Polarographic studies of ion binding in solutions of pepsin. II. Binding of thallous ions

The results of the application of a modified polarographic method to the study of binding of thallous ions in solutions of pepsin are reported. Isoionic solutions of pepsin were converted by using a cation exchange resin to solutions of thallous pepsin. The extent of ion binding was obtained as in the case of cadmium pepsin by assuming that only free, i.e., unbound, ions contribute to the diffusion and migration currents, respectively. However, the relations used previously for computing the degree of ion binding have been refined on the basis of experience acquired in a parallel study of solutions of thallous acrylate and polyacrylate, respectively.     

Analytical solutions of Poisson's equation for realistic geometrical shapes of membrane ion channels.

Analytical solutions of Poisson's equations satisfying the Dirichlet boundary conditions for a toroidal dielectric boundary are presented. The electric potential computed anywhere in the toroidal conduit by the analytical method agrees with the value derived from an iterative numerical method. We show that three different channel geometries, namely, bicone, catenary, and toroid, give similar potential profiles as an ion traverses along their central axis. We then examine the effects of dipoles in the toroidal channel wall on the potential profile of ions passing through the channel. The presence of dipoles eliminates the barrier for one polarity of ion, while raising the barrier for ions of the opposite polarity. We also examine how a uniform electric field from an external source is affected by the protein boundary and a mobile charge. The channel distorts the field, reducing it in the vestibules, and enhancing it in the constricted segment. The presence of an ion in one vestibule effectively excludes ions of the same polarity from that vestibule, but has little effect in the other vestibule. Finally, we discuss how the solutions we provide here may be utilized to simulate a system containing a channel and many interacting ions.     

Anomalous mole fraction effect, electrostatics, and binding in ionic channels.

Ionic channels bathed in mixed solutions of two permeant electrolytes often conduct less current than channels bathed in pure solutions of either. For many years, this anomalous mole fraction effect (AMFE) has been thought to occur only in single-file pores containing two or more ions at a time. Most thinking about channels incorporates this view. We show here that the AMFE arises naturally, as an electrostatic consequence of localized ion specific binding, if the average current through a channel is described by a theory (Poisson-Nernst-Planck, PNP) that computes the average electric field from the average concentration of charges in and near the channel. The theory contains only those ion-ion interactions mediated by the mean field, and it does not enforce single filing. The AMFE is predicted by PNP over a wide range of mean concentrations of ions in the channel; for example, it is predicted when (on the average) less, or much less, than one ion is found in the channel's pore. In this treatment, the AMFE arises, in large measure, from a depletion layer produced near a region of ion-specific binding. The small excess concentration of ions in the binding region repels all nearby ions of like charge, thereby creating a depletion layer. The overall conductance of the channel arises in effect from resistors in series, one from the binding region, one from the depletion zone, and one from the unbinding region. The highest value resistor (which occurs in the depletion zone) limits the overall series conductance. Here the AMFE is not the result of single filing or multiple occupancy, and so previous views of permeation need to be revised: the presence of an AMFE does not imply that ions permeate single file through a multiply occupied pore.     

The Electrical Conductance of Semipermeable Membranes: III. Bipolar Flow-Symmetric Electrolytes

The first paper of this series presented a general formulation of the problem of stationary ion flow through membranes. The second treated in detail the special case of unipolar flow across membranes separating symmetric electrolytes. In this, the third paper of the series, we deal with another special case, that of bipolar flow between symmetric electrolytes. Here it is assumed that the total current is carried by both positive and negative permeant ions. The restriction to symmetric electrolytes implies that all ions present in the membrane and surrounding solutions have valences of identical absolute magnitude. After extracting from the general development a set of equations appropriate to the special case being considered, we outline a procedure for the numerical solution of the conductance problem for this case. Numerical results, presented as part of a discussion of approximate analytic methods of solution, establish the utility of these methods. A discussion of the significance of this work for membrane studies is presented in conclusion.     

