Modeling membranes under a transmembrane potential

Accurate modeling of ion transport through synthetic and biological transmembrane channels has been so far a challenging problem. We introduce here a new method that allows one to study such transport under realistic biological conditions. We present results from molecular dynamics simulations of an ion channel formed by a peptide nanotube, embedded in a lipid bilayer, and subject to transmembrane potentials generated by asymmetric distributions of ions on both sides of the membrane. We show that the method is efficient for generating ionic currents and allows us to estimate the intrinsic conductance of the channel.     

Cation selective ion channels formed by macrodiolide antibiotic elaiophylin in lipid bilayer membranes

The macrodiolide antibiotic elaiophylin (1) forms stable, long-lasting cation selective ion channels in planar lipid bilayer membranes prepared from soybean phosphatidylcholine. Current of the single ion channel displayed two sublevels corresponding to the two substates of the channel conductance: a slow substate, with about 5 s of mean dwell time in the open state at 40% level of the total amplitude conductance, and a fast substate of higher conductance with dwell times in the open and closed state of about 0.1 s. Amplitude conductances of the single ion channels in 200 mM of LiCl, NaCl, KCl, RbCl and CsCl were 75, 140, 220, 240 and 226 pS, and the conductance was linear function of the electrolyte concentration. Ratios of cation to anion permeabilities of the channel for NaCl and KCl were 8+/-2 and >24, respectively. A molecular model of the channel structure is suggested.     

Effect of agents modifying the membrane dipole potential on properties of syringomycin E channels

We evaluated the effect of agents modifying the membrane dipole potential: phloretin, 6-ketocholestanol and RH 421 on the properties of single channels formed by lipodepsipeptide syringomycin E (SRE) in planar lipid bilayers. SRE forms two conductive states in lipid bilayers: "small" and "large." Large SRE channels are clusters of several small ones, demonstrating synchronous openings and closures. The increase in the membrane dipole potential led to (i) an increase in SRE channel conductance, (ii) an increase in the channel's lifetime, and (iii) a decrease in a number of synchronously operating small channels in the clusters. Overall, the results support the model of the small SRE channel synchronization in the cluster as voltage-dependent orientation of the lipid dipoles associated with the channel pores.     

Design of a recursively-structured valveless device for microfluidic manipulation

A recursively-structured apparatus based on a pneumatic pumping structure has been investigated numerically and experimentally in the present study. For the T-connected channels, this apparatus demonstrated the ability to manipulate the liquid drop from a first channel to a second channel, while simultaneously preventing flow into the third channel. The microTAS research aimed at biochemical analysis miniaturization and integration has recently made explosive progress. However, there is still a considerable technical challenge in integrating these procedures into a multiple-step system. An important issue for this integration is microfluid management techniques. The microTAS method must be designed considering special transport mechanisms to move samples and reagents through the microchannels. The structure of this apparatus was simple and easily fabricated. Moreover, because there is a continuous airflow at the "outlet" during fluid manipulation, it is possible to avoid contamination of the air source similar to the "laminar flow hook" in biological experiments. Utilizing the concept of a recursive structure, one can easily design a device wherein more than three channels are included in the flow network, either intersecting in a single junction or in multiple junctions.     

Modulating ion channel properties of transmembrane peptide nanotubes through heteromeric supramolecular assemblies

A selective heteromeric supramolecular assembly process is devised to create functional single channels of altered ion conductance, charge selectivity, and rectification. The hollow transmembrane tubular structure produced spontaneously from the self-assembly of cyclic-d,l-alpha-peptides in lipid bilayers is modified by designer cyclic peptide "cap" subunits that bind site-selectively at the mouth of the channel assembly.     

