Effect of dolichyl monophosphate on the permeability properties of lipid membranes

The effect of dolichyl monophosphate on the permeability properties of dimyristoylphosphatidylcholine bilayers to alkaline cations, Ca²⁺ and glucose has been determined by stop-flow spectrophotometry. The results show that, in con trast to free dolichol effects, the monophosphate derivative increased the permeability following a decreasing order of the permeating particle size. Phase diagrams indicate that dolichyl monophosphate is fully incorporated into the phosphatidylcholine bilayer around 0.75% weight/weight ratio. For these ratios, the permeation of ions is higher in the gel than in the liquid crystalline state.     

Influence of dolichyl phosphate on permeability and stability of bilayer lipid membranes

The ionic permeability coefficients, ionic transference numbers, activation energy of ion transport and breakdown voltage of bilayer lipid membranes made from dioleoylphosphatidylcholine or its mixtures with dolichyl 12-phosphate have been studied. The electrical measurements showed that dolichyl phosphate in phospholipid bilayers decreases membrane permeability, changes membrane ionic selectivity and increases membrane stability. These results are discussed in light of the aggregation behavior and the intramolecular clustering of a dolichyl phosphate molecule in phospholipid membranes. From our data we suggest that the hydrophilic part of dolichyl phosphate molecules regulates their behavior in membranes.     

Voltage-dependent behaviour of dolichyl phosphate-phosphatidylcholine bilayer lipid membranes

The current-voltage steady-state characteristics, cyclic voltammograms and capacitance-voltage steady-state relationships of bilayer lipid membranes made from dioleoylphosphatidylcholine or its mixtures with dolichyl-12 phosphate have been studied. Sustained fluctuations of the capacitance of dolichyl phosphate modified bilayers under applied voltage were observed. The results suggest that the dynamics of dolichyl phosphate molecules in membranes can be regulated by transmembrane electrical potential.     

Effect of phloretin on the permeability of thin lipid membranes

Phloretin dramatically increases cation conductances and decreases anion conductances of membranes treated with ion carriers (nonactin, valinomycin, carbonyl-cyanide-m-chlorophenylhydrazone [CCCP], and Hg(C6F5)2) or lipophilic ions (tetraphenylarsonium [tphAs+] and tetraphenylborate [TPhB-]). For example, on phosphatidylethanolamine membranes, 10(-4) M phloretin increases K+ -nonactin and TPhAs+ conductances and decreases CCCP- and TPhB- conductances 10(3)-fold; on lecithin: cholesterol membranes, it increases K+-nonactin conductance 10(5)-fold and decreases CCCP- conductance 10(3)-fold. Similar effects are obtained with p- and m-nitrophenol at 10(-2) M. These effects are produced by the un-ionized form of phloretin and the nitrophenols. We believe that phloretin, which possesses a large dipole moment, adsorbs and orients at the membrane surface to introduce a dipole potential of opposite polarity to the preexisting positive one, thus increasing the partition coefficient of cations into the membrane interior and decreasing the partition coefficient of anions. (Phloretin may also increase the fluidity of cholesterol-containing membranes; this is manifested by its two- to three-fold increase in nonelectrolyte permeability and its asymmetrical effect on cation and anion conductances in cholesterol-containing membranes.) It is possible that pholoretin's inhibition of chloride, urea, and glucose transport in biological membranes results from the effects of these intense intrafacial dipole fields on the translocator(s) of these molecules.     

Enzymatic dephosphorylation of endogenous and exogenous dolichyl monophosphate by calf brain membranes

Calf brain membranes have been shown to enzymatically dephosphorylate endogenous and partially purified, exogenous dolichyl [³²P]monophosphate. The properties and specificity of the dolichyl monophosphatase activity have been studied by following the release of [³²P]phosphate from exogenous dolichyl [³²P]monophosphate added in a dispersion with Triton X-100. The calf brain phosphatase (1) is inhibited by Mn²⁺, Mg²⁺, Ca²⁺, fluoride, and phosphate; (2) exhibits a neutral pH optimum; and (3) has an apparent K_m of 200 μm for dolichyl monophosphate. Dolichyl monophosphatase activity can be distinguished from phosphatidate phosphatase on the basis of their responses to fluoride and phosphate. Based on differential thermolability and the effects of divalent cations and EDTA, the calf brain dolichyl monophosphatase can also be discriminated from the general phosphatase activity assayed with p-nitrophenyl phosphate. Dolichyl monophosphatase activity can be solubilized by treating microsomes with Triton X-100. The enzymatic dephosphorylation of exogenous dolichyl [³²P]monophosphate catalyzed by particulate and detergent-solubilized preparations is negligibly affected by equimolar concentrations of ATP and an assortment of phosphomonoesters, including phosphatidic acid and hexadecyl phosphate. A reduction of approximately 40% in dolichyl monophosphatase activity is observed in the presence of equimolar amounts of retinyl monophosphate. Overall, these results represent good evidence for the presence of a neutral polyisoprenyl monophosphatase in central nervous tissue.     