Interactions des ions métalliques avec le DNA. I. Comportement de polyélectrolyte et liaison des ions compensateurs

The theory of polyelectrolyte solution of Alexandrowicz: and Katchalsky is used to calculate the electrostatic potential of single stranded polynucleotides for different ionic strength. We have considered the potential of double stranded DNA as the superposition of the different potentials produced by each chain, provided the average distance between the strands is higher than an ionic strength-dependent parameter b. For ionic strength lower than 5 × 10^?2M, the assumption is no longer valid, and a cylindrical model with a uniform charge density must be used. The continuity between the two models was tested, and thus we can calculate the electrical potential at the vicinity of a phosphate group in the whole range of ionic-strength where the double helix is stable. It was therefore possible to determine the theoretical number of ions bound electrostatically to DNA and we found an increase of ion binding with a decrease of ionic strength. Such a model was further applied to the change of specific volume in different salt solutions. Comparison is made with recent pycnometric data on Na^? and Cs^? salts of DNA. Agreement is good in the case of Cs⁺, but for Na⁺, cation binding is likely to be accompanied by a change of the hydration of DNA, which depends on ionic strength. With the same model we can see easily the ion-trapping properties of DNA which play an important role in any formation of complex between heavy ions and bases.     

Mono- and trivalent ions around DNA: a small-angle scattering study of competition and interactions

The presence of small numbers of multivalent ions in DNA-containing solutions results in strong attractive forces between DNA strands. Despite the biological importance of this interaction, e.g., DNA condensation, its physical origin remains elusive. We carried out a series of experiments to probe interactions between short DNA strands as small numbers of trivalent ions are included in a solution containing DNA and monovalent ions. Using resonant (anomalous) and nonresonant small angle x-ray scattering, we coordinated measurements of the number and distribution of each ion species around the DNA with the onset of attractive forces between DNA strands. DNA-DNA interactions occur as the number of trivalent ions increases. Surprisingly good agreement is found between data and size-corrected numerical Poisson-Boltzmann predictions of ion competition for non- and weakly interacting DNAs. We also obtained an estimate for the minimum number of trivalent ions needed to initiate DNA-DNA attraction.     

Mass Transfer in the Cornea: I. Interacting Ion Flows in an Arbitrarily Charged Membrane

The formalisms of irreversible thermodynamics are used to describe multi-ionic nonconvective flow through an arbitrarily charged membrane. Interactions between oppositely charged ions are included and are measured by a single phenomenological coefficient. The consequent generalized Nernst-Planck flux equations are integrated to yield a relation between the species fluxes and the composition of the solutions bounding the membrane. It is assumed in the derivation that activity coefficient gradients within the membrane and direct interactions between ions of like charge are negligible. Some special cases are examined. To illustrate the use of the final equations, a single membrane separating solutions of differing composition is modeled, and the effect of ion-ion interactions on the membrane potential and the ion fluxes is demonstrated for several values of diffusion current density and membrane charge density.     

Current-Voltage Curves of Porous Membranes in the Presence of Pore-Blocking Ions: I. Narrow Pores Containing No More Than One Moving Ion

We propose a physical model for voltage-dependent conductance changes of excitable cell membranes. It is based on competition of uni- and bivalent ions for chains of stable sites extending through the membrane. These one-dimensional pathways (pores) have different profiles of chemical potential for the two ionic species so that bivalent ions can block the passage of univalent ions at large membrane potentials. We treat the special case that each pore is either empty or, because of electrostatic repulsion, contains no more than one uni- or bivalent ion at a time. A system of linear differential equations describes the time-dependent probabilities of the various possible pore states. The states are limited by transition rate constants involving the profile of the chemical potential, the membrane voltage, the ionic concentrations in the adjacent baths, and electrostatic interactions between the ions. The steady-state solutions (Kirchhoff-Hill theorem) yield expressions for the relationship between the small signal conductance of univalent ions and the concentration of these ions in the external bathing medium (a saturation curve) and for the ionic currents and the steady-state current-voltage curve (N-shaped). From the latter curve we compute the shift of theshold potential caused by concentration changes of the external bathing medium. The model yields a number of predictions which can be tested experimentally.     

