31P-NMR studies on phospholipid structure in membranes of intact, functionally-active, rat liver mitochondria.
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Animals↗
Biomedical subjects
B De Kruijff
Publications and source records attributed to B De Kruijff.
Effects of lysophosphatidylcholines on phosphatidylcholine and phosphatidylcholine/cholesterol liposome systems as revealed by 31P-NMR, electron microscopy and permeability studies.
(1) The effect of incorporation of different lysophosphatidylcholine species on the structure, barrier properties and dynamics of bilayers made of various phosphatidylcholines both the presence and absence of cholesterol have been investigated by 31P-NMR, freeze-fracture electron microscopy and K+-permeability measurements. (2) In a dispersion of lysophosphatidylcholine : cholesterol (1 : 1) the lipids are organized in extended bilayers. Upon cooling a micellar solution of 1-palmitoyllysophosphatidylcholine below the chain-melting temperature a transition to a lamellar, most likely interdigitating organization is observed. 31P-NMR shows in both situations a marked decrease in effective chemical shift anisotropy. (3) 1-Palmitoyllysophosphatidylcholine can be incorporated up to 30 mol% into liquid crystalline bilayers of dipalmitoylphosphatidylcholine and up to 35 mol% into dioleoylphosphatidylcholine bilayers. Above this concentration micellization of the bilayers occurs. In the gel state, bilayer structure is maintained up to 60 mol% of the lysocompound. (4) 1-Oleoyllysophosphatidylcholine can be incorporated to higher concentrations into liquid crystalline phosphatidylcholine bilayers than the palmitoyl analogue, which can be explained by the more cylindrical shape of the 1-oleoyllysophosphatidylcholine. (5) In marked contrast, incorporation of only 1 mol% of 1-oleoyllysophosphatidylcholine into gel state dipalmitoylphosphatidylcholine already destabilizes bilayer structure and makes the membranes completely permeable for K+. These results are discussed with respect to the mixing properties of the various lysophosphatidylcholines. (6) In general these effects are accompanied by a loss of K+-permeability barrier, which however occurs at lower lysophosphatidylcholine concentrations than needed for the start of micellization. (7) Cholesterol incorporation counteracts the bilayer destabilizing role of lysophosphatidylcholines. (8) 31P-NMR demonstrates with increasing lysophosphatidylcholine concentrations in the bilayers of phosphatidylcholines a decrease in the effective chemical shift anisotropy. As the rigid lattice spectra of lysophosphatidylcholine and phosphatidylcholine are identical, this reflects a change in the conformational and/or motional properties of the phospholipid head groups. This phenomenon might play a role in the observed permeability changes.
Cholesterol↗
A possible role of rhodopsin in maintaining bilayer structure in the photoreceptor membrane.
31P-NMR measurements demonstrate that at 37 degrees C, independent of the photolytic state of the photopigment rhodopsin, the lipids in the photo-receptormembrane are almost exclusively organised in a bilayer. In strong contrast, the 31P-NMR spectra of the extracted lipids are characteristic for the hexagonal HII phase and an isotropic phase. The isotropic phase is characterised by freeze-fracture electron microscopy as particles and pits on smooth surfaces, possibly indicating inverted micelles. These results suggest a structural role for rhodopsin in maintaining the photoreceptor membrane lipids in a bilayer configuration.
Animals↗
Effect of the phase transition on the transbilayer movement of dimyristoyl phosphatidylcholine in unilamellar vesicles.
Dimyristoyl phosphatidylcholine rapidly exchanges between vesicles at 37 degrees C without vesicle fusion. The rate of the transbilayer movement of dimyristoyl phosphatidylcholine in sonicated vesicles has been measured employing 13C NMR using N-13CH3-labeled lipids which are introduced into the outer monolayer of non-labeled vesicles by a phosphatidylcholine exchange protein. The rate of transbilayer movement of dimyristoyl phosphatidylcholine shows a distinct maximum (half-time 4 h) in the temperature range at which the hydrocarbon phase transition occurs. The activation energy of the flip-flop rate above the phase transition is 23.7 +/- 2.0 kcal/mol.
Biological Transport↗
Polymorphic phase behaviour of lipid mixtures as detected by 31P NMR. Evidence that cholesterol may destabilize bilayer structure in membrane systems containing phosphatidylethanolamine.
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Cholesterol↗
Rapid transbilayer movement of phospholipids induced by an asymmetrical perturbation of the bilayer.
Phospholipase D is used to convert egg phosphatidylcholine to phosphatidic acid in unilamellar vesicles. The transbilayer distribution of both lipids is determined by 31P NMR using paramagnetic ions. Phosphatidic acid formed in the outer monolayer is translocated to the inner monolayer with a halftime of 30-40 min or less. This is accompanied by an equally fast movement of part of the phosphatidylcholine from the inner to the outer monolayer. During these fast transbilayer movements the barrier properties of the vesicle bilayer are maintained.
Magnetic Resonance Spectroscopy↗
Induction of a relatively fast transbilayer movement of phosphatidylcholine in vesicles. A 13CNMR study.
[N-13CH3]Phosphatidylcholines are introduced into the outer monolayer of phosphatidylcholine vesicles with the phosphatidylcholine exchange protein from bovine liver. The transbilayer distribution of the [N-13CH3]phosphatidyl-choline is measured with 13C NMR. The transbilayer movements of [N-13CH3]-dioleoyl phosphatidylcholine and [N-13CH3]dimyristoyl phosphatidylcholine at 30 degrees C in vesicles composed of these phosphatidylcholines are extremely slow processes with estimated half-times of days. [N-13CH3]Dioleoyl phosphatidyl-choline introduced into dimyristoyl phosphatidylcholine vesicles migrates from the outer to the inner monolayer with a half-time of less than 12 h. The data suggest that differential changes in the lateral packing of the two monolayers might be a driving force for transbilayer transport of phospholipids.