Anisotropic Electron Conductance Driven by Reaction Byproducts on a Porous Network of Dibromobenzothiadiazole on Cu(110)

Efficiency in charge‐transport is a fundamental but demanding prerequisite to allow better exploitation of molecular functionalities in organic electronics and energy‐conversion systems. Here, we report on a mechanism that enables a one‐dimensional conductance structure by connecting discrete molecular states at 2.1 eV through the pores of a metal–organic network on Cu(110). Two adjacent, periodic and isoenergetic contributions, namely a molecular resonance and the confined surface‐state, add‐up leading to anisotropic structures, as channels, observable in real‐space conductance images. The adsorption configurations of Br atoms, inorganic byproduct of the redox‐reacted 4,7‐dibromobenzo[c]‐1,2,5‐thiadiazole (2Br‐BTD) molecules on the copper surface, drive the confinement of the Cu surface state within the pores and critically control the channel continuity. Small displacements of the Br atoms change the local surface potential misaligning the energy levels. This work visualizes the effect of order‐disorder transitions caused by the movement of single atoms in the electronic properties of two‐dimensional organic networks.     

An Improved Method of Determining the zeta-Potential and Surface Conductance

In the classical "slope-intercept" method of determining the zeta potential and the surface conductance, the relationship between DeltaP and E(s) is measured experimentally at a number of different channel sizes (e.g., the height of a slit channel, h). The parameter (epsilon(r)epsilon(0)DeltaP/μE(s)lambda(b)) is then plotted as a function of 1/h and linear regression is performed. The y-intercept of the regressed line is then related to the zeta-potential and its slope to the surface conductance. However, in this classical method, the electrical double layer effect or the electrokinetic effects on the liquid flow are not considered. Consequently, this technique is valid or accurate only when the following conditions are met: (1) relatively large channels are used; (2) the electrical double layer is sufficiently thin; and (3) the streaming potential is sufficiently small that the electroosmotic body force on the mobile ions in the double layer region can be ignored. In this paper a more general or improved slope-intercept method is developed to account for cases where the above three conditions are not met. Additionally a general least-squares analysis is described which accounts for uncertainty in the measured channel height as well as unequal variance in the streaming potential measurements. In this paper, both the classical and the improved slope-intercept techniques have been applied to streaming potential data measured with slit glass channels, ranging in height from 3 μm to 66 μm, for several aqueous electrolyte solutions. The comparison shows that the classical method will always overestimate both the zeta-potential and the surface conductance. Significant errors will occur when the classical method is applied to systems with small channel heights and low ionic concentrations. Furthermore, it is demonstrated that traditional regression techniques where the uncertainty is confined only to the dependent variable and each measurement is given equal weight may produce physically inconsistent results. Copyright 2000 Academic Press.     

Method to determine the effective ζ potential in a microchannel with an embedded gate electrode

We present a theoretical model to determine the effective zeta potential ζ(eff) in microfluidic channels where an embedded, insulated gate electrode allows for external tuning of a portion of the channel surface charge. In addition, we derive a method to determine ζ(eff) in such channels, for any value of salt concentration, using the solution displacement technique. To do so, we simulate typical current-monitoring measurements using our model, and highlight the experimental parameters that lead to inaccurate results using this procedure with an heterogenous channel. Our method corrects for such inaccuracies by using our model with experimental data to find the correct value of ζ(eff) . Finally, we perform experiments to demonstrate our method and the use of our model with a silica-PMDS microchannel system with an embedded Ti-Au-Ti gate electrode that covers 50% of the bottom surface of the channel. We show that our theory captures the salient features of our experiments, thereby offering a useful tool to predict effective zeta potential in channels with a nonuniform zeta potential.     

A spectral overlap method for self monitoring quench correction in double isotope liquid scintillation counting

A method for quench correction of samples with double radioactive labelling is described. Each nuclide makes a contribution to the counting rate of three channels of a liquid scintillation counter. This channel overlap is an essential requirement of the calibration procedure rather than a limitation, and allows more freedom in the choice of counting conditions. After calibration with suitable standards the method will tolerate wide variations in the ratio of one isotope to the other extending to single label samples of either isotope. This is the outstanding advantage over the channel ratio method which requires a statistically significant counting rate for the higher energy isotope. The method takes advantage of the facilities offered by a computer which may be on line or remote.¹⁴C and tritium are used to demonstrate the utility of the method.     

