Effect Of Transmembrane Potential Research Articles

Cell type Identification based on Ion Transmembrane Transport Considering Extracellular Electrical Double Layer (-Effect of Transmembrane Potential by Cell Membrane Electric Double Layer)

Cell type Identification based on Ion Transmembrane Transport Considering Extracellular Electrical Double Layer (-Effect of Transmembrane Potential by Cell Membrane Electric Double Layer)

Effects of transmembrane potential and pH gradient on the cytochrome c-promoted fusion of mitochondrial mimetic membranes

The present study investigated the effects of ΔΨ and ΔpH (pH gradient) on the interaction of cytochrome c with a mitochondrial mimetic membrane composed of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cardiolipin (CL) leading to vesicle fusion. ΔpH generated by lowered bulk pH (pH(out)) of PCPECL liposomes, with an internal pH (pH(in)) of 8.0, favored vesicle fusion with a titration sigmoidal profile (pK(a) ~ 6.9). Conversely, ΔpH generated by enhanced pH(in) of PCPECL at a pH(out) of 6.0 favored the fusion of vesicles with a linear profile. We did not observe a significant amount of liposome fusion when ΔpH was generated by lowered pH(in) at a pH(out) of 8.0. At bulk acidic pH, ΔΨ generated by Na⁺ gradient also favored cyt c-promoted vesicle fusion. At acidic and alkaline pH(out), the presence of ΔpH and ΔΨ did not affect cytochrome c binding affinity measured by pyrene quenching. Therefore, cytochrome c-mediated PC/PE/CL vesicle fusion is dependent of ionization of the protein site L (acidic pH) and the presence of transmembrane potential. The effect of transmembrane potential is probably related to the generation of defects on the lipid bilayer. These results are consistent with previous reports showing that cytochrome c release prior to the dissipation of the ΔΨ(M) blocks inner mitochondrial membrane fusion during apoptosis.

Near-IR Resonance Raman Spectroscopy of Archaerhodopsin 3: Effects of Transmembrane Potential

Archaerhodopsin 3 (AR3) is a light driven proton pump from Halorubrum sodomense that has been used as a genetically targetable neuronal silencer and an effective fluorescent sensor of transmembrane potential. Unlike the more extensively studied bacteriorhodopsin (BR) from Halobacterium salinarum, AR3 readily incorporates into the plasma membrane of both E. coli and mammalian cells. Here, we used near-IR resonance Raman confocal microscopy to study the effects of pH and membrane potential on the AR3 retinal chromophore structure. Measurements were performed both on AR3 reconstituted into E. coli polar lipids and in vivo in E. coli expressing AR3 in the absence and presence of a negative transmembrane potential. The retinal chromophore structure of AR3 is in an all-trans configuration almost identical to BR over the entire pH range from 3 to 11. Small changes are detected in the retinal ethylenic stretching frequency and Schiff Base (SB) hydrogen bonding strength relative to BR which may be related to a different water structure near the SB. In the case of the AR3 mutant D95N, at neutral pH an all-trans retinal O-like species (O(all-trans)) is found. At higher pH a second 13-cis retinal N-like species (N(13-cis)) is detected which is attributed to a slowly decaying intermediate in the red-light photocycle of D95N. However, the amount of N(13-cis) detected is less in E. coli cells but is restored upon addition of carbonyl cyanide m-chlorophenyl hydrazone (CCCP) or sonication, both of which dissipate the normal negative membrane potential. We postulate that these changes are due to the effect of membrane potential on the N(13-cis) to M(13-cis) levels accumulated in the D95N red-light photocycle and on a molecular level by the effects of the electric field on the protonation/deprotonation of the cytoplasmic accessible SB. This mechanism also provides a possible explanation for the observed fluorescence dependence of AR3 and other microbial rhodopsins on transmembrane potential.

Structure and Orientation of the Mammalian Antibacterial Peptide Cecropin P1 within Phospholipid Membranes

Cecropins are positively charged antibacterial peptides that act by permeating the membrane of susceptible bacteria. To gain insight into the mechanism of membrane permeation, the secondary structure and the orientation within phospholipid membranes of the mammalian cecropin P1 (CecP) was studied using attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy and molecular dynamics simulations. The shape and frequency of the amide I and II absorption peaks of CecP within acidic PE/PG multibilayers (phosphatidylethanolamine/phosphatidylglycerol) in a 7:3 (w/w) ratio (a phospholipid composition similar to that of many bacterial membranes), indicated that the peptide is predominantly α-helical. Polarized ATR-FTIR spectroscopy was used to determine the orientation of the peptide relative to the bilayer normal of phospholipid multibilayers. The ATR dichroic ratio of the amide I band of CecP peptide reconstituted into oriented PE/PG phospholipid membranes indicated that the peptide is preferentially oriented nearly parallel to the surface of the lipid membranes. A similar secondary structure and orientation were found when zwitterionic phosphatidylcholine phospho lipids were used. The incorporation of CecP did not significantly change the order parameters of the acyl chains of the multibilayer, further suggesting that CecP does not penetrate the hydrocarbon core of the membranes. Molecular dynamics simulations were used to gain insight into possible effects of transmembrane potential on the orientation of CecP relative to the membrane. The simulations appear to confirm that CecP adopts an orientation parallel to the membrane surface and does not insert into the bilayer in response to a cispositive transmembrane voltage difference. Taken together, the results further support a “carpet-like” mechanism, rather than the formation of transmembrane pores, as the mode of action of CecP. According to this model, formation of a layer of peptide monomers on the membrane surface destablizes the phospholipid packing of the membrane leading to its eventual disintegration.

