Alkaline Earth Divalent Cations Research Articles

Divalent Cation Modulation of Ion Permeation in TMEM16 Proteins.

Intracellular divalent cations control the molecular function of transmembrane protein 16 (TMEM16) family members. Both anion channels (such as TMEM16A) and phospholipid scramblases (such as TMEM16F) in this family are activated by intracellular Ca2+ in the low µM range. In addition, intracellular Ca2+ or Co2+ at mM concentrations have been shown to further potentiate the saturated Ca2+-activated current of TMEM16A. In this study, we found that all alkaline earth divalent cations in mM concentrations can generate similar potentiation effects in TMEM16A when applied intracellularly, and that manipulations thought to deplete membrane phospholipids weaken the effect. In comparison, mM concentrations of divalent cations minimally potentiate the current of TMEM16F but significantly change its cation/anion selectivity. We suggest that divalent cations may increase local concentrations of permeant ions via a change in pore electrostatic potential, possibly acting through phospholipid head groups in or near the pore. Monovalent cations appear to exert a similar effect, although with a much lower affinity. Our findings resolve controversies regarding the ion selectivity of TMEM16 proteins. The physiological role of this mechanism, however, remains elusive because of the nearly constant high cation concentrations in cytosols.

Effects of Cation Binding to the Intracellular Vestibule of TMEM16 Ion Transport Pathways

TMEM16A and TMEM16F are prototype molecules for the two functional categories of TMEM16 family members, respectively, Ca2+-activated chloride channels and phospholipid scramblases. Upon binding of sub-µM to low µM intracellular Ca2+, TMEM16A conducts anionic currents while TMEM16F is permeable to both cations and anions. In this study, we found another divalent cation, Co2+, while unable to induce current in TMEM16A, inhibits the Ca2+-induced current via competing with Ca2+ for the high-affinity (sub-µM to low µM) channel activation sites. In addition, Co2+ also potentiates the TMEM16A current induced by Ca2+. All alkaline earth divalent cations, including Ca2+ itself, can generate this current potentiation with low affinities—at least hundreds of µM to mM concentrations are required for observing the effect. On the other hand, divalent cations at mM concentrations generate very little potentiation in TMEM16F, likely because the changes in local anion and cation concentrations caused by divalent cation bindings cancel out each other. Nonetheless, divalent cations change the cation/anion selectivity for the TMEM16F current conduction. Manipulating membrane phospholipids alters this low-affinity effect of divalent cations. Recent studies have shown that membrane phospholipids participate in various gating and permeation functions of TMEM16 molecules. Our results suggest that intracellular divalent cations bind to membrane phospholipids with a low affinity to alter the electrostatic potential near or in the ion-transport pathways of TMEM16 molecules. Monovalent cations and polyamine molecules also exert similar effects, although with different apparent affinities. The physiological significance of the TMEM16 current modulation by altering membrane surface potential will be discussed.

Ion-Specific Effects of Divalent Ions on the Structure of Polyelectrolyte Brushes.

Polyelectrolyte brushes consist of charged polymer chains attached on one end to a surface at high densities. They are relevant for many practical applications ranging from biosensors to drug delivery to colloidal stability. Their structure and functionality can be dramatically influenced by multivalent counterions in the solution environment. In this work, the surface forces apparatus (SFA) and atomic force microscopy (AFM) were used to investigate the effects of three alkaline earth divalent cations, Mg2+, Ca2+, and Ba2+, on the structures of polystyrenesulfonate (PSS) brushes tethered to mica and silicon oxide surfaces. While all these ions caused significant shrinkage of the height of the PSS brushes, strong ion-specific effects were observed. Mg2+ and Ca2+ caused homogeneous shrinkage; Ba2+ led to pinned-micelle like inhomogeneous structures. Isothermal titration calorimetry (ITC) demonstrated that this ion specificity was mainly caused by the difference in binding energy between sulfonate groups and the divalent cations. Considering the abundance of divalent cations in industrial processes, natural environments, and biological systems, the understanding of strong ion-specific effects of divalent counterions is of great importance for theoretical studies and various applications involving polyelectrolytes.

