, percepts, and the perceived layout of space; and (5) and why the retinal image is a poor starting point for perceptual processing. These newer approaches argue that stimuli must be considered as dynamic: humans process the systematic changes in patterned light when objects move and when they themselves move. Furthermore, the processing of visual stimuli must be active and interactive, so that perceivers can construct panoramic and stable percepts from an interaction of stimulus information and expectancies of what is contained in the visual environment. These developments all suggest a very different approach to the computational analyses of object location and identification, and of the visual guidance of locomotion.

  2. Cyanide does more to inhibit heme enzymes, than merely serving as an active-site ligand

    SciTech Connect

    Parashar, Abhinav; Venkatachalam, Avanthika; Gideon, Daniel Andrew; Manoj, Kelath Murali

    2014-12-12

    Highlights: • Cyanide (CN) is a well-studied toxic principle, known to inhibit heme-enzymes. • Inhibition is supposed to result from CN binding at the active site as a ligand. • Diverse heme enzymes’ CN inhibition profiles challenge prevailing mechanism. • Poor binding efficiency of CN at low enzyme concentrations and ligand pressures. • CN-based diffusible radicals cause ‘non-productive electron transfers’ (inhibition). - Abstract: The toxicity of cyanide is hitherto attributed to its ability to bind to heme proteins’ active site and thereby inhibit their activity. It is shown herein that the long-held interpretation is inadequate to explain several observations in heme-enzyme reaction systems. Generation of cyanide-based diffusible radicals in heme-enzyme reaction milieu could shunt electron transfers (by non-active site processes), and thus be detrimental to the efficiency of oxidative outcomes.

  3. Denaturation studies of active-site labeled papain using electron paramagnetic resonance and fluorescence spectroscopy.

    PubMed Central

    Ping, Z A; Butterfiel, D A

    1991-01-01

    A spin-labeled p-chloromercuribenzoate (SL-PMB) and a fluorescence probe, 6-acryloyl-2-dimethylaminonaphthalene (Acrylodan), both of which bind to the single SH group located in the active site of papain, were used to investigate the interaction of papain (EC 3.4.22.2) with two protein denaturants. It was found that the active site of papain was highly stable in urea solution, but underwent a large conformational change in guanidine hydrochloride solution. Electron paramagnetic resonance and fluorescence results were in agreement and both paralleled enzymatic activity of papain with respect to both the variation in pH and denaturation. These results strongly suggest that SL-PMB and Acrylodan labels can be used to characterize the physical state of the active site of the enzyme. PMID:1657229

  4. Catalysis-dependent selenium incorporation and migration in the nitrogenase active site iron-molybdenum cofactor

    PubMed Central

    Spatzal, Thomas; Perez, Kathryn A; Howard, James B; Rees, Douglas C

    2015-01-01

    Dinitrogen reduction in the biological nitrogen cycle is catalyzed by nitrogenase, a two-component metalloenzyme. Understanding of the transformation of the inert resting state of the active site FeMo-cofactor into an activated state capable of reducing dinitrogen remains elusive. Here we report the catalysis dependent, site-selective incorporation of selenium into the FeMo-cofactor from selenocyanate as a newly identified substrate and inhibitor. The 1.60 Å resolution structure reveals selenium occupying the S2B site of FeMo-cofactor in the Azotobacter vinelandii MoFe-protein, a position that was recently identified as the CO-binding site. The Se2B-labeled enzyme retains substrate reduction activity and marks the starting point for a crystallographic pulse-chase experiment of the active site during turnover. Through a series of crystal structures obtained at resolutions of 1.32–1.66 Å, including the CO-inhibited form of Av1-Se2B, the exchangeability of all three belt-sulfur sites is demonstrated, providing direct insights into unforeseen rearrangements of the metal center during catalysis. DOI: http://dx.doi.org/10.7554/eLife.11620.001 PMID:26673079

  5. Catalysis-dependent selenium incorporation and migration in the nitrogenase active site iron-molybdenum cofactor.