Acyl-chain remodeling of dioctanoyl-phosphatidylcholine in Saccharomyces cerevisiae mutant defective in de novo and salvage phosphatidylcholine synthesis

A yeast strain, in which endogenous phosphatidylcholine (PC) synthesis is controllable, was constructed by the replacement of the promoter of PCT1, encoding CTP:phosphocholine cytidylyltransferase, with GAL1 promoter in a double deletion mutant of PEM1 and PEM2, encoding phosphatidylethanolamine methyltransferase and phospholipid methyltransferase, respectively. This mutant did not grow in the glucose-containing medium, but the addition of dioctanoyl-phosphatidylcholine (diC8PC) supported its growth. Analyses of the metabolism of ¹³C-labeled diC8PC ((methyl-¹³C)₃-diC8PC) in this strain using electrospray ionization tandem mass spectrometry revealed that it was converted to PC species containing acyl residues of 16 or 18 carbons at both sn-1 and sn-2 positions. In addition, both acyl residues of (methyl-¹³C)₃-diC8PC were replaced with 16:1 acyl chains in the in vitro reaction using the yeast cell extract in the presence of palmitoleoyl-CoA. These results indicate that PC containing short acyl residues was remodeled to those with acyl chains of physiological length in yeast.     

Mimicry of phorbol ester responses by diacylglycerols. Differential effects on phosphatidylcholine biosynthesis, cell-cell communication and epidermal growth factor binding

The biosynthesis of phosphatidylcholine (PC) in HEL-37 cells was followed by measuring the incorporation of [32P]Pi into PC. Incorporation was stimulated by 12-O-tetradecanoylphorbol 13-acetate (TPA) and by the synthetic diacylglycerol, sn-1,2-dioctanoylglycerol (diC8), but not by sn-1-oleoyl-2-acetylglycerol or sn-1,2-dihexanoylglycerol (diC6). DiC8 was rapidly metabolised by HEL-37 cells to the corresponding PC and phosphatidic acid derivatives. diC8, diC6 and oleoylacetylglycerol effectively displaced [3H]phorbol-12,13-dibutyrate bound to a soluble cell extract from HEL-37 cells, but only diC8 was able to displace the labelled phorbol ester from prelabelled cells. TPA, diC8, diC6 and oleoylacetylglycerol were all effective inhibitors of 125I-labelled epidermal growth factor binding to, and gap junctional communication between, HEL-37 cells. It is concluded that only cell-permeable diacylglycerols stimulate PC biosynthesis which may therefore require interaction with membranes other than the plasma membrane.     

1,2-Diacylglycerols overcome cyclic AMP-mediated inhibition of phosphatidylcholine synthesis in GH3 pituitary cells.

Previous studies showed that phorbol esters and thyrotropin-releasing hormone (TRH) stimulated phosphatidylcholine synthesis via protein kinase C in GH3 pituitary cells [Kolesnick (1987) J. Biol. Chem. 262, 14525-14530]. In contrast, 1,2-diacylglycerol-stimulated phosphatidylcholine synthesis appeared independent of protein kinase C. The present studies compare phosphatidylcholine synthesis stimulated by these agents with inhibition via the cyclic AMP system. The potent phorbol ester phorbol 12-myristate 13-acetate (PMA, 10 nM) increased [32P]Pi incorporation into phosphatidylcholine at 30 min to 159 +/- 6% of control. The adenylate cyclase activator cholera toxin (CT; 10 nM) and the cyclic AMP analogue dibutyryl cyclic AMP (1 mM) abolished this effect. CT similarly abolished TRH-induced phosphatidylcholine, but not phosphatidylinositol, synthesis. This is the first report of inhibiton of receptor-mediated phosphatidylcholine synthesis by the cyclic AMP system. The 1,2-diacylglycerol 1,2-dioctanoylglycerol (diC8) also stimulated concentration-dependent phosphatidylcholine synthesis. DiC8 (3 micrograms/ml) induced an effect quantitatively similar to that of maximal concentrations of PMA and TRH, whereas a maximal diC8 concentration (30 micrograms/ml) stimulated an effect 3-4-fold greater than these other agents. CT decreased the effect of diC8 (3 micrograms/ml) by 80%. Higher diC8 concentrations overcame the CT inhibition. Similar results were obtained with dibutyryl cyclic AMP. Additional differences were found between low and high concentrations of diC8. Low concentrations of diC8 failed to induce additive phosphatidylcholine synthesis with maximal concentrations of PMA, whereas high concentrations were additive. Hence, low concentrations of 1,2-diacylglycerols appear to be regulated similarly to phorbol esters, and higher concentrations appear to act via a pathway unavailable to phorbol esters.     