Theoretical Studies on Selectivity of Dibenzo-18-Crown-6-Ether for Alkaline Earth Divalent Cations

Crown ether is one of well-known host molecules and able to selectively sequester metal cation. We employed M06-2X density functional theory with IEFPCM and SMD continuum solvation models to study selectivity of dibenzo-18-crown-6-ether (DB18C6) for alkaline earth dications, , , , and in the gas phase and in aqueous solution. showed predominantly strong binding affinity in the gas phase because of strong polarization of CO bonds by cation. In aqueous solution, binding free energy differences became smaller among these dications. However, had the best binding, being incompatible with experimental observations in aqueous solution. The enthalpies of the dication exchange reaction between DB18C6 and water cluster molecules were computed as another estimation of selectivity in aqueous solution. These results also demonstrated that bound to DB18C6 better than . We speculated that the species determining selectivity in water could be 2:1 complexes of two DB18C6s and one dication.

Divalent counterion-induced condensation of triple-strand DNA

Understanding and manipulation of the forces assembling DNA/RNA helices have broad implications for biology, medicine, and physics. One subject of significance is the attractive force between dsDNA mediated by polycations of valence ≥ 3. Despite extensive studies, the physical origin of the "like-charge attraction" remains unsettled among competing theories. Here we show that triple-strand DNA (tsDNA), a more highly charged helix than dsDNA, is precipitated by alkaline-earth divalent cations that are unable to condense dsDNA. We further show that our observation is general by examining several cations (Mg(2+), Ba(2+), and Ca(2+)) and two distinct tsDNA constructs. Cation-condensed tsDNA forms ordered hexagonal arrays that redissolve upon adding monovalent salts. Forces between tsDNA helices, measured by osmotic stress, follow the form of hydration forces observed with condensed dsDNA. Probing a well-defined system of point-like cations and tsDNAs with more evenly spaced helical charges, the counterintuitive observation that the more highly charged tsDNA (vs. dsDNA) is condensed by cations of lower valence provides new insights into theories of polyelectrolytes and the biological and pathological roles of tsDNA. Cations and tsDNAs also hold promise as a model system for future studies of DNA-DNA interactions and electrostatic interactions in general.

Size mismatch: A crucial factor for generating a spin-glass insulator in manganites

The structural, electronic, and magnetic properties of the highly mismatched perovskite oxides, ${\mathrm{Th}}_{0.35}{A}_{0.65}{\mathrm{MnO}}_{3},$ where A is for the alkaline earth divalent cations (Ca, Ba, Sr), which are all characterized by the same large tolerance factor $(t=0.934),$ have been investigated by using electron microscopy, electrical resistivity, magnetic susceptibility, and magnetization. It is clearly established that a transition from ferromagnetic metallic towards spin-glass insulator samples is induced as the A-site cationic size mismatch is increased. Moreover, the magnetoresistance (MR) properties of these manganites are strongly reduced for the spin-glass insulators, demonstrating that the A-site cationic disorder is detrimental for the colossal MR properties. Based on these results, a new electronic and magnetic diagram is established that shows that the A-site disorder, rather than the A-site average cationic size (or t) is the relevant factor for generating spin-glass insulating manganites.

Differences in cation sensitivity of ligand binding to Y 1 and Y 2 subtype of neuropeptide Y receptor of rat brain

The binding of selective ligands to the Y 1 subtype of neuropeptide Y receptor in rat brain particulates was promoted by Ca 2+ and also stimulated by Sr 2+, but reversibly reduced by Ba 2+, Mg 2+, Mn 2+, by the organic polycations neomycin and spermidine, and by chelating agents. The alkali monovalent cations inhibited the Ca 2+-enabled Y 1 subtype binding with some selectivity (Cs +≥NH 4 +>Li +>Na +,K +), with half-inhibition between 70–120 mM. The specific Y 2 subtype binding was enhanced by all alkaline-earth divalent cations, Mn 2+, neomycin and spermidine in the range of 0.1–10 mM, and by alkali cations at up to 100 mM, and also by Na + salts of the chelators EGTA and EDTA. The large disparity in cation sensitivity indicates substantial differences in the structure of the binding sites of the Y 1 and Y 2 receptors, predictable from known distinct features of ligand epitopes and of primary structure of the receptors.

