Conformational Dynamics of the Human Propeller Telomeric DNA Quadruplex on a Microsecond Time Scale

The human telomeric DNA sequence with four repeats can fold into a parallel-stranded propeller-type topology. NMR structures solved under molecular crowding experiments correlate with the crystal structures found with crystal-packing interactions that are effectively equivalent to molecular crowding. This topology has been used for rationalization of ligand design and occurs experimentally in a number of complexes with a diversity of ligands, at least in the crystalline state. While G-quartet stems have been well characterised, the interactions of the TTA loop with the G-quartets are much less defined. To better understand the conformational variability and structural dynamics of the propeller-type topology, we performed molecular dynamics simulations in explicit solvent up to 1.5 µs. The analysis provides a detailed atomistic account of the dynamic nature of the TTA loops highlighting their interactions with the G-quartets including formation of an A:A base pair, triad, pentad and hexad. The results present a threshold in quadruplex simulations, with regards to understanding the flexible nature of the sugar-phosphate backbone in formation of unusual architecture within the topology. Furthermore, this study stresses the importance of simulation time in sampling conformational space for this topology.

Mode of cembranoid action on embryonic muscle acetylcholine receptor

The mechanism of eupalmerin acetate (EUAC) actions on the embryonic muscle nicotinic acetylcholine receptor (nAChR) in BC3H-1 cells was studied by using whole-cell and single-channel patch-clamp current measurements. With whole-cell currents, EUAC did not act as an agonist on this receptor. Coapplication of 30 mu M EUAC with 50 mu M, 100 N, or 500 mu M carbamoylcholine (CCh) reversibly inhibited the current amplitude, whereas, with 20 mu M CCh, current was increased above control values in the presence of EUAC. EUAC concentration curves (0.01-40 N) obtained with 100 mu M and 500 mu M CCh displayed slope coefficients, n(H), significantly smaller than one, suggesting that EUAC bound to several sites with widely differing affinities on the receptor molecule. The apparent rate of receptor desensitization in the presence of EUAC and CCh was either slower than or equal to that obtained with CCh alone. The major finding from single-channel studies was that EUAC did not affect single-channel conductance or the ability of CCh to interact with the receptor. Instead, EUAC acted by increasing the channel closing rate constant. The results are not consistent with the competitive model for EUAC inhibition, with the sequential open-channel block model, or with inhibition by increased desensitization. The data are best accounted for by a model in which EUAC acts by closed-channel block at low concentrations, by positive modulation at intermediate concentrations, and by negative allosteric modulation of the open channel at high concentrations. (c) 2007 Wiley-Liss, Inc.

Submillisecond events in protein folding.

The pathway of protein folding is now being analyzed at the resolution of individual residues by kinetic measurements on suitably engineered mutants. The kinetic methods generally employed for studying folding are typically limited to the time range of > or = 1 ms because the folding of denatured proteins is usually initiated by mixing them with buffers that favor folding, and the dead time of rapid mixing experiments is about a millisecond. We now show that the study of protein folding may be extended to the microsecond time region by using temperature-jump measurements on the cold-unfolded state of a suitable protein. We are able to detect early events in the folding of mutants of barstar, the polypeptide inhibitor of barnase. A preliminary characterization of the fast phase from spectroscopic and phi-value analysis indicates that it is a transition between two relatively solvent-exposed states with little consolidation of structure.

Entanglement and entropy operator for strings in pp-wave time dependent background

In this Letter new aspects of string theory propagating in a pp-wave time dependent background with a null singularity are explored. It is shown the appearance of a 2d entanglement entropy dynamically generated by the background. For asymptotically flat observers, the vacuum close to the singularity is unitarily inequivalent to the vacuum at tau = -infinity and it is shown that the 2d entanglement entropy diverges close to this point. As a consequence. The positive time region is inaccessible for observers in tau = -infinity. For a stationary measure, the vacuum at finite time is seen by those observers as a thermal state and the information loss is encoded as a heat bath of string states. (c) 2006 Elsevier B.V. All rights reserved.

Complex crystallization dynamics in amorphous germanium observed with dynamic transmission electron microscopy

Crystallization of amorphous germanium (a-Ge) by laser or electron beam heating is a remarkably complex process that involves several distinct modes of crystal growth and the development of intricate microstructural patterns on the nanosecond to ten microsecond time scales. Here we use dynamic transmission electron microscopy (DTEM) to study the fast, complex crystallization dynamics with 10 nm spatial and 15 ns temporal resolution. We have obtained time-resolved real-space images of nanosecond laser-induced crystallization in a-Ge with unprecedentedly high spatial resolution. Direct visualization of the crystallization front allows for time-resolved snapshots of the initiation and roughening of the dendrites on submicrosecond time scales. This growth is followed by a rapid transition to a ledgelike growth mechanism that produces a layered microstructure on a time scale of several microseconds. This study provides insights into the mechanisms governing this complex crystallization process and is a dramatic demonstration of the power of DTEM for studying time-dependent material processes far from equilibrium.