Partial area of cholesterol in monounsaturated diacylphosphatidylcholine bilayers

The influence of cholesterol on the structural parameters of phosphatidylcholine bilayers is studied by small-angle neutron scattering on unilamellar liposomes. Monounsaturated diacylphosphatidylcholines diCn:1PC with the length of acyl chains n = 14, 18 and 22 carbons are used. We confirm that the bilayer thickness increases with increasing concentration of cholesterol for all studied diCn:1PCs. However, partial areas per diCn:1PC and cholesterol molecule on lipid–water interface are found not to depend of cholesterol concentration. The partial area per cholesterol molecule is 0.24 nm². In addition, the partial area per diC18:1PC is larger than that for diC14:1PC and diC22:1PC.     

Alamethicin Aggregation in Lipid Membranes

X-ray scattering features induced by aggregates of alamethicin (Alm) were obtained in oriented stacks of model membranes of DOPC(diC18:1PC) and diC22:1PC. The first feature obtained near full hydration was Bragg rod in-plane scattering near 0.11 ?⁻¹ in DOPC and near 0.08 ?⁻¹ in diC22:1PC at a 1:10 Alm:lipid ratio. This feature is interpreted as bundles consisting of n Alm monomers in a barrel-stave configuration surrounding a water pore. Fitting the scattering data to previously published molecular dynamics simulations indicates that the number of peptides per bundle is n = 6 in DOPC and n ≥ 9 in diC22:1PC. The larger bundle size in diC22:1PC is explained by hydrophobic mismatch of Alm with the thicker bilayer. A second diffuse scattering peak located at q _r ≈ 0.7 ?⁻¹ is obtained for both DOPC and diC22:1PC at several peptide concentrations. Theoretical calculations indicate that this peak cannot be caused by the Alm bundle structure. Instead, we interpret it as being due to two-dimensional hexagonally packed clusters in equilibrium with Alm bundles. As the relative humidity was reduced, interactions between Alm in neighboring bilayers produced more peaks with three-dimensional crystallographic character that do not index with the conventional hexagonal space groups.     

Incorporation and remodeling of phosphatidylethanolamine containing short acyl residues in yeast

Phosphatidylethanolamine (PE) is one of the essential phospholipids in the yeast Saccharomyces cerevisiae. We have previously shown that a yeast strain, the endogenous PE synthesis of which was controllable, grew in the presence of PE containing decanoyl residues (diC10PE) when PE synthesis was repressed. In this study, we investigated the fate of diC10PE, its uptake and remodeling in yeast. Deletion of the genes encoding Lem3p/Ros3p or P-type ATPases, Dnf1p and Dnf2p, impaired the growth of the mutants in the medium containing diC10PE, suggesting the involvement of these proteins in the uptake of diC10PE. Analysis of the metabolism of deuterium-labeled diC10PE by electrospray ionization tandem mass spectrometry revealed that it was rapidly converted to deuterium-labeled PEs containing C16 or C18 acyl residues. The probable intermediate PEs that contained decanoic acid and C16 or C18 fatty acids as acyl residues were also detected. In addition, a substantial amount of decanoic acid was released into the culture medium during growth in the presence of diC10PE. These results imply that diC10PE was remodeled to PEs with longer acyl residues and used as membrane components. Defects in the remodeling of diC10PE in the deletion mutants of ALE1 and SLC1, products of which were capable of acyl-transfer to the sn− 2 position of lyso-phospholipids, suggested their involvement in the introduction of acyl residues to the sn− 2 position of lyso-phosphatidylethanolamine in the remodeling reaction of diC10PE. Our results also suggest the presence of a mechanism to maintain the physiological length of PE acyl residues in yeast.     