Theory of transport in nanofluidic channels with moderately thin electrical double layers: effect of the wall potential modulation on solutions of symmetric and asymmetric electrolytes

Electrokinetic phenomena play an important role for the transport in submicrometer-size channels since the electric double layers formed at the walls can occupy a substantial part of the channel volume. This presents a theoretical difficulty and specific problems are usually treated numerically or not comprehensively. In our work we present a theoretical model that allows one to obtain analytical expressions for the transport of fluid (electro-osmotic flow), ions (electric current), and dissolved charged molecules (analytes). The model is based on the weak double layer approximation and has a wide range of validity. An important feature of this theoretical approach is that it is applicable not only to symmetric but also to asymmetric 2:1 and 1:2 electrolytes which exhibit very interesting properties in nanoscale channels. The possibility of affecting the wall electrokinetic zeta potential by applying a transverse voltage bias is analyzed. This transverse bias is used in an attempt to control the transport in the channel and such devices are often called "fluidic field-effect transistors." Our model quantifies the effect of the voltage bias on the zeta potential of the channel wall and therefore can be used for prediction of transport and optimization of separations in such fluidic devices.     

Potential of microbore HPLC within a multiresidue method for the trace analysis of plant-protective substances in water

The performance of microbore HPLC as a "measurement channel" within a true multiclass/multiresidue method for monitoring plant protectants in raw and potable water is demonstrated. The method has a modular design and consists of a non-selective sampling and preparation line generating 250 microL of an "extract" from a 100-mL water sample; this extract can be introduced to up to four measurement channels, as required by the analytical task. The microbore HPLC channel can be used to quantify 34 plant protectants in the 0.1 microg L(-1) concentration range by use of diode-array detection at seven different wavelengths. A solvent change is necessary to link sample preparation to microbore HPLC; this uses 50 microL of the "extract" and is accomplished directly in an autosampler vial. Performance characteristics were evaluated for tap water spiked at 0.2 microg L(-1). Average recoveries were between 65 and 100% and method detection limits were 0.07 microg L(-1) or better. The ability to provide comparable and accurate results was proven by participation in an interlaboratory comparison trial. The procedure for preparing microbore columns from 750 microm i.d. PEEK tubing is described in detail to enable the reader to prepare his own columns. The reproducibility of this preparation procedure was proven by an analysis-of-variance test.     

Staphylococcus aureus bicomponent gamma-hemolysins, HlgA, HlgB, and HlgC, can form mixed pores containing all components

Staphylococcal gamma-hemolysins are bicomponent toxins forming a protein family with leucocidins and alpha-toxin. Two active toxins (AB and CB) can be formed combining one of the class-S components, HlgA or HlgC, with the class-F component HlgB. These two gamma-hemolysins form pores with marked similarities to alpha-toxin in terms of conductance, nonlinearity of the current-voltage curve, and channel stability in the open state. AB and CB pores, however, are cation-selective, whereas alpha-toxin is anion-selective. gamma-Hemolysins' pores are hetero-oligomers formed by three or four copies of each component (indicated as 3A3B and 3C3B or 4A4B and 4C4B). Point mutants located on a beta-strand of the class-S component that forms part of the protomer-protomer contact region can prevent oligomer assembly. Interestingly, these mutants inhibit growth of pores formed not only by their natural components but also by nonstandard components. This lead to the hypothesis that mixed ABC pores could also be formed. By studying the conductance of pores, assembled in the presence of all three components (in different ratios), it was observed that the magnitudes expected for mixed pores were, indeed, present. We conclude that the gamma-hemolysin/leucocidin bicomponent toxin family may form a larger than expected number of active toxins by cross-combining various S and F components.     