Specific ion effects on the growth rates of Staphylococcus aureus and Pseudomonas aeruginosa

Motivated by recent advances in the physical and chemical basis of the Hofmeister effect, we measured the rate cell growth of S. aureus--a halophilic pathogenic bacterium--and of P. aeruginosa, an opportunistic pathogen, in the presence of different aqueous salt solutions at different concentrations (0.2, 0.6 and 0.9 M). Microorganism growth rates depend strongly on the kind of anion in the growth medium. In the case of S. aureus, chloride provides a favorable growth medium, while both kosmotropes (water structure makers) and chaotropes (water structure breakers) reduce the microorganism growth. In the case of P. aeruginosa, all ions affect adversely the bacterial survival. In both cases, the trends parallel the specific ion, or Hofmeister, sequences observed in a wide range of physico-chemical systems. The correspondence with specific ion effect obtained in other studies, on the activities of a DNA restriction enzyme, of horseradish peroxidase, and of Lipase A (Aspergillus niger) is particularly striking. This work provides compelling evidence for Hofmeister effects, physical chemistry in action, in these organisms.     

Ion disturbance and clustering in the NaCl water solutions

Ion clustering and the solvation properties in the NaCl solutions are explored by molecular dynamics simulations with several popular force fields. The existence of ions has a negligible disturbance to the hydrogen bond structures and rotational mobility of water beyond the first ion solvation shells, which is suggested by the local hydrogen bond structures and the rotation times of water. The potential of mean force (PMF) of ion pair in the dilute solution presents a consistent view with the populations of ion clusters in the electrolyte solutions. The aggregation level of ions is sensitive to the force field used in the simulations. The ion-ion interaction potential plays an important role in the forming of the contact ion pair. The entropy of water increases as the ion pair approaches each other and the association of ion pair is driven by the increment of water entropy according to the results from the selected force fields. The kinetic transition from the single solvent separated state to the contact ion pair is controlled by the enthalpy loss of solution.

Velocity auto-correlation function of ions and water molecules in different concentrations,anions and ion clusters

The characteristics of ion solvation are important for electrochemical and biophysical phenomena because all such phenomena occur under the presence of solvated ions. In this study, we performed an all-atom molecular dynamics simulation of aqueous NaCl ranging from 0.5 to 3.0 M, and aqueous NaF, NaBr and NaI in 2.0 M, to investigate the time-averaged velocity auto-correlation function (TAVAF) of ions and water molecules. By comparing the concentrations and ion pairs, we observed three behaviours: (i) in the case of NaCl, the velocity auto-correlation of Cl^?  becomes weaker as the concentration increases, whereas those of Na⁺ are not clearly different, (ii) the intensity of fluctuations of the TAVAF gradually decreases following the decrease in ionic radius and (iii) every TAVAF of water molecules in ionic solutions is clearly lower than that of bulk because of the cage effect. Furthermore, we observed that the first minimum of the TAVAF in the cluster is smaller than that of the isolated ions. These results indicate that the diffusion of ions and water molecules is affected by cage effect, and that the generation of ion cluster affects the diffusion of ions.     

A method for controlling the activities of free metal,hydrogen, and phosphate ions in hydroponic solutions using ion exchange and chelating resins

Summary An ion exchange and chelating resin system was developed to buffer the activities of selected free cations and phosphate in hydroponic solutions at concentrations similar to those that occur naturally in soil solutions. Free-ion activities of Cd, Cu, Ni, Zn, Mn, H, Ca, Mg, and K were maintained by ion exchange and chelating resins in a controlled ionic strength system. Iron was controlled by EDDHA and chelating resin, and P by a cation-exchange resin containing adsorbed polynuclear hydroxyaluminum. This mixed-resin hydroponic system was used to establish a range of ionic Cd activities similar to that found in soil solutions of soils amended with sewage sludge. Activities of other nutrients were maintained at realistic soil-solution levels. A metal complexing agent (EDTA) was used to increase total metal concentration in the hydroponic solutions without significantly altering the ionic activities of the metals maintained in solution. This allowed differentiation of the effects of free ions and complexed metals on metal uptake. Concentrations of metal complexes in solution were controlled by the ion activities of the metals maintained and the concentrations and selectivities of the complexing agent. The mixed-resin system supplied sufficient nutrients for the growth of tomato plants (Lycopersicon esculentum, cv. Wisconsin-55) in hydroponic culture. Research supported by the College of Agricultural and Life Sciences, University of Wisonsin-Madison and by United States Environmental Protection Agency through Grant CR807270010.     

