gramicidin-a and tetraphenylphosphonium

The possibility that simple lipophilic cations such as tetraphenylphosphonium (TPA+), triphenylmethylphosphonium (TPMP+), and diphenyldimethylphosphonium (DDP+) are substrates for the multidrug-resistance transport protein, P-glycoprotein, was tested. Hamster cells transfected with and overexpressing mouse mdr1 or mouse mdr3 exhibit high levels of resistance to TPP+ and TPA+ (20-fold) and somewhat lower levels of resistance to TPMP+ and DDP+ (3-12-fold). Transfected cell clones expressing mdr1 or mdr3 mutants with decreased activity against drugs of the MDR spectrum (e.g., Vinca alkaloids and anthracyclines) also show reduced resistance to lipophilic cations. Studies with radiolabeled TPP+ and TPA+ demonstrate that increased resistance to cytotoxic concentrations of these lipophilic cations is correlated quantitatively with a decrease in intracellular accumulation in mdr1- and mdr3-transfected cells. This decreased intracellular accumulation is shown to be strictly dependent on intact intracellular nucleotide triphosphate pools and is reversed by verapamil, a known competitive inhibitor of P-glycoprotein. Taken together, these results demonstrate that lipophilic cations are a new class of substrates for P-glycoprotein and can be used to study its mechanism of action in homologous and heterologous systems.

Topics: Adenosine Triphosphate; Animals; Arsenicals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Cations; Cell Survival; CHO Cells; Cricetinae; Electrophoresis, Polyacrylamide Gel; Gramicidin; Membrane Glycoproteins; Mutation; Onium Compounds; Organophosphorus Compounds; Substrate Specificity; Trityl Compounds; Verapamil

Two diamines were prepared to investigate the structure-activity relationship required for an increase in the permeability of the outer membrane of Escherichia coli. It was found that diamine (a), bis[4-(2-methylaminoethoxy)phenyl]methane dihydrochloride, increased the permeability of the membrane, while diamine (b), 1,4-bis(2-methylaminoethoxy)benzene dihydrochloride, did not. The result indicated that the existence of bulky hydrophobic moiety is important to cause an increase in the permeability.

Topics: Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Membrane Permeability; Diamines; Escherichia coli; Gramicidin; Kinetics; Nitriles; Onium Compounds; Organophosphorus Compounds; Potassium

The rate of protonophore-mediated decay of pH gradient across lipid vesicular membranes was found to be enhanced by orders of magnitude by valinomycin-K+. Experiments in the presence of gramicidin have shown that the observed rate enhancement by valinomycin-K+ is not due to collapse of the diffusion potential alone. The enhancement of the rate showed hyperbolic dependence on the concentration of valinomycin. Rate enhancement was observed in the presence of the membrane permeant cation tetraphenylphosphonium (TTP+) also. Several factors which might enhance the intrinsic H+ conductivity of protonophores were analyzed. The level of partitioning of the protonophore into the membrane and the pK of membrane-bound protonophores were measured. Valinomycin-K+ did not alter both these parameters significantly. TPP+ increased the partitioning of protonophores and decreased the pK values of membrane-bound protonophores. However, these changes were too small to explain the observed rate enhancements. We suggest that valinomycin-K+ and TPP+ enhance the H+ conductivity of protonophores by increasing the permeability of the ionized form of protonophores by forming an ion pair.

Topics: Boron Compounds; Cations; Cell Membrane Permeability; Gramicidin; Hydrogen; Hydrogen-Ion Concentration; Kinetics; Onium Compounds; Organophosphorus Compounds; Potassium; Tetraphenylborate; Valinomycin

Topics: Animals; Cell Membrane; Diffusion; Gramicidin; Humans; In Vitro Techniques; Ions; Membrane Potentials; Mice; Onium Compounds; Organophosphorus Compounds; Ouabain; Phenytoin; Potassium; Thiocyanates

Using the penetrating ions of tetraphenylphosphonium (TPP+) and tetraphenylborone (TPB-), the membrane potential of the Bacillus subtilis and Escherichia coli cells was shown that the TPP+ absorption by the cells is an energy-coupled process. The TPB- anions are released from the cells after addition of an energy substrate. The value of the membrane potential calculated from the distribution pattern of the penetrating ions in the cells and the incubation medium lies within the interval of --100--150 mV (intracellular negative electric potential). The value of the membrane potential strongly depends on pH of the incubation medium; our attempts to measure the membrane potential in the E. coli cells at ph 6.0 were unsuccessful; however, at pH 8.5 it was found to be equal to --100 mV. Treatment of the cells with nigericin partially prevents the decrease of the membrane potential in an acidic medium and increases the potential in neutral and alkaline media. The formation of the membrane potential is suppressed by valinomycin and gramicidine, as well as by the oxidative phosphorylation uncouplers; the inhibiting effect of valinomycin requires the presence of K+ in the incubation medium. The membrane potential of the B. subtilis cells is insensitive to the effect of cyanide in the absence of arsenate. It is concluded that the membrane potential of B. subtilis and E. coli is formed both via respiration and by hydrolysis of intracellular ATP.

Topics: Bacillus subtilis; Boron Compounds; Escherichia coli; Gramicidin; Hydrogen-Ion Concentration; Kinetics; Membrane Potentials; Onium Compounds; Organophosphorus Compounds; Tetraphenylborate; Valinomycin