Europium ion labeling anti-HBs

A microsecond time-resolved laser fluorescence spectroscopic analysis set was developed, A chelate-cyclic anhydride of diethylenetrimin pentaacetic acid anhydride (DTPAA) was synthesized. An anti-HBs antibody was purified, A EU3+ -DTPAA-anti-HBs label was prepared by two step procedure. We described the optimal condtion with EU3+ as marker and DTPAA as chelate bounding to antibody molecule. Labeling parameters such as solvent pH, protein and chelate molar ratio, reaction time, separation method were discussed in detail.

Pressure-jump X-ray studies of liquid crystal transitions in lipids

In this paper, we give an overview of our studies by static and time-resolved X-ray diffraction of inverse cubic phases and phase transitions in lipids. In 1, we briefly discuss the lyotropic phase behaviour of lipids, focusing attention on non-lamellar structures, and their geometric/topological relationship to fusion processes in lipid membranes. Possible pathways for transitions between different cubic phases are also outlined. In 2, we discuss the effects of hydrostatic pressure on lipid membranes and lipid phase transitions, and describe how the parameters required to predict the pressure dependence of lipid phase transition temperatures can be conveniently measured. We review some earlier results of inverse bicontinuous cubic phases from our laboratory, showing effects such as pressure-induced formation and swelling. In 3, we describe the technique of pressure-jump synchrotron X-ray diffraction. We present results that have been obtained from the lipid system 1:2 dilauroylphosphatidylcholine/lauric acid for cubic-inverse hexagonal, cubic-cubic and lamellar-cubic transitions. The rate of transition was found to increase with the amplitude of the pressure-jump and with increasing temperature. Evidence for intermediate structures occurring transiently during the transitions was also obtained. In 4, we describe an IDL-based 'AXCESS' software package being developed in our laboratory to permit batch processing and analysis of the large X-ray datasets produced by pressure-jump synchrotron experiments. In 5, we present some recent results on the fluid lamellar-Pn3m cubic phase transition of the single-chain lipid 1-monoelaidin, which we have studied both by pressure-jump and temperature-jump X-ray diffraction. Finally, in 6, we give a few indicators of future directions of this research. We anticipate that the most useful technical advance will be the development of pressure-jump apparatus on the microsecond time-scale, which will involve the use of a stack of piezoelectric pressure actuators. The pressure-jump technique is not restricted to lipid phase transitions, but can be used to study a wide range of soft matter transitions, ranging from protein unfolding and DNA unwinding and transitions, to phase transitions in thermotropic liquid crystals, surfactants and block copolymers.

Heterosite effects in novel heteronuclear clusters [OS2Ru(CO)(11)(PPh3)] and Os2Ru(CO)(10)(2-acetylpyridine-N-isopropylimine)]

A new synthetic route towards the mixed-metal cluster [OS2Ru(CO)(12)] is described together with the syntheses of its PPh3 and iPr-AcPy (iPr-AcPy = 2-acetylpyridine-N-isopropylimine) derivatives. The molecular structures of the novel clusters [Os2Ru(CO)(11)(PPh3)] and [Os2Ru(CO)(10)(iPr-AcPy)] were determined on the basis of crystalline solid solutions of the Os2Ru and corresponding Os-3 species. The structures reveal that coordination of the Lewis bases occurs exclusively at the ruthenium site of [Os2Ru(CO)(12)], which is in agreement with density functional theory (DFT) calculations on several structural isomers of these compounds. According to the time-dependent DFT results, the lowest optically accessible excited state of [Os2Ru(CO)(10)(iPr-AcPy)] has a prevailing sigma(Ru-Os-2)pi*(iPr-AcPy) character, with a partial sigma sigma*(Ru-Os-2) contribution. In weakly coordinating 2-chlorobutane, the excited state has a lifetime tau = 10.4 +/- 1.2 ps and produces biradicals considerably faster than observed for [Os-3(CO)10(iPr-AcPy) (tau = 25.3 +/- 0.7ps)]. In coordinating acetonitrile, the excited state of [Os2Ru(CO)(10)(iPr-AcPy)] decays mono-exponentially with a lifetime tau = 2.1 +/- 0.2 ps. In contrast to [Os-3(CO)(10)(iPr-AcPy)] that forms biradicals as the main primary photoproduct even in strongly coordinating solvents, zwitterion formation from the solvated lowest excited state is observed for the heterometallic cluster. This is concluded from time-resolved absorption studies in the microsecond time domain. Due to the lower tendency of the coordinatively unsaturated Ru+(CO)(2)(iPr-AcPy-/0) moiety to bind a Lewis base, the heteronuclear biradical and zwitterionic photoproducts live significantly shorter than their triosmium counterparts. The influence of the weaker Os-2-Ru(iPr-AcPy) bond on the redox reactivity is clearly reflected in very reactive radical anions formed upon electrochemical reduction of [Os2Ru(CO)(10)(iPr-AcPy)]. The dimer [-OS(CO)(4)-Os(CO)(4)-Ru(CO)(2)(iPr-AcPy)](2)(2-) is the only IR-detectable intermediate reduction product. The dinuclear complex [Os-2(CO)(8)](2-) and insoluble [Ru(CO)(2)(iPr-AcPy)](n), are the ultimate reduction products, proving fragmentation of the OS2Ru core.