Analysis of Li<Superscript>+</Superscript> cation migration paths in oxygen-containing compounds

Novel method for the studying of voids and channels in crystal structures is developed on the basis of the TOPOS structure-topology software. The method is using the Voronoi-Dirichlet crystal space partition. All ternary and quaternary lithium-containing inorganic compounds whose structure has been studied (822 compounds of the Li _a X _b O _z composition and 1349 compounds of the Li _a X1 _b X2 _c O _z composition, where X, X1, and X2 is any chemical element) are analyzed for the first time. The dimensionality of systems of the channels capable of transporting lithium ions is revealed. For all compounds, the migration patterns are constructed, which characterize systems of the conductance channels with the dimensionality 1, 2, and 3; the theoretically calculated coordinates of lithium atom positions in the voids’ centers agree well (accurate within 0.06 nm) with the known structure data. It is found, that 275 compounds have infinite channel system. Of this sampling, 249 compounds (125 structural types) have been described as solid electrolytes; the rest (26 compounds) can be thought of as potential ionic conductors with one-dimensional (6 types), two-dimensional (2 types), or three-dimensional (18 types) conductance.     

Ionic channel behavior of modified cyclodextrins inserted in lipid membranes

We study the insertion and behavior of modified amphiphilic cyclodextrins in suspended bilayer lipid membranes by electrophysiological methods. We observe that our molecules build single well-defined ionic channels. The pore conductance is measured in two lipid membranes differing by their composition. These measurements reveal two distinct behaviors. In the case of thin membranes, we observe single channels, whereas in the case of thick membranes, we only detect a large number of aggregated channels. In a few experiments, we have been able to monitor the transition between the two behaviors by modifying slightly the swelling of the lipid bilayers by decane. The precise structure of the channels is yet unknown; however, we deduce from our measurements an estimation of the channel diameter.     

Strong effects of molecular structure on electron transport in carbon/molecule/copper electronic junctions

Carbon/molecule/copper molecular electronic junctions were fabricated by metal deposition of copper onto films of various thicknesses of fluorene (FL), biphenyl (BP), and nitrobiphenyl (NBP) covalently bonded to flat, graphitic carbon. A "crossed-wire" junction configuration provided high device yield and good junction reproducibility. Current/voltage characteristics were investigated for 69 junctions with various molecular structures and thicknesses and at several temperatures. The current/voltage curves for all cases studied were nearly symmetric, scan rate independent, repeatable at least thousands of cycles and exhibited negligible hysteresis. Junction conductance was strongly dependent on the dihedral angle between phenyl rings and on the nature of the molecule/copper "contact". Junctions made with NBP showed a decrease in conductivity of a factor of 1300 when the molecular layer thickness increased from 1.6 to 4.5 nm. The slope of ln(i) vs layer thickness for both BP and NBP was weakly dependent on applied voltage and ranged from 0.16 to 0.24 A(-1). These attenuation factors are similar to those observed for similar molecular layers on modified electrodes used to study electrochemical kinetics. All junctions studied showed weak temperature dependence in the range of approximately 325 to 214 K, implying activation barriers in the range of 0.06 to 0.15 eV. The carbon/molecule/copper junction structure provides a robust, reproducible platform for investigations of the dependence of electron transport in molecular junctions on both molecular structure and temperature. Furthermore, the results indicate that junction conductance is a strong function of molecular structure, rather than some artifact resulting from junction fabrication.     

Mechanical compression in cofacial porphyrin cyclophane pincers

Intra- and intermolecular interactions are dominating chemical processes, and their concerted interplay enables complex nonequilibrium states like life. While the responsible basic forces are typically investigated spectroscopically, a conductance measurement to probe and control these interactions in a single molecule far out of equilibrium is reported here. Specifically, by separating macroscopic metal electrodes, two π-conjugated, bridge-connected porphyrin decks are peeled off on one side, but compressed on the other side due to the covalent mechanical fixation. We observe that the conductance response shows an exceptional exponential rise by two orders of magnitude in individual breaking events during the stretching. Theoretical studies atomistically explain the measured conductance behavior by a mechanically activated increase in through-bond transport and a simultaneous strengthening of through-space coupling. Our results not only reveal the various interacting intramolecular transport channels in a molecular set of levers, but also the molecules'' potential to serve as molecular electro-mechanical sensors and switches.

