Showing papers on "Phosphatidylethanolamine published in 1979"

Influence of thermal acclimation on membrane lipid composition of rainbow trout liver

Abstract: Rainbow trout (Salmo gairdneri) acclimated to 5 degrees C possessed larger livers and less neutral lipid per gram of liver than 20 degrees C-acclimated animals; quantities of liver glycolipid, phospholipid, and cholesterol did not vary significantly with acclimation temperature. The relative proportions of phosphatidylethanolamine increased significantly following cold exposure, whereas the quantities of sphingomyelin and cardiolipin declined. For all phosphatides examined (phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, phosphatidylinositol, lysolecithin, cardiolipin, sphingomyelin) cold acclimation resulted in 1) an increase in the quantity of polyunsaturated fatty acids, 2) a reduction in the level of saturated fatty acids, and 3) little change in the total content of monoenes and dienes. The increased content of polyunsaturated fatty acids in choline and ethanolamine phosphatides following cold acclimation was confined to the 2-position and occurred at the expense of monoenes and dienes. The relative proportions of n - 3 fatty acids, and less frequently n - 6 fatty acids, increased in phosphatides of cold-acclimated trout, whereas the relative proportions of n - 9 fatty acids declined. These data suggest a preferential incorporation of fatty acids belonging to the linolenic acid family at reduced temperatures. Temperature-induced changes in the chemical composition of trout liver phospholipids counteracted the effects of acute temperature change on nonelectrolyte permeability of isolated liposomes.

Phospholipid methylation unmasks cryptic beta-adrenergic receptors in rat reticulocytes

Abstract: The effect of phospholipid methylation on the number of beta-adrenergic receptor binding sites was examined in rat reticulocyte membranes. Stimulation of phosphatidylcholine synthesis by the introduction of the methyl donor S-adenosyl-L-methionine into reticulocyte ghosts increased the number of beta-adrenergic receptor sites. The appearance of beta-adrenergic binding sites was dependent on the formation of phosphatidylcholine by the enzyme that converts phosphatidyl-N-monomethylethanolamine from phosphatidylethanolamine. Both the synthesis of phosphatidylcholine and the unmasking of cryptic receptors were time and temperature dependent and did not occur in the presence of the methyl transferase inhibitor, S-adenosyl-L-homocysteine.

Rhodopsin-lipid associations in bovine rod outer segment membranes. Identification of immobilized lipid by spin-labels.