Aircraft routing with minimal climate impact: the REACT4C climate cost function modelling approach (V1.0)

In addition to CO2, the climate impact of aviation is strongly influenced by non-CO2 emissions, such as nitrogen oxides, influencing ozone and methane, and water vapour, which can lead to the formation of persistent contrails in ice-supersaturated regions. Because these non-CO2 emission effects are characterised by a short lifetime, their climate impact largely depends on emission location and time; that is to say, emissions in certain locations (or times) can lead to a greater climate impact (even on the global average) than the same emission in other locations (or times). Avoiding these climate-sensitive regions might thus be beneficial to climate. Here, we describe a modelling chain for investigating this climate impact mitigation option. This modelling chain forms a multi-step modelling approach, starting with the simulation of the fate of emissions released at a certain location and time (time-region grid points). This is performed with the chemistry–climate model EMAC, extended via the two submodels AIRTRAC (V1.0) and CONTRAIL (V1.0), which describe the contribution of emissions to the composition of the atmosphere and to contrail formation, respectively. The impact of emissions from the large number of time-region grid points is efficiently calculated by applying a Lagrangian scheme. EMAC also includes the calculation of radiative impacts, which are, in a second step, the input to climate metric formulas describing the global climate impact of the emission at each time-region grid point. The result of the modelling chain comprises a four-dimensional data set in space and time, which we call climate cost functions and which describes the global climate impact of an emission at each grid point and each point in time. In a third step, these climate cost functions are used in an air traffic simulator (SAAM) coupled to an emission tool (AEM) to optimise aircraft trajectories for the North Atlantic region. Here, we describe the details of this new modelling approach and show some example results. A number of sensitivity analyses are performed to motivate the settings of individual parameters. A stepwise sanity check of the results of the modelling chain is undertaken to demonstrate the plausibility of the climate cost functions.

Interfacial Electron Transfer Dynamics Following Laser Flash Photolysis of [Ru(bpy)(2)((4,4 `-PO(3)H(2))(2)bpy)](2+) in TiO(2) Nanoparticle Films in Aqueous Environments

Nanosecond laser flash photolysis has been used to investigate injection and back electron transfer from the complex [(Ru-(bpy)(2)(4,4`-(PO(3)H(2))(2)bpy)](2+) surface-bound to TiO(2) (TiO(2)-Ru(II)). The measurements were conducted under conditions appropriate for water oxidation catalysis by known single-site water oxidation catalysts. Systematic variations in average lifetimes for back electron transfer, - were observed with changes in pH, surface coverage, incident excitation intensity, and applied bias. The results were qualitatively consistent with a model involving rate-limiting thermal activation of injected electrons from trap sites to the conduction band or shallow trap sites followed by site-to-site hopping and interfacial electron transfer, TiO(2)(e(-))-Ru(3+) -> TiO(2)-Ru(2+). The appearance of pH-dependent decreases in the efficiency of formation of TiO(2)-Ru(3+) and in incident-photon-to-current efficiencies with the added reductive scavenger hydroquinone point to pH-dependent back electron transfer processes on both the sub-nanosecond and millisecond-microsecond time scales, which could be significant in limiting long-term storage of multiple redox equivalents.