Effect of peptide PV on the ionic permeability of lipid bilayer membranes

This paper reports the effects of peptide PV (primary structure: cyclo-(D-val-L-pro-L-val-D-pro)_δ) on the electrical properties of sheep red cell lipid bilayers. The membrane conductance (G_m) induced by PV in either Na⁺ or K⁺ medium is proportional to the concentration of PV in the aqueous phase. The PV concentration required to produce a comparable increase in G_m in K⁺ medium is about 10⁴ times greater than for its analogue, valinomycin (val). Although the selectivity sequence for PV and val is similar, K⁺ ≳ Rb⁺ > Cs⁺ > NH₄ ⁺ > TI⁺ > Na⁺ > Li⁺; the ratio of GGm in K⁺ to that in Na⁺ is about 10 for PV compared to > 10³ for val. When equal concentrations of PV are added to both sides of a bilayer, the membrane current approaches a maximum value independent of voltage when the membrane potential exceeds 100 mV. When PV is added to only one side of a bilayer separating identical salt solutions of either Na⁺ or K⁺ salts, rectification occurs such that the positive current flows more easily away rather than toward the side containing the carrier. Under these conditions, a large, stable, zero-current potential (VVm) is also observed, with the side containing PV being negative. The magnitude of this VVm is about 90 mV and relatively independent of PV concentration when the latter is larger than 2 Times; 10^–5 M. From a model which assumes that V_m equals the equilibrium potential for the PV-cation complexes (MS ⁺) and that the reaction between PV and cations is at equilibrium on the two membrane surfaces, we compute the permeability of the membrane to free PV to be about 10^–5 cm s^–1, which is about 10^–7 times the permeability of similar membranes to free val. This interpretation is supported by the fact that the observed values of V_m are in agreement with the calculated equilibrium potential for MS⁺ over a wide range of ratios of concentrations of total PV in the two bathing solutions, if the unstirred layers are taken into account in computing the MS⁺ concentrations at the membrane surfaces.     

Effect of exogenous dolichyl monophosphate on a developmental change in mannosylphosphoryldolichol biosynthesis

The initial rate of mannosylphosphoryldolichol formation by pig brain white matter is 2.9 to 3.3-fold higher in membranes from actively myelinating animals as compared to similar preparations from adults. Exogenous dolichyl monophosphate stimulated mannolipid synthesis in both preparations indicating that the level of the acceptor lipid was rate-limiting. The relative enhancement, however, was higher in membranes from adult animals reducing the ratio of initial rates for young/adult. Exogenous dolichyl monophosphate also stimulated the labeling of a mannosylated oligosaccharide lipid and mannoproteins, including a polypeptide (apparent molecular weight of 100,000) not labeled by gray matter membranes.     

The effect of synthetic polymers on the electrical and permeability properties of lipid membranes

1. The effect of two series of hydrophilic and hydrophobic polymers on the stability, conductivity and permeability towards water and leucine of black lipid membranes and liposomes is reported. 2. The changes in properties of these membrane preparations is related to bulk phase viscosity and dielectric measurements together with monolayer studies. 3. The hydrophobic polymers dramatically increase membrane stability, had no effect on conductivity, but increased the permeability coefficient of leucine. 4. The hydrophilic polymers produced minor, but significant changes to membrane properties. 5. It is concluded that not only basic polymers but also neutral and acidic macromolecules can interact strongly with lipid membranes.     

Water permeability of thin lipid membranes

The osmotic permeability coefficient, P_f, and the tagged water permeability coefficient, P_d, were determined for thin (<100 A) lipid membranes formed from ox brain lipids plus DL-α-tocopherol; their value of approximately 1 x 10^-3 cm/sec is within the range reported for plasma membranes. It was established that P_f = P_d. Other reports that P_f > P_d can be attributed to the presence of unstirred layers in the experimental determination of P_d. Thus, there is no evidence for the existence of aqueous pores in these thin phospholipid membranes. The adsorption onto the membrane of a protein that lowers its electrical resistance by a factor of 10³ was found not to affect its water permeability; however, glucose and sucrose were found to interact with the membrane to modify P_f. Possible mechanisms of water transport across these films are discussed, together with the implications of data obtained on these structures for plasma membranes.     