Abstract: Rhodopsin-lipid interactions have been studied in bovine rod outer segment (ROS) membranes by using spin-labels. Spin-labeled fatty acid, sterol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol, and phosphatidic acid molecules all display a two-component spectrum when probing ROS membranes. One of the spectral components represents 33-43% of the total spectral intensity and is characteristic of a strongly immobilized nitroxide spin-label. This immobilized component is resolved from -4 to 37 O C . The remaining 67-57% of the integrated spectral intensity has a very similar form to the spectra of the same spin-labels in bilayers of extracted ROS membrane lipid. A small selectivity for the immobilized regions of ROS M a n y important biological functions are performed by proteins organized in lipid bilayer membranes (Sandemann, 1978), and an understanding of the interactions between these proteins and their lipids is therefore of particular importance. Rod outer segment (ROS’) membranes are well suited to the study of the interactions between integral membrane proteins and the lipid matrix for a variety of reasons. Firstly, rhodopsin constitutes 85-90% of the total membrane protein of bovine ROS membranes (Montal & Korenbrot, 1976; O’Brien, 1978; Daemen, 1973; Papermaster et al., 1976), and therefore any protein-lipid interactions will, in all probability, be directly due to rhodopsin-lipid associations. Such a level of single protein enrichment is normally achieved by reconstitution, recombination, or specific enrichment procedures which suffer from the hazards of either protein aggregation or denaturation or both and of detergent removal, which is often a long and incomplete process. ROS membranes are isolated without the use of detergents or of enrichment processes involving delipidation or protein loss. Secondly, a limited amount of structural information for rhodopsin is available. This has come from X-ray diffraction studies (Charbre, 1975), X-ray scattering (Sardet et al., 1976) and neutron-scattering (Osborne et al., 1978) data, and ultracentrifugation experiments (Lewis et al., 1974), and some dimensional information has been determined (Sardet et al., 1976; Osborne et al., 1978). Such parameters are useful when interpreting observations of the interactions between lipids and integral membrane proteins in structural terms. Thirdly, ROS membranes are particularly interesting since rhodopsin, within its membrane environment, is responsible for the primary step in visual perception, leading from light absorption to nerve excitation. This process involves both the full photolytic cycle of rhodopsin, including regeneration, and the consequent modulation of the cytoplasmic activity of an internal transmitter through conformational changes taking From the Max-Planck-Institut fur biophysikalische Chemie, Abteilung Spektroskopie, D-3400 Gottingen-Nikolausberg, Federal Republic of Germany. Received May 29, 1979. I.D.V. was the recipient of a DFG exchange stipend to the Abteilung Biochemische Kinetik. $Permanent address: Institute of Photobiology, Academy of Science of B.S.S.R., Minsk, U.S.S.R. membranes is shown by phosphatidylser‘ne, while the fatty acid, phosphatidylcholine, phosphatidy / ethanolamine, and sterol spin-labels partition almost equally into these regions. The selectivity is not solely due to the head-group charge on phosphatidylserine since the negatively charged phosphatidylglycerol and phosphatidic acid spin-labels do not display an enhanced selectivity. These results are interpreted in terms of two populations of lipid existing in ROS membranes, the major one being fluid bilayer in exchange with and surrounding the immobilized lipid which is in direct contact with rhodopsin. On the basis of available information on the size of rhodopsin, it is calculated that the immobilized lipid is sufficient to form a single complete shell around the protein. place in rhodopsin (Saibil et al., 1976; Liebman et al., 1974; Downer & Englander, 1975; McDowell & Williams, 1976; Ostroy, 1977; Hubbell et al., 1977). The membrane lipid could be involved in both of these stages: in stabilizing the structure of rhodopsin during the various states of the photolytic and regenerative cycles and in regulating the conformational changes leading to the transmitter response. In the present work, we demonstrate the existence of an immobilized lipid component in ROS membranes as monitored by a number of lipid spin-labels. This immobilized lipid is revealed as one component of a distinct two-component ESR spectrum of spin-labeled ROS membranes, the observation of which in itself demonstrates that the two components exist for longer than s. Similar experiments with bilayers of extracted ROS lipid show only a one-component spectrum, typical of the spin-labels undergoing anisotropic motion in a lipid bilayer. Such fluid bilayer spectra closely resemble the other components observed from spin-labeled ROS membranes. It is suggested that the “immobilized” component arises from motionally perturbed bilayer lipid. From spectral subtraction and integration it is shown that the amount of immobilized lipid is 24 lipid molecules/protein, which is probably sufficient to form a single shell around rhodopsin, the major protein of ROS membranes. Experiments with fatty acid, sterol, and a number of phospholipid spin-labels demonstrate a limited preference for the immobilized region by phosphatidylserine molecules. The organization in the ROS membrane, the composition of the immobilized lipids around rhodopsin, and the approximate size of the intramembranous portion of rhodopsin are discussed. Experimental Section

Activation of phospholipid metabolism during mediator release from stimulated rat mast cells.

Abstract: Phospholipid metabolism was studied during mediator release from highly purified rat mast cells. The incorporation of 32 PO 4 into individual phospholipids was determined after isolation of phospholipid classes by two-dimensional thin layer chromatography. Various stimulators of histamine release (anti-IgE antibody, concanavalin A [Con A], compound 48/80, or the ionophore A23187) increased 32 PO 4 incorporation into phosphatidic acid (PA), phosphatidylinositol (PI), and phosphatidylcholine (PC) 4- to 10-fold within 15 min. No change in 32 PO 4 incorporation into phosphatidylserine (PS), phosphatidylethanolamine, or sphingomyelin was detected. Anti-IgE antibody caused significantly increased labeling of PA within 8 sec and of PI and PC within 30 sec. The concentrations of anti-IgE and Con A causing threshold, half maximal, and maximal phospholipid labeling were slightly less than those required for comparable changes in histamine release. Both phospholipid labeling and mediator release in response to anti-IgE or Con A were enhanced 3- to 4-fold by PS. The addition of 50 mM α-methylmannoside to mast cells 10 min after challenge with Con A rapidly halted both ongoing mediator release and PA labeling. The results of these studies indicate that marked and selective changes in phospholipid metabolism occur before as well as during mediator release from mast cells and that these reactions may be an intrinsic part of the biochemical mechanisms that control mediator release.