Linear Bayes policy for learning in contextual-bandits

Machine and Statistical Learning techniques are used in almost all online advertisement systems. The problem of discovering which content is more demanded (e.g. receive more clicks) can be modeled as a multi-armed bandit problem. Contextual bandits (i.e., bandits with covariates, side information or associative reinforcement learning) associate, to each specific content, several features that define the “context” in which it appears (e.g. user, web page, time, region). This problem can be studied in the stochastic/statistical setting by means of the conditional probability paradigm using the Bayes’ theorem. However, for very large contextual information and/or real-time constraints, the exact calculation of the Bayes’ rule is computationally infeasible. In this article, we present a method that is able to handle large contextual information for learning in contextual-bandits problems. This method was tested in the Challenge on Yahoo! dataset at ICML2012’s Workshop “new Challenges for Exploration & Exploitation 3”, obtaining the second place. Its basic exploration policy is deterministic in the sense that for the same input data (as a time-series) the same results are obtained. We address the deterministic exploration vs. exploitation issue, explaining the way in which the proposed method deterministically finds an effective dynamic trade-off based solely in the input-data, in contrast to other methods that use a random number generator.

Cytochrome c′ folding triggered by electron transfer: Fast and slow formation of four-helix bundles

Reduced (FeII) Rhodopseudomonas palustris cytochrome c′ (Cyt c′) is more stable toward unfolding ([GuHCl]1/2 = 2.9(1) M) than the oxidized (FeIII) protein ([GuHCl]1/2 = 1.9(1) M). The difference in folding free energies (ΔΔGf° = 70 meV) is less than half of the difference in reduction potentials of the folded protein (100 mV vs. NHE) and a free heme in aqueous solution (≈−150 mV). The spectroscopic features of unfolded FeII–Cyt c′ indicate a low-spin heme that is axially coordinated to methionine sulfur (Met-15 or Met-25). Time-resolved absorption measurements after CO photodissociation from unfolded FeII(CO)–Cyt c′ confirm that methionine can bind to the ferroheme on the microsecond time scale [kobs = 5(2) × 104 s−1]. Protein folding was initiated by photoreduction (two-photon laser excitation of NADH) of unfolded FeIII–Cyt c′ ([GuHCl] = 2.02–2.54 M). Folding kinetics monitored by heme absorption span a wide time range and are highly heterogeneous; there are fast-folding (≈103 s−1), intermediate-folding (102–101 s−1), and slow-folding (10−1 s−1) populations, with the last two likely containing methionine-ligated (Met-15 or Met-25) ferrohemes. Kinetics after photoreduction of unfolded FeIII–Cyt c′ in the presence of CO are attributable to CO binding [1.4(6) × 103 s−1] and FeII(CO)–Cyt c′ folding [2.8(9) s−1] processes; stopped-flow triggered folding of FeIII–Cyt c′ (which does not contain a protein-derived sixth ligand) is adequately described by a single kinetics phase with an estimated folding time constant of ≈4 ms [ΔGf° = −33(3) kJ mol−1] at zero denaturant.

Cocaine: mechanism of inhibition of a muscle acetylcholine receptor studied by a laser-pulse photolysis technique.

Effects of cocaine on the muscle nicotinic acetylcholine receptor were investigated by using a chemical kinetic technique with a microsecond time resolution. This membrane-bound receptor regulates signal transmission between nerve and muscle cells, initiates muscle contraction, and is inhibited by cocaine, an abused drug. The inhibition mechanism is not well understood because of the lack of chemical kinetic techniques with the appropriate (microsecond) time resolution. Such a technique, utilizing laser-pulse photolysis, was recently developed; by using it the following results were obtained. (i) The apparent cocaine dissociation constant of the closed-channel receptor form is approximately 50 microM. High carbamoylcholine concentration and, therefore, increased concentrations of the open-channel receptor form, decrease receptor affinity for cocaine approximately 6-fold. (ii) The rate of the receptor reaction with cocaine is at least approximately 30-fold slower than the channel-opening rate, resulting in a cocaine-induced decrease in the concentration of open receptor channels without a concomitant decrease in the channel-opening or -closing rates. (iii) The channel-closing rate increases approximately 1.5-fold as the cocaine concentration is increased from 20 to 60 microM but then remains constant as the concentration is increased further. The results are consistent with a mechanism in which cocaine first binds rapidly to a regulatory site of the receptor, which can still form transmembrane channels. Subsequently, a slow step (t1/2 approximately 70 ms) leads to a receptor form that cannot form transmembrane channels, and acetylcholine receptor-mediated signal transmission is, therefore, blocked. Implications for the search for therapeutic agents that alleviate cocaine poisoning are mentioned.

Polarization switching in stretched pulse fiber laser

We studied experimentally polarization dynamics in a carbon nanotube mode locked stretched pulse fiber laser. For the first time, polarization locked, regular and irregular polarization switching have been observed at the microsecond time scale. © 2014 OSA.