A two-order conductance increase upon stretching in porphyrin cyclophane pincer junctions is measured. Atomistic studies explain experimental observations by characteristic intramolecular changes in through-space and through-bond transport.     

Alamethicin-leucine zipper hybrid peptide: a prototype for the design of artificial receptors and ion channels.

In this report, we describe a novel concept of extramembrane control of channel peptide assembly and the eventual channel current modulation. Alamethicin is a peptide antibiotic, which usually forms ion channels in various association states. By introducing an extramembrane leucine zipper segment (Alm-LeuZ), the association number of alamethicin was effectively controlled to produce a single predominant channel open state. The assembly was estimated to be a tetramer, by comparison of the channel conductance with that of the template-assembled Alm-LeuZ tetramer, which was prepared by the conjugation of a maleimide-functionalized peptide template with cysteine-derivatized Alm-LeuZ segments. Employment of an extramembrane segment of a random conformation provided higher levels of channel conductance. The result exemplified the possibility of channel current control by a conformational switch of the extramembrane segments.     

Ionic conductance of synthetic channels: analysis, lessons, and recommendations

Synthetic ion channels have been known for nearly three decades, but it is only in the past decade that analysis of the currents these ionic conductors carry has become a standard technique. A broad range of structural types have been explored and these reports have produced a very diverse collection of ion channel conductance behaviours. In this critical review we describe a notational method to extract salient information from reported ion channel experiments. We use an activity grid to represent quantitative information on conductance and opening duration with a five-level colour code to represent qualitative information on the nature of the conductance-time profile. Analysis of the cumulative dataset suggests that the reported conductance data can reflect the structural features of the compounds prepared, but does also reflect the energetic landscape of the bilayer membrane in which synthetic ion channels function (143 references).     

Anisotropic Conductivity at the Single‐Molecule Scale

In most junctions built by wiring a single molecule between two electrodes, the electrons flow along only one axis: between the two anchoring groups. However, molecules can be anisotropic, and an orientation‐dependent conductance is expected. Here, we fabricated single‐molecule junctions by using the electrode potential to control the molecular orientation and access individual elements of the conductivity tensor. We measured the conductance in two directions, along the molecular plane as the benzene ring bridges two electrodes using anchoring groups (upright) and orthogonal to the molecular plane with the molecule lying flat on the substrate (planar). The perpendicular (planar) conductance is about 400 times higher than that along the molecular plane (upright). This offers a new method for designing a reversible room‐temperature single‐molecule electromechanical switch that controllably employs the electrode potential to orient the molecule in the junction in either “ON” or “OFF” conductance states.     

Further characterization of cation channels present in the chicken red blood cell membrane

In this paper, we provide an update on cation channels in nucleated chicken erythrocytes. Patch-clamp techniques were used to further characterize the two different types of cation channels present in the membrane of chicken red blood. In the whole-cell mode, with Ringer in the bath and internal K+ saline in the pipette solution, the membrane conductance was generated by cationic currents, since the reversal potential was shifted toward cations equilibrium when the impermeant cation NMDG was substituted to small cations. The membrane conductance could be increased by application of mechanical deformation or by the addition of agonists of the cAMP-dependent pathway. At the unitary level, two different types of cationic channels were revealed and could account for the cationic conductance observed in whole-cell configuration. One of them belongs to the family of stretch-activated cationic channel showing changes in activity under conditions of membrane deformation, whereas the second one belongs to the family of the cAMP activated cationic channels. These two channels could be distinguished according to their unitary conductances and drug sensitivities. The stretch-activated channel was sensitive to Gd(3+) and the cAMP-dependent channel was sensitive to flufenamic acid. Possible role of these channels in cell volume regulation process is discussed.