Keith W. Butler

The effects of alcohols on lipid bilayers: A spin label study☆

Aliphatic alcohos produce changes in the structural arrangement of lipids in bilayers as indicated by studies using the steroid spin probe 3-spiro-2′-(N-oxyl-4′,4′-dimethyloxazolidine)) cholestane. The concentrations of alcohol corresponding to the onset of perceptible changes in organization correlate well with those causing anesthesia in biological systems, suggesting that anesthesia may be caused by small disruptions of lipid bilayer structure in biological membranes. The data indicate also that lipid bilayer disruption or reorganization is involved in alcohol effects on black lipid membrane conductivity and erythrocyte antihemolysis.

The fluidity and organization of mitochondrial membrane lipids of the brown adipose tissue of cold-adapted rats and hamsters as determined by nitroxide spin probes☆

Abstract A detailed study of lipid fluidity and organization in the mitochondria of the brown adipose tissue from warm- and cold-adapted rats (nonhibernators) and hamsters (hibernators) is made in order to delineate any relationship between lipid properties and the ability to lower body temperature after cold-adaptation. Complete phospholipid analyses are presented; the data are very similar for cold- and warm-adapted rats, and for cold- and warm-adapted hamsters, but the rat lipids have a higher degree of unsaturation than those of the hamsters. Spin probe analogs of stearic acid and cholestane were used to investigate at the molecular level the fluidity and order of the mitochondrial lipids. Studies were made on intact mitochondria, and in liposomes and oriented multibilayers of extracted lipids. In no case was evidence found for a phase transition in the lipids, or for a relationship between the lipid fluidity in brown adipose tissue mitochondria and the ability to survive at lowered body temperatures. The spin probes generally had a decreased mobility in mitochondria relative to extracted lipids. The electron spin resonance spectra were analyzed to include order- and time-dependent phenomena by a recent stochastic method. The results show that more approximate analyses for order parameters and correlation times can yield incorrect conclusions. As segmental motion decreases in rate, order parameters will be overestimated. Decreasing rates of pseudoisotropic motion lead to incorrect estimates of rotational correlation times. Either of the above can result in the inference of an artifactual phase transition in the lipids.

Sterol structure and ordering effects in spin-labelled phospholipid multibilayer structures.

Abstract : Cholesterol is a common component of vertebrate cellular membranes. Sterols with a similar structure (3beta-OH group, hydrocarbon chain at position 17) are also found in vascular plants, algae, fungi, and microorganisms. The biological function of these compounds is not completely understood. The authors investigated the effects of steroid structure on the degree of order in multibilayer structures of polar membrane lipids using a spin label technique. The results indicate that cholesterol and structurally related sterols increase the degree of order of the spin label, and hence that of the lipids in the lamellar structure. (Author)

Studies on the thermotropic behavior of aqueous phosphatidylethanolamines.

The thermal response of aqueous dispersions of a series of synthetic saturated phosphatidylethanolamines was studied by differential scanning calorimetry and by infrared spectroscopy. Dispersions which had not been previously heated above tm, the temperature of the gel to liquid crystalline transition, showed transitions at a higher temperature, tm+h, having a considerably greater enthalpy change. It is demonstrated that the higher temperature transition is due to the simultaneous hydration and acyl chain melting of these saturated phosphatidylethanolamines. This transition has not been observed in the corresponding phosphatidylcholines.

The location of spin probes in two phase mixed lipid systems

Abstract We observed electron spin resonance (ESR) spectra of either a cholestane or a stearic acid spin probe in liposomes formed of egg, yeast or dipalmitoyllecithins individually or in mixtures and found: 1. (1) The “liquid lines” often observed with the stearic acid probe and liposomes originate with the probe free in solution. 2. (2) The lipid-water partition coefficient of the stearic acid probe increases with increasing fluidity of the lipid. In Mixed systems this causes the probe concentration to be higher in the more fluid component and leads to overestimation of the fluidity of the system. 3. (3) The lipid-water equilibrium of the stearic acid probe is rapid. This prevents removal of the free probe from the system by dialysis or washing and ensures that movement of the probe to the more fluid environment upon phase separation in the lipid will be rapid. 4. (4) Even with fluid lipids “liquid lines” can appear in the ESR spectra at high dilutions or high spin probe: lipid ratios. 5. (5) The cholestane spin probe produces spectra wit no “liquid lines”, which in most cases represent the fluid and crystalline phases of mixed systems in proportion to their relative concentrations.