Futile cycling at the plasma membrane: a hallmark of low-affinity nutrient transport

Plant nutrient acquisition from concentrated soil solutions is governed by low-affinity transport systems in the plasma membranes of root cells. In this Opinion article, we illustrate that for six major nutrient ions, in addition to influx mediation by low-affinity transporters, high rates of ion cycling at the plasma membrane are a hallmark of nutrient transport at elevated external concentrations. This phenomenon is characterized by high rates of ion influx and concurrent high efflux of the same ion, resulting in efflux:influx ratios close to 1. Prolonged maintenance of futile cellular ion cycling can be energetically unfavorable and detrimental to plant growth and survival. We discuss how ion cycling can contribute to the toxicities of ions such as Na(+) and NH(4)(+) in the low-affinity range of ion provision. We also argue that cellular ion cycling makes the reliable measurement of ion influxes in the low-affinity range a formidable challenge.     

Rate theory models for ion transport through rigid pores. I. Time-dependent analysis in the case of vanishing interactions

In this first of a series of papers concerning the theoretical analysis of rate theory models for ion transport through rigid pores, the case of vanishing interactions is investigated. "Rigidity" means that ions crossing membranes through pores see a fixed structure of the pores, not changing in time. A single pore is considered to be a sequence of (n + 1) activation barriers separated by n energy minima. The explicit analytical treatment is restricted to pores with regular internal barrier structure, including the nonequilibrium situation of an applied electric field. In this case the connection with continuum diffusion models is demonstrated by performing in the limit n leads to infinity (n = number of binding sites within the pores) the transition to continuum. Thus, from diffusion equations describing a discrete number of jumps, the corresponding diffusion-like partial differential equations and boundary conditions are generated. For regular pores, from the time dependent solutions of the discrete equations, the corresponding solutions of the continuum equations are explicitly generated. The time-dependent relaxation behaviour of the discrete model is in good agreement with the continuum model if one assumes more than two binding sites in the pores.     

Interaction between quaternary ammonium ions in the pore of potassium channels. Evidence against an electrostatic repulsion mechanism

We have examined the interaction between internal and external ions in the pore of potassium channels. We found that external tetraethylammonium was able to antagonize block of Shaker channels by internal TEA when the external and internal solutions contained K(+) ions. This antagonism was absent in solutions with Rb(+) as the only permeant ion. An externally applied trivalent TEA analogue, gallamine, was less effective than the monovalent TEA in inhibiting block by internal TEA. In addition, block by external TEA was little affected by changes in the concentration of internal K(+) ions, but was increased by the presence of internal Na(+) ions in the pore. These results demonstrate that external and internal TEA ions, likely located at opposite ends of the pore selectivity filter, do not experience a mutual electrostatic repulsion. We found that these results can be simulated by a simple 4-barrier-3-site permeation model in which ions compete for available binding sites without long-range electrostatic interactions.     

Multi-ion conduction and selectivity in the high-conductance Ca++-activated K+ channel from skeletal muscle.

Open-channel ion permeation properties were investigated for Ca++-activated K+ (CaK) channels in solutions of K+ and its analogues T1+, Rb+, and NH4+. Single CaK channels were inserted into planar lipid bilayers composed of neutral phospholipids, and open-channel current-voltage (I-V) relations were measured in symmetrical and asymmetrical solutions of each of these individual ions. For all concentrations studied, the zero-voltage conductance falls in the sequence K+ greater than T1+ greater than NH4+ greater than Rb+. The shape of the I-V curve in symmetrical solutions of a single permeant ion is non-ohmic and is species-dependent. The I-V shape is sublinear for K+ and T1+ and superlinear for Rb+ and NH4+. As judged by reversal potentials under bi-ionic conditions with K+ on one side of the bilayer and the test cation on the other, the permeability sequence is T1+ greater than K+ greater than Rb+ greater than NH4+ at 300 mM, which differs from the conductance sequence. Symmetrical mixtures of K+ or NH4+ with Rb+ show a striking anomalous mole fraction behavior, i.e., a minimum in single-channel conductance when the composition of a two-ion mixture is varied at constant total ion concentration. This result is incompatible with present models that consider the CaK channel a single-ion pore. In total, the results show that the CaK channel finely discriminates among K+-like ions, exhibiting different energy profiles among these species, and that several such ions can reside simultaneously within the conduction pathway.     

Shear deformation effects in enzyme catalysis. Metal ion effect in the shear inactivation of urease.