Divalent cation conduction in the ryanodine receptor channel of sheep cardiac muscle sarcoplasmic reticulum.

The conduction properties of the alkaline earth divalent cations were determined in the purified sheep cardiac sarcoplasmic reticulum ryanodine receptor channel after reconstitution into planar phospholipid bilayers. Under bi-ionic conditions there was little difference in permeability among Ba2+, Ca2+, Sr2+, and Mg2+. However, there was a significant difference between the divalent cations and K+, with the divalent cations between 5.8- and 6.7-fold more permeant. Single-channel conductances were determined under symmetrical ionic conditions with 210 mM Ba2+ and Sr2+ and from the single-channel current-voltage relationship under bi-ionic conditions with 210 mM divalent cations and 210 mM K+. Single-channel conductance ranged from 202 pS for Ba2+ to 89 pS for Mg2+ and fell in the sequence Ba2+ greater than Sr2+ greater than Ca2+ greater than Mg2+. Near-maximal single-channel conductance is observed at concentrations as low as 2 mM Ba2+. Single-channel conductance and current measurements in mixtures of Ba(2+)-Mg2+ and Ba(2+)-Ca2+ reveal no anomalous behavior as the mole fraction of the ions is varied. The Ca(2+)-K+ reversal potential determined under bi-ionic conditions was independent of the absolute value of the ion concentrations. The data are compatible with the ryanodine receptor channel acting as a high conductance channel displaying moderate discrimination between divalent and monovalent cations. The channel behaves as though ion translocation occurs in single file with at most one ion able to occupy the conduction pathway at a time.

Characterization of cationic binding sites of neurotoxins from venom of the scorpion ( Centruroides sculpturatus Ewing) using lanthanides as binding probes

R. M. David, N. R. Krishna and D. D. Watt. Characterization of cationic binding sites of neurotoxins from venom of the scorpion ( Centruroides sculpturatus Ewing) using lanthanides as binding probes. Toxicon 29, 645–662, 1991.—Binding sites for cations were probed in the structures of protein neurotoxins from Centruroides sculpturatus by enhancement of terbium(III) fluorescence, detected by emission at 552 nm, when aromatic side-chains of the toxins were activated at 286 nm. Gadolinium, Gd(III), was used as a cation binding probe by observing its effects on nuclear magnetic resonance (NMR) spectra. Toxins CsE-v2 and v3, when bound to Tb(III), enhance luminescence of Tb(III) 20-fold whereas CsE-v1 enhances Tb(III) luminescence about 15-fold. Toxins CsE-I and V have no effect on the luminescence of Tb(III) implying that these latter two toxins have structures incompatible with efficient energy transfer from activated aromatic side-chains. Enhancement of fluorescence is pH dependent and is competitively inhibited by alkaline earth divalent cations and by other lanthanide(III) ions. Neodymium, Nd(III), with an ionic radius of 0.995 Å is the most efficient of the lanthanide ions and the divalent cations in displacement of Tb(III) from the toxins. Relaxation enhancements of aromatic C H resonances by Gd(III) are apparent with tyrosines 4, 42, 38, 14–40 peak and tryptophan 47. Results from pH vs fluorescence studies suggest that carboxyl groups are involved in binding of Tb(III). Association constants ( K a) of the Tb(III)-CsE-v2 and v3 complexes are respectively 2.5 × 10 3 and 2.4 × 10 3 M −1 determined by fluoresence enhancement and 2.4 × 10 3 and 2.3 × 10 3 M −1 by equilibrium dialysis. Similarly K a values for toxins CsE I and V are respectively 1.9 × 10 3 and 1.8 × 10 3 M −1 determined by equilibrium dialysis. Experimental evidence suggests that at least two Tb(III)s are bound per toxin molecule. The results from these studies are discussed in relation to the tertiary structure of toxin CsE-v3.