The effects of local anesthetics on lipid multilayers: A spin probe study

Abstract The effects of a series of local anesthetics on multilayers formed from ox brain white matter lipids were investigated using an intercalated spin-labeled analog of cholestane as a monitor of molecular organization. Local anesthetics could disorder or disrupt these films at pH values approaching or above the p K of the anesthetic. At a constant concentration of a local anesthetic this effect increased with increasing pH. In films formed from lipids with a reduced cholesterol content, local anesthetics promoted the formation of ordered multilamellar arrays and increased their thermal stability. This effect required a lower concentration of local anesthetic than did the disordering effect, and each local anesthetic exhibited an optimal pH range. Depending upon the lipid, the concentration of anesthetic, and the pH of the bathing solution, local anesthetics can either stabilize or disrupt lipid bilayers.

Cation-induced organization changes in a lipid bilayer model membrane☆

Abstract Cation-induced changes in organization of the lipids in a bilayer model membrane were monitored using a spin label probe. The degree of organization of the lipids was found to be dependent on the concentration and valence of the cation as well as upon the presence of a net negative charge on the lipids. These results suggest that the influence of inorganic cations on biological membranes can take place via ion-induced organizational changes in the lipids.

The properties of gel state lipid in membranes of acholeplasma laidlawii as observed by 2H NMR

Deuterium nuclear magnetic resonance (‘H NMR) has proven to be an informative, perturbation-free monitor of the molecular organization of lipids in model [ 1,2] and biological [3,4] membranes. However, in most cases studies have been limited to the liquid crystalline squid) state of the membrane lipids due to the difficulty of observing the broad spectrum of the gel phase. With the innovation of the quadrupole echo technique [S] , and attention to various aspects of spectrometer design, it is now possible to observe these broad ‘H powder spectra [4,6]. We describe here a detailed study of the palmitoyl chains in the lipids of A. 2aidlawiz’membranes over the temperature range 145°C. We have shown that the gel and liquid crystalline states of the lipid coexist in slow exchange over the range of the calorimetricallyobserved gel-liquid crystal transition. Only -25% of the palmitoyl chains are still in the liquid crystalline state at the low temperature end of the calorimetricallydetermined transition region. At this temperature, the orientations of those chains in the gel phase still undergo considerable motion. There is a distribution of o~entation~ order parameters, the average being -50% that of completely immobilized chains. This average value is that expected for chains in the alltrans conformation rotating about their long axes. As the temperature is lowered below the transition region, the number of immobilized acyl chainsincreases steadily down to 1°C where almost all the chains are

Molecular order in acholeplasma laidlawii membranes as determined by deuterium magnetic resonance of biosynthetically-incorporated specifically-labelled lipids

The first application of deuterium magentic resonance of specifically labelled lipids to the study of a natural biological membrane is described. Palmitic acid labelled at the terminal methyl group with deuterium was incorporated biosynthetically into the lipids of the plasma membrane of Acholeplasma laidlawii. The deuterium nuclear magnetic resonance spectra contain quadrupole splittings which yield directly order parameters for this region of the membrane. Below the growth temperature (37 degrees C) the spectra are indicative of lipid in both gel and liquid crystalline states. Above this temperature they demonstrate the existence of an entirely liquid crystalline membrane whose order parameter decreases rapidly with increasing temperature. Comparison with egg phosphatidylcholine over the same temperature range shows a more rapid change in order with temperature for the A. laidlawii membranes.

Acholeplasma laidlawii membranes: An electron spin resonance study of the influence on molecular order of fatty acid composition and cholesterol

Abstract The effects on membrane structure of including various fatty acids and cholesterol in the growth medium of Acholeplasma laidlawii were investigated by the use of spin-labeled fatty acids. Although the order-mobility parameters varied significantly at some temperatures with the nature of the fatty acid incorporated, the value measured at the growth temperature was only slightly affected by changes in the fatty acid composition of the membranes. The data confirm previous assertions that despite a high level of incorporation of fatty acids of various chain lengths or degree of unsaturation, A. laidlawii regulates its overall membrane fluidity within close limits at the growth temperature. Incorporation of cholesterol increased the degree of order at all temperatures. The coexistence of two lipid phases, one protein-dependent, could be observed in membranes. The order-mobility parameter of spin probes proved less satisfactory for the observation of a gel to liquid crystal transition of the membrane lipid than the partition parameter of a fatty acid spin probe. Order parameters measured by fatty acid spin probes were somewhat higher than those measured by the analogous 2H nmr probes.