The mechanism of the inactivation of the enzyme urease produced by subjecting its dilute solutions to hydrodynamic shear stresses in the range 0.5-2.5 Pa has been determined. By studying the kinetics of urease-catalyzed urea hydrolysis during application of hydrodynamic shear under varying chemical environments, we demonstrate that micromolar quantities of metal ions, in this case adventitious Fe, can accelerate the oxidation of thiol groups on urease and thus inactivate it when the protein is subjected to a shearing stress of order 1.0 Pa. In the absence of metal ion this stress level is ineffectual. It is proposed that this type of synergy between deformation and chemical environment may be crucial in many situations where biological macromolecules are subjected to mechanical stress.     

Evaluation of surface tension and ion occupancy effects on gramicidin A channel lifetime.

The surface tension of glycerylmonooleate-hexadecane lipid bilayer membranes and the lifetime of gramicidin A channels were measured at various concentrations of the surrounding solutions. For HCl the surface tension is essentially constant at approximately 5 mN/m up to approximately 1 M, whereas the average lifetime increases approximately 40-fold. At higher concentrations the surface tension decreases markedly. For CsCl the surface tension is constant up to about 1 M then increases with salt level. The average lifetime in this case increases about sixfold. In both cases the lifetime levels off and even decreases at higher salt levels. The increase in lifetime observed with ion activity is therefore qualitatively different from, and not explained by, the established dependence of lifetime on membrane properties (Elliot, J.R., D. Needham, J.P. Dilger, and D.A. Haydon. 1983. Biochim. Biophys. Acta. 735:95-103). We have previously proposed that ion occupancy is a determinant of channel stability, and to test this hypothesis the voltage dependence of channel lifetime was measured in asymmetrical solutions. For the case of a potassium chloride solution on one side of the membrane and a hydrogen chloride solution, on the other, the voltage dependence of the lifetime is asymmetrical. The asymmetry is such that when the electrical field is applied in the direction of the chemical gradient for each of the ions, the channel lifetime approaches, at increasing field strengths, that of a symmetrical solution of the respective ion. The voltage dependence of the surface tension, on the other hand, is negligible for the range of voltages used.(ABSTRACT TRUNCATED AT 250 WORDS)     

Tuning the voltage dependence of tetraethylammonium block with permeant ions in an inward-rectifier K+ channel.

To understand the role of permeating ions in determining blocking ion-induced rectification, we examined block of the ROMK1 inward-rectifier K+ channel by intracellular tetraethylammonium in the presence of various alkali metal ions in both the extra- and intracellular solutions. We found that the channel exhibits different degrees of rectification when different alkali metal ions (all at 100 mM) are present in the extra- and intracellular solution. A quantitative analysis shows that an external ion site in the ROMK1 pore binds various alkali metal ions (Na+, K+, Rb+, and Cs+) with different affinities, which can in turn be altered by the binding of different permeating ions at an internal site through a nonelectrostatic mechanism. Consequently, the external site is saturated to a different level under the various ionic conditions. Since rectification is determined by the movement of all energetically coupled ions in the transmembrane electrical field along the pore, different degrees of rectification are observed in various combinations of extra- and intracellular permeant ions. Furthermore, the external and internal ion-binding sites in the ROMK1 pore appear to have different ion selectivity: the external site selects strongly against the smaller Na+, but only modestly among the three larger ions, whereas the internal site interacts quite differently with the larger K+ and Rb+ ions.     

PADC nuclear track detector for ion spectroscopy in laser-plasma acceleration

The transparent polymer polyallyl-diglycol-carbonate (PADC), also known as CR-39, is widely used as detector for heavy charged particles at low fluence. It allows for detection of single protons and ions via formation of microscopic tracks after etching in NaOH or KOH solutions. PADC combines a high sensitivity and high specificity with inertness towards electromagnetic noise. Present fields of application include laser-ion acceleration, inertial confinement fusion, radiobiological studies with cell cultures, and dosimetry of nuclear fragments in particle therapy. These require precise knowledge of the energy-dependent response of PADC to different ion species. We present calibration data for a new type of detector material, Radosys RS39, to protons (0.2–3 MeV) and carbon ions (0.6–12 MeV). RS39 is less sensitive to protons than other types of PADC. Its response to carbon ions, however, is similar to other materials. Our data indicate that RS39 allows for measuring carbon ion energies up to 10 MeV only from the track diameters. In addition, it can be used for discrimination between protons and carbon ions in a single etching process.