Electrical properties and glucose permeability of bilayer lipid membranes on incorporation of erythrocyte membrane extracts

1. Extracts of the human erythrocyte membrane have been prepared by solubilization with Triton X-100 and analysed by electrophoresis and gel filtration techniques. 2. The extracts have been incorporated asymmetrically into lecithincholesterol-n-decane planar bilayers. 3. The electrical characteristics and glucose permeabilities of the bilayers have been measured. 4. The extracts increased the electrical conductance of the bilayers and also markedly enhanced the D-glucose permeability but not the L-glucose permeability. 5. The enhanced D-glucose permeability was inhibited by monosaccharide transport inhibitors. 6. The results support the claim that a monosachharide facititated diffusion system has been set-up in vitro which has many of the characteristics of the transport system in the human erythrocyte membrane. 7. The data indicates that the trans membrane polypeptides of band 3 of the electrophoretogram of the erythrocyte membrane proteins (notation of Fairbanks, G., Steck, T.L. and Wallach, D.F.H. (1971) Biochemistry 10, 2606-2616) are implicated in D-glucose transport, although the possibility that relatively minor component of the membrane could be responsible for glucose transport cannot be eliminated.     

Effect of general anesthetics on the properties of lipid membranes of various compositions

Computer simulations of four lipid membranes of different compositions, namely neat DPPC and PSM, and equimolar DPPC-cholesterol and PSM-cholesterol mixtures, are performed in the presence and absence of the general anesthetics diethylether and sevoflurane both at 1 and 600 bar. The results are analyzed in order to identify membrane properties that are potentially related to the molecular mechanism of anesthesia, namely that change in the same way in any membrane with any anesthetics, and change oppositely with increasing pressure. We find that the lateral lipid density satisfies both criteria: it is decreased by anesthetics and increased by pressure. This anesthetic-induced swelling is attributed to only those anesthetic molecules that are located close to the boundary of the apolar phase. This lateral expansion is found to lead to increased lateral mobility of the lipids, an effect often thought to be related to general anesthesia; to an increased fraction of the free volume around the outer preferred position of anesthetics; and to the decrease of the lateral pressure in the nearby range of the ester and amide groups, a region into which anesthetic molecules already cannot penetrate. All these changes are reverted by the increase of pressure. Another important finding of this study is that cholesterol has an opposite effect on the membrane properties than anesthetics, and, correspondingly, these changes are less marked in the presence of cholesterol. Therefore, changes in the membrane that can lead to general anesthesia are expected to occur in the membrane domains of low cholesterol content.     

The nonelectrolyte permeability of planar lipid bilayer membranes

The permeability of lecithin bilayer membranes to nonelectrolytes is in reasonable agreement with Overton's rule. The is, Pd alpha DKhc, where/Pd is the permeability coefficient of a solute through the bilayer, Khc is its hydrocarbon:water partition coefficient, and D is its diffusion coefficient in bulk hydrocarbon. The partition coefficients are by far the major determinants of the relative magnitudes of the permeability coefficients; the diffusion coefficients make only a minor contribution. We note that the recent emphasis on theoretically calculated intramembranous diffusion coefficients (Dm'S) has diverted attention from the experimentally measurable and physiologically relevant permeability coefficients (Pd'S) and has obscured the simplicity and usefulness of Overton's rule.     

Water and nonelectrolyte permeability of lipid bilayer membranes

Both the permeability coefficients (Pd's) through lipid bilayer membranes of varying composition (lecithin [L], lecithin:cholesterol [LC], and spingomyelin:cholesterol [SC]) and the n-hexadecane:water partition coefficients (Knc's) of H2O and seven nonelectrolytes (1,6 hexanediol, 1,4 butanediol, n-butyramide, isobutyramide, acetamide, formamide, and urea) were measured. For a given membrane compositiin, Pd/DKnc (where D is the diffusion constant in water) is the same for most of the molecules tested. There is no extraordinary dependence of Pd on molecular weight; thus, given Pd(acetamide), Pd(1,6 hexanediol) is correctly predicted from the Knc and D values for the two molecules. The major exceptions are H2O, whose value of Pd/DKnc is about 10-fold larger, and urea, whose value is about 5-fold smaller than the general average. In a "tight" membrane such as SC, Pd(n- butyramide)/Pd(isobutyramide)=2.5; thus this bilayer manifests the same sort of discrimination between branched and straight chain molecules as occurs in many plasma membranes. Although the absolute values of the Pd's change by more than a factor of 100 in going from the tightest membrane (SC) to the loosest (L), the relative values remain approximately constant. The general conclusion of this study is that H2O and nonelectrolytes cross lipid bilayer membranes by a solubility- diffusion mechanism, and that the bilayer interior is much more like an oil (a la Overton) than a rubber-like polymer (a la Lieb and Stein).