Cross-linking phosphatidylinositol-specific phospholipase C traps two activating phosphatidylcholine molecules on the enzyme

Bacillus thuringiensis phosphatidylinositol-specific phospholipase C (PI-PLC), a bacterial model for the catalytic domain of mammalian PI-PLC enzymes, was cross-linked by 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride to probe for the aggregation and/or conformational changes of PI-PLC when bound to activating phosphatidylcholine (PC) interfaces. Dimers and higher order multimers (up to 31% of the total protein when cross-linked at pH 7) were observed when the enzyme was cross-linked in the presence of PC vesicles. Aggregates were also detected with PI-PLC bound to diheptanoyl-PC (diC(7)PC) micelles, although the fraction of cross-linked multimers (19% at pH 7) was lower than when the enzyme was cross-linked in the presence of vesicles. PI-PLC cross-linked in the presence of a diC(7)PC interface exhibited an enhanced specific activity for PI cleavage. The extent of this cross-linking-enhanced activation was reduced in PI-PLC mutants lacking either tryptophan in the rim (W47A and W242A) of this (betaalpha)(8)-barrel protein. The higher activity of the native protein cross-linked in the presence of diC(7)PC correlated with an increased affinity of the protein for two diC(7)PC molecules as detected by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry. In contrast to wild type protein, W47A and W242A had only a single diC(7)PC tightly associated when cross-linked in the presence of that activator molecule. These results indicate that (i) each rim tryptophan residue is involved in binding a PC molecule at interfaces, (ii) the affinity of the enzyme for an activating PC molecule is enhanced when the protein is bound to a surface, and (iii) this conformation of the enzyme with at least two PC bound that is stabilized by chemical cross-linking interacts more effectively with activating interfaces, leading to higher observed specific activities for the phosphotransferase reaction.     

Protein kinase C does not regulate diacylglycerol metabolism in aortic smooth muscle cells

Summary The effect of a reduction in protein kinase C activity on the metabolism of exogenous [³H]diC8 by freshly isolated smooth muscle cells from rabbit aorta and cultured A10 smooth muscle cells was determined. The metabolism of [³H]diC8 by both smooth muscle cell preparations was predominantly by hydrolysis to yield monoC8 and glycerol (lipase pathway); very little radioactivity was incorporated into phospholipids. Diacylglycerol lipase activity measured in vitro with A10 cell homogenates was much greater than diacylglycerol kinase activity. The addition of the protein kinase C inhibitor H-7 to incubations of isolated aortic smooth muscle cells and cultured A10 cells had no significant effect on the metabolism of [³H]diC8. Protein kinase C activity in cultured A10 cells preincubated for 20 h with a phorbol ester was reduced to 14% of control as a consequence of down-regulation, but diC8 metabolism was not changed. Therefore, protein kinase C does not regulate the metabolism of diacylglycerols in aortic smooth muscle cells.Abbreviations IP₃ inositol 1,4,5-trisphosphate - DG diacylglycerol - MG monoacylglycerol - PL phospholipid(s) - diC8 dioctanoylglycerol - H-7 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride - monoC8 monooctanoylglycerol - PS phosphatidylserine - PDBu phorbol 12,13-dibutyrate     

Alamethicin in lipid bilayers: Combined use of X-ray scattering and MD simulations