Animals↗
The preference of cholesterol for phosphatidylcholine in mixed phosphatidylcholine-phosphatidylethanolamine bilayers.
The following phosphatidylethanolamines were studied by differential scanning calorimetry: 1,2-dipalmitoleoyl-, 1,2-dioleoyl-, 1,2-dilauroyl-, 1,2-dielaidyl-, 1,2-dimyristoyl- and 1,2-dipalmitoyl-sn-glycero-3-phosphoryl-ethanolamine. The saturated and trans-unsaturated species underwent thermotropic phase transitions at temperatures about 20-30 degrees C higher than the corresponding phosphatidylcholines but the enthalpy changes were nearly identical. The transition temperatures for the cis-unsaturated species were about the same as those of the corresponding phosphatidylcholines but here the enthalpy change was markedly decreased as compared with the phosphatidylcholines. Freeze-fracture electron microscopy revealed phase changes from a lamellar to a hexagonal phase for 1,2-dipalmitoleoyl- and 1,2-dioleoyl-sn-glycero-phosphorylethanolamine at 20 and 0 degrees C respectively. At these temperatures no transitions were apparent in the calorimeter scan. Incorporation of increasing amounts of cholesterol into phosphatidylethanol-amine bilayers gradually decreased the enthalpy changes of the phase transition in the same manner as was demonstrated before for phosphatidylcholine/cholesterol mixtures. This was studied both for 1,2-dipalmitoleoyl- and 1,2-dimyristoyl-sn-glycerophosphorylethanolamine. In an equimolar mixture of 1,2-dioleoyl- and 1,2-dipalmitoylphosphoryl-ethanolamine, which showed phase separation, cholesterol preferentially decreased the transition of the lowest melting component. In equimolar mixtures of phosphatidylethanolamines and phosphatidylcholines, which showed phase separation, cholesterol preferentially abolished the transition of the phosphatidylcholine component present. This occurred both in experiments where the phosphatidylcholine was the lowest melting and where it was the highest melting component present in the mixture. These experiments strongly suggest that in phosphatidylcholine-phosphatidylethanolamine mixtures at temperatures where both components are in the liquid-crystalline state cholesterol is preferently associated with the phosphatidylcholine component in the mixture.
Cholesterol↗
Outside-inside distributions and sizes of mixed phosphatidylcholine-cholesterol vesicles.
(1) The effect of the incorporation of cholesterol upon the distribution of various molecular species of phosphatidylcholine across the bilayers of mixed sonicated liposomes (vesicles) has been studied with 31P-MNR. (2) The outside-inside ratio of both saturated and unsaturated phosphatidylcholine species was not much affected by the incorporation of up to 30 mol% cholesterol. Above 30 mol% cholesterol the outside-inside ratio strongly increased for phosphatidylcholines with cis unsaturated fatty acid chains. In contrast the outside-inside ratio of trans unsaturated and fully saturated phophatidylcholine species was either not affected or decreased by the incorporation of more than 30 mol% cholesterol. (3) a simple relationship between the size of the vesicle and the linewidth of the 31P-NMR resonance is described. From the measured linewidths the sizes of the various cholesterol containing vesicles have been obtained. It is found that incorporation of 0-30 mol% cholesterol does not significantly affect the size of the vesicle whereas above 30 mol% cholesterol does not significantly affect the size of the vesicle whereas above 30 mol% cholesterol the size of all phosphatidylcholine vesicles sharply increases. The increase in size is the largest for the more saturated phosphatidylcholine species. (4) From the outside-inside ratio and the size of the vesicle the composition of the outer and inner layer of the mixed vesicles could be obtained. Below 30 mol% cholesterol the composition of outer and inner layer is nearly identical. Above 30 mol% cholesterol the distribution of lipid across the bilayer of all visicles becomes assymetric with a disporportionately larger amount of cholesterol present in the inside monolayer.
Binding Sites↗
Phosphorus nuclear magnetic resonance of Acholeplasma laidlawii cell membranes and derived liposomes.
1. The 129 MHz 31P-NMR spectrum of Acholeplasma laidlawii membranes is very similar to the spectrum of the derived liposomes and is a typical "solid state" spectrum in which the major contribution to the linewidth is made by the chemical shift anisotropy. From the value of the chemical shift anisotropy an order parameter of 0.15 is estimated for the lipid phosphates in both membranes. 2. The 31P-NMR spectrum of the A. laidlawii membrane is insensitive to pronase digestion of 4-60% of the membrane proteins and subsequent cytochrome C binding. These results indicate that either no strong lipid polar headgroup-protein interactions occur in the membrane or that the lipid-protein "complexes" in the membrane have a fast rotation (Tc shorter than 10(-6)S) along an axis perpendicular to the plane of the membrane. 3. Phospholipase A2 degrades all the phosphatidylglycerol in the membrane. The resulting membrane contains a phosphoglycolipid as the sole phosphorus-containing compound. The 31P-NMR spectrum of these membranes is identical to the spectrum of the native membranes suggesting a similar motion for the phosphate groups in both lipids. 4. Ca2+ binding to liposomes prepared from either the total polar lipids or the total phosphorus-containing lipids isolated from the A. laidlawii membrane does not affect the 21P-NMR spectrum. 5. The 31P-NMR spectrum of the membranes and derived liposomes, however, is sensitive to lipid phase transitions. When the membrane lipids are in the gel state a broadening of the 31P resonance occurs demonstrating that the polar head group motion in a biological membrane is more restricted below the lipid-phase transition temperature.
Acholeplasma laidlawii↗