Benzodiazepine and β -adrenergic receptor ligands independently stimulate phospholipid methylation

Abstract: The stepwise methylation of phosphatidylethanolamine by two methyltransferase synthesises phosphatidylcholine in membranes1. Although the CDP-choline pathway is the major route for phosphatidylcholine synthesis, recent work in our laboratory has found that the methylation pathway is metabolically highly active. The phospholipid methyltransferases, their substrates and their products are asymmetrically distributed in the cell membrane. Methyltransferase I, located on the cytoplasmic side of the membrane, transfers a methyl group from S-adenosylmethionine to phosphatidylethanolamine to form phosphatidyl-N-monomethylethanolamine. Methyltransferase II, on the exterior surface of the bilayer, catalyses two additional N-methylations to form phosphatidylcholine. The asymmetric distribution of methyltransferases results in the synthesis and translocation of phosphatidylcholine to the exterior bilayer surface2. The synthesis and translocation of methylated phos-pholipid affects many membrane events, such as fluidity3, coupling of the β-adrenergic receptor with adenylate cyclase4, the number of β-adrenergic receptors5, Ca2+-ATPase activity6, leukocyte chemotaxis7, mast cell histamine secretion8 and lymphocyte mitogenesis9. We now report that stimulation of a benzodiazepine receptor with anti-anxiety benzodiazepines, and stimulation of the β-adrenergic receptor with β-agonists, increase phospholipid methylation in C6 astrocytoma cells. Furthermore, simultaneous addition of benzodiazepine and β-adrenergic agonists increases methylation in an additive manner.

Interactions of lysophospholipids and mast cells

Abstract: PHOSPHOLIPID and protein are the major components of biological membranes, and much is known about their organisation in various cell membranes1,2. An important question concerns the existence and significance of specific protein–phospholipid interactions in membranes. Among the methods used to elucidate these interactions is the study of the regulatory function of specific phospholipids in intact cells. Phosphatidylserine (PS) enhances the secretory response of the mast cell3 in a highly specific manner. Other diacyl phospholipids, phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol, phosphatidylinositol and phosphatidic acid, are ineffective3–5 and several N-substituted derivatives of PS are competitive inhibitors of the effect of PS on mast cells6. Furthermore, PS selectively enhances the response of cells to IgE-dependent secretagogues7,8 without increasing the response of the cells to IgE-independent agents such as polymyxin B9, chymotrypsin3, compound 48/80 (refs 3, 9), or A23187 (D. L. and T. W. M., unpublished). As histamine secretion involves a major alteration in the organisation of membrane components10, the effect of PS on secretion makes it possible to study a complex membrane event with an established phospholipid specificity. Previously, we have used PS of various fatty acid compositions11: dimyristoyl PS, dipalmitoyl PS, and distearoyl PS. We now provide evidence that lysophosphatidyl-serine (lyso-PS) generated from bovine brain PS by the action of phospholipase A2 activates mast cell secretion induced by concanavalin A (Con A) at 1/30th to 1/50th the concentration of PS required for an equivalent response. Although some characteristics of the activation of mast cell secretion by lyso-PS resemble those observed with PS, its mode of interaction with the cell may be different; PS acts on mast cells in a micellar state12, but lyso-PS acts to potentiate secretion well below its critical micelle concentration (CMC).

Lipid alterations in cell envelopes of polymyxin-resistant Pseudomonas aeruginosa isolates.

Abstract: The lipid composition of cells of Pseudomonas aeruginosa strains resistant to polymyxin was compared with the lipid composition of cells of polymyxin-sensitive strains as to their content of readily extractable lipids (RELs), acid-extractable lipids, the fatty acid composition of RELs, and the contents of various phospholipids in the REL fraction. The polymyxin-resistant strains had an increased content of RELs, but a decreased phospholipid content. The REL fraction from the polymyxin-resistant strains had an increased content of unsaturated fatty acids accompanied by a decreased content of cyclopropane fatty acids as compared with the fatty acid composition of RELs from polymyxin-sensitive strains. The phosphatidylethanolamine content was greatly reduced in the polymyxin-resistant strains, whereas the content of an unidentified lipid, thought to be a neutral lipid lacking either a phosphate, free amino, or choline moiety, was greatly increased. Cell envelopes of the polymyxin-resistant strains contained reduced concentrations of Mg2+ and Ca2+ as compared with the cell envelopes of polymyxin-sensitive strains. It appears that polymyxin resistance in these strains is associated with a significant alteration in the lipid composition and divalent cation content of the cell envelope.

