Depolarization of the cell membrane causes inhibition of cell locomotion and pinocytosis inAcanthamoeba castellanii andAmoeba proteus

Summary 10 M CCCP protonophore in an acidic medium causes depolarization of the cell membrane and immediate cessation of locomotion inAcanthamoeba castellanii andAmoeba proteus. In the basic media there is no depolarization or inhibition of cell locomotion. Other depolarizing agents (alkali cations, crown molecules) also stop locomotion and induce pinocytosis in amoeba. Pinocytotic uptake of horseradish peroxidase byAcanthamoeba castellanii is increased by 69% in the presence of CCCP in the medium at pH 5.7 but is not influenced at higher pH values. This might indicate that both amoeboid locomotion and pinocytosis are controlled by membrane potential.     

Ionophores and intact cells I. Valinomycin and nigericin act preferentially on mitochondria and not on the plasma membrane of Saccharomyces cerevisiae

Valinomycin and nigericin prevented growth of 13 strains of the yeast Saccharomyces cerevisiae on non-fermentable substrate glycerol without affecting much fermentative growth on glucose. The two antibiotics did not induce swelling and lysis of yeast protoplasts in potassium acetate and did not modify uptake and release of Rb⁺ by the yeast cells. Both antibiotics were taken up by yeast cells at a relatively low rate. Nigericin accelerated the glucose-induced changes of fluorescence of a cyanine dye absorbed by yeast cells, which had been previously ascribed to a depolarization-repolarization cycle of the mitochondrial membrane. The data suggest that valinomycin and nigericin act as ionophores in the inner mitochondrial membrane and not in the plasma membrane of intact yeast cells.     

Highly robust lipid membranes on crystalline S-layer supports investigated by electrochemical impedance spectroscopy

In the present work, S-layer supported lipid membranes formed by a modified Langmuir-Blodgett technique were investigated by electrochemical impedance spectroscopy (EIS). Basically two intermediate hydrophilic supports for phospholipid- (DPhyPC) and bipolar tetraetherlipid- (MPL from Thermoplasma acidophilum) membranes have been applied: First, the S-layer protein SbpA isolated from Bacillus sphaericus CCM 2177 recrystallized onto a gold electrode; and second, as a reference support, an S-layer ultrafiltration membrane (SUM), which consists of a microfiltration membrane (MFM) with deposited S-layer carrying cell wall fragments. The electrochemical properties and the stability of DPhyPC and MPL membranes were found to depend on the used support. The specific capacitances were 0.53 and 0.69 μF/cm² for DPhyPC bilayers and 0.75 and 0.77 μF/cm² for MPL monolayers resting on SbpA and SUM, respectively. Membrane resistances of up to 80 MΩ cm² were observed for DPhyPC bilayers on SbpA. In addition, membranes supported by SbpA exhibited a remarkable long-term robustness of up to 2 days. The membrane functionality could be demonstrated by reconstitution of membrane-active peptides such as valinomycin and alamethicin. The present results recommend S-layer-supported lipid membranes as promising structures for membrane protein-based biosensor technology.     

Differential effects of 2,4-dinitrophenol and valinomycin (+K+) on uncoupler-stimulated ATPase of human tumor mitochondria

The uncoupler-stimulated mitochondrial ATPase of four human tumors, mouse kidney, brain and fetal liver exhibited a characteristic behavior when preincubated with the H⁺-conducting uncouplers, dinitrophenol, CCCP, S-13 and gramicidin. The ATPase activity was considerably lower with preincubation than without. Preincubation with valinomycin (+K⁺), on the other hand, did not result in a significant decrease of the ATPase activity. These results may be contrasted with those obtained with liver or heart mitochondria, the ATPase activity of which did not suffer any loss when preincubated with dinitrophenol. The effect of preincubation with dinitrophenol on the tumor mitochondria could not be accounted for by dinitrophenol-induced Mg²⁺ efflux, since the differential effects of dinitrophenol and valinomycin (+K⁺) remained even when ATPase activity was determined in presence of Mg²⁺. Small amounts of ATP and ADP in the preincubation mixture containing dinitrophenol protected against the decay of the ATPase activity, implicating the exchangeable adenine nucleotides in the tumor mitochondria. In a model system where liver mitochondria were depleted of their adenine nucleotides, a lower ATPase activity was indeed obtained. However, direct determination of the concentations of adenine nucleotides in dinitrophenol- and valinomycin-treated tumor mitochondria revealed only slight differences.     

Location of valinomycin in lipid vesicles

The location of the cyclododecadepsipeptide, valinomycin in vesicles formed from two synthetic lipids is studied by differential scanning calorimetry, spin-label partitioning electron paramagnetic resonance and [¹H]-nuclear magnetic resonance. The results show that valinomycin is located near the head group region of dipalmitoyl phosphatidyl choline vesicles and in the hydrophobic core of the dimyristoyl phosphatidyl choline vesicles in the liquid crystalline phase.