P-C4-14 The effect of transmembrane potential on the structure of lipid bilayer: Malinsky J., Herman P., Vecer J., Plasek J. Institute of Physics, Charles University, Prague (CZ)

P-C4-14 The effect of transmembrane potential on the structure of lipid bilayer: Malinsky J., Herman P., Vecer J., Plasek J. Institute of Physics, Charles University, Prague (CZ)

Transmembrane reduction of methylviologen by ascorbate photosensitized by bacteriopheophytin a in liposomes. Effect of transmembrane potential

Operation of a bacteriopheophytin a (BPh) reductive mechanism for electron transfer in liposome media has been demonstrated. The electron transfer through the liposome wall can be accomplished mainly by an electron exchange mechanism between BPh in − and BPh out at opposing sides of the membrane. The rate of electron transfer across the bilayer and the rate of backward recombination of BPh in − and ascorbate radicals on the inner liposome surface are influenced by the generation of transmembrane potentials with K + gradients in the presence of valinomycin. The ratio of the transmembrane electron transfer rate constants was measured at positive, negative and zero transmembrane potentials by laser photolysis.

Glycine transport by intestinal brush border vesicles of the amphibian Discoglossus pictus

1. 1. In order to determine the different components of glycine uptake by the intestine of the frog, Discoglossus plctus, we have used brush border membrane vesicles isolated by a classical precipitation technique. 2. 2. Enzymatic tests showed that a good purification was obtained. The concentration ratio of alkaline phosphatase was 14.8. 3. 3. Glycine entry in vesicles as a function of time, in presence or absence of sodium, indicated an overshoot which decreased when incubation time was prolonged. The overshoot was dependent on the presence of sodium. 4. 4. The nature of the anion associated to sodium had little effect on glycine uptake. Nevertheless, chloride and thiocyanate appeared more efficient than glutarate. 5. 5. The effect of transmembrane potential was studied by using valinomycin associated with a potassium gradient. The addition of this substance stimulated glycine transport by 43%. 6. 6. The transport at different glycine concentrations showed two components: one non-saturable with weak affinity and the other saturable with strong affinity ( K t = 0.338 mM). 7. 7. In conclusion, glycine transport by the brush border of D. plctus intestine presents a saturable component depending on sodium and on transmembrane electrical potential.

The effect of transmembrane potential on the dynamic behavior of cell membranes

The relationship between transmembrane potential and lipid dynamics in the cytoplasmic membrane of mouse thymus cells has been investigated. Changes of transmembrane potential was followed by measuring the fluorescence emission of the anionic dye, bis-(1,3-dibutylbarbiturate)trimethine exonol (diBa-C 4-(3)). Assessment of lipid fluidity was carried out applying three fluorescent lipid probes, 1-[4-(trimethylammonium)phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH), 12-(9-anthroyloxy)stearic acid (12-AS) and 1,6-diphenyl-1,3,5-hexatriene (DPH) used to monitor different structural regions of the bilayer. The fluorescence anisotropy of these probes was measured as a function of temperature at two values of transmembrane potential. In the case of DPH it proved to depend on the membrane potential in the higher temperature range (above 28°C), while no such a dependence could be observed for DPH below this temperature range and for TMA-DPH and 12-AS in between 20 and 37°C. These data suggest that changes in transmembrane potential are accompanied with some local alteration in membrane lipid dynamics and/or structure.

Effect of transmembrane potential on the deformability of RBC's suspended in carbohydrate-saline solutions

The effect of transmembrane potential on the deformability of red blood cells (RBC) suspended in isotonic glucose-saline and galactose-saline solutions was determined using the micropipette aspiration technique. In this method, a negative pressure was applied to the RBC membrane via a micropipette and the resulting deformation was analyzed using a Kelvin model to yield a membrane elastic modulus E. When glucose concentration was increased from 0 to 0.3 M/dm 3 in the extracellular media, the membrane elastic modulus increased seven-fold ( p < 0.0001) with most increase occurring in the 0–0.1 M/dm 3 range. Whereas, galactose had no significant effect on the membrane elasticity up to a concentration of 0.28 M/dm 3. However, in pure galactose (isotonic) solution (0.308 M/dm 3), the membrane elastic modulus decreased by a factor of 3 ( p < 0.001). Increase in the RBC elastic modulus in the presence of glucose may be due to binding of glucose to the membrane and intracellular proteins, and decrease in the presence of pure galactose may be due to the transmembrane potential generated by chloride ions.

Effect of transmembrane potential on the manifestations of ouabain toxicity in sheep cardiac Purkinje fibres.

The effect of membrane potential on the manifestations of ouabain-induced alterations in the electrical activity of shortened sheep cardiac Purkinje fibres was studied by a microelectrode technique. The level of membrane potential was modulated by passing intracellular polarizing current. It was found that the toxic electrical effects induced by exposure to ouabain vary according to the level of membrane potential.