A Role of Membranes in the Activation of a New Multifunctional Protein Kinase System

Abstract: A new multifunctional protein kinase, which normally exists as an inactive form in the soluble fraction in mammalian tissues, attaches to membranes to exhibit full enzymatic activity. A low concentration of Ca2+ is absolutely necessary for this activation. This process is reversible. cAMP shows no effect. The active factors in membranes are phosphatidylinositol, phosphatidylserine, phosphatidic acid, diphosphatidylglycerol, and phosphatidylethanolamine in that order. Phosphatidylcholine and sphingomyelin are far less effective. Cytoplasmic as well as other membrane fractions from various tissues are active in supporting the enzymatic activity. A possible role of this Ca2+ and phospholipid-activated protein kinase system in transmembrane control is proposed.

Effect of phosphatidylserine on acetylcholine output from the cerebral cortex of the rat.

Abstract: — The effect of sonicated suspensions of phosphatidylserine, phosphatidylethanolamine and phosphatidylcholine injected intravenously on acetylcholine release from the cerebral cortex was investigated in urethane anaesthetized rats. The electroeorticogram was also recorded. Phosphatidylserine caused a dose dependent, calcium dependent increase in acetylcholine output with no electrocorticografic changes. The increase, 75% peak effect after 150 mg/kg, was abolished by septal lesions and pretreatment with pimozide. Phosphatidylserine had no effect on acetylcholine release from brain slices in vitro. Phosphatidylethanolamine was approximately half as active as phosphatidylserine and phosphatidylcholine had no effect on acetylcholine output in vivo. It is concluded that phosphatidylserine exerts an indirect stimulating action on a septio-cortical cholinergic pathway.

The calcium-dependent phosphatidylinositol-phosphodiesterase of rat brain. Mechanisms of suppression and stimulation.

Abstract: 1. The activity of the soluble, calcium-dependent phosphatidylinositol-specific phosphodiesterase (EC 3.1.4.10) against [32P] phosphatidylinositol has been investigated. 2. KCl (only at neutral pH), Mg2+, positively-charged proteins such as histone, and phospholipids containing a choline headgroup are all inhibitory to the enzyme. Choline-phospholipids cause a 90% inhibition at an equimolar ratio to phosphatidylinositol. 3. Other phospholipids (phosphatidylglycerol, phosphatidylserine, phosphatidylethanolamine and phosphatidic acid) are all potent stimulators of the enzyme: maximum stimulation being observed at a ratio of 1 mol activator/5–10 mol phosphatidylinositol. 4. Unsaturated amphiphiles such as oleic and oleoyl alcohol also stimulate the activity, maximum stimulation being observed at about an equimolar ratio to phosphatidylinositol. Saturated amphiphiles (such as stearic acid and stearoyl alcohol) are less effective. 5. The activation by acidic phospholipids and unsaturated amphiphiles appear to be independent as they are additive and, under certain conditions, synergistic. 6. Both types of stimulator (independently or together) can reverse the inhibition caused by histone or phosphatidylcholine. 7. Possible mechanisms of the suppression of the phosphatidylinositol phosphodiesterase in vivo, of its activation, and of the amplification of phosphatidylinositol breakdown are discussed.

The Relipidation of Delipidated Na,K‐ATPase

Abstract: Enzymatically inactive, delipidated Na,K-ATPase from dogfish rectal glands was titrated with dioleoylphosphatidylcholine and with dioleoylphosphatidylethanolamine The process of relipidation has the following characteristic properties Enzymatic activities reappear independently of each other: first the phosphatase, then the ATPase The properties of the phosphatase regenerated depend on the ratio of lipid/protein used; the ATPase seems to be independent of this ratio The simplest model that is consistent with the above results and with the shapes of the titration curves, has the following requirements Firstly, the enzyme is composed of two subunits that, as far as lipid binding is concerned, are identical and independent of each other Secondly, lipid adds onto the enzyme as preformed clumps of 25 molecules of phosphatidylcholine or 18 molecules of phosphatidylethanolamine Thirdly, each subunit binds two clumps of lipid, and binding shows positive cooperativity Fourthly, when either subunit becomes saturated with lipid, the enzyme exhibits one form of phosphatase Fifthly, when both subunits are saturated with lipid, the enzyme exhibits a second form of phosphatase and ATPase The data and their analysis according to this model lead to the suggestion that Na,K-ATPase is a functional dimer, the interaction between subunits being influenced by the Na+ and K+ concentrations in the medium: K+ favouring the functional independence of the subunits and Na+favouring their functional interaction