We study fully hydrated bilayers of two di-monounsaturated phospholipids diC18:1PC (DOPC) and diC22:1PC with varying amounts of alamethicin (Alm). We combine the use of X-ray diffuse scattering and molecular dynamics simulations to determine the orientation of alamethicin in model lipids. Comparison of the experimental and simulated form factors shows that Alm helices are inserted transmembrane at high humidity and high concentrations, in agreement with earlier results. The X-ray scattering data and the MD simulations agree that membrane thickness changes very little up to 1/10 Alm/DOPC. In contrast, the X-ray data indicate that the thicker diC22:1PC membrane thins with added Alm, a total decrease in thickness of 4 Å at 1/10 Alm/diC22:1PC. The different effect of Alm on the thickness changes of the two bilayers is consistent with Alm having a hydrophobic thickness close to the hydrophobic thickness of 27 Å for DOPC; Alm is then mismatched with the 7 Å thicker diC22:1PC bilayer. The X-ray data indicate that Alm decreases the bending modulus (K_C) by a factor of ∼ 2 in DOPC and a factor of ∼ 10 in diC22:1PC membranes (P/L ∼ 1/10). The van der Waals and fluctuational interactions between bilayers are also evaluated through determination of the anisotropic B compressibility modulus.     

FGF‐8b induces growth and rich vascularization in an orthotopic PC‐3 model of prostate cancer

Fibroblast growth factor 8 (FGF‐8) is expressed at an increased level in a high proportion of prostate cancers and it is associated with a poor prognosis of the disease. Our aim was to study the effects of FGF‐8b on proliferation of PC‐3 prostate cancer cells and growth of PC‐3 tumors, and to identify FGF‐8b‐associated molecular targets. Expression of ectopic FGF‐8b in PC‐3 cells caused a 1.5‐fold increase in cell proliferation in vitro and a four‐ to fivefold increase in the size of subcutaneous and orthotopic prostate tumors in nude mice. Tumors expressing FGF‐8b showed a characteristic morphology with a very rich network of capillaries. This was associated with increased spread of the cancer cells to the lungs as measured by RT‐qPCR of FGF‐8b mRNA. Microarray analyses revealed significantly altered, up‐ and downregulated, genes in PC‐3 cell cultures (169 genes) and in orthotopic PC‐3 tumors (61 genes). IPA network analysis of the upregulated genes showed the strongest association with development, cell proliferation (CRIP1, SHC1), angiogenesis (CCL2, DDAH2), bone metastasis (SPP1), cell‐to‐cell signaling and energy production, and the downregulated genes associated with differentiation (DKK‐1, VDR) and cell death (CYCS). The changes in gene expression were confirmed by RT‐qPCR. In conclusion, our results demonstrate that FGF‐8b increases the growth and angiogenesis of orthotopic prostate tumors. The associated gene expression signature suggests potential mediators for FGF‐8b actions on prostate cancer progression and metastasis. J. Cell. Biochem. 107: 769–784, 2009. © 2009 Wiley‐Liss, Inc.     

Influence of N-dodecyl-N,N-dimethylamine N-oxide on the activity of sarcoplasmic reticulum Ca(2+)-transporting ATPase reconstituted into diacylphosphatidylcholine vesicles: efects of bilayer physical parameters

Sarcoplasmic reticulum Ca-transporting ATPase (EC 3.6.1.38) was isolated from rabbit white muscle, purified and reconstituted into vesicles of synthetic diacylphosphatidylcholines with monounsaturated acyl chains using the cholate dilution method. In fluid bilayers at 37 degrees C, the specific activity of ATPase displays a maximum (31.5+/-0.8 IU/mg) for dioleoylphosphatidylcholine (diC18:1PC) and decreases progressively for both shorter and longer acyl chain lengths. Besides the hydrophobic mismatch between protein and lipid bilayer, changes in the bilayer hydration and lateral interactions detected by small angle neutron scattering (SANS) can contribute to this acyl chain length dependence. When reconstituted into dierucoylphosphatidylcholine (diC22:1PC), the zwitterionic surfactant N-dodecyl-N,N-dimethylamine N-oxide (C12NO) stimulates the ATPase activity from 14.2+/-0.6 to 32.5+/-0.8 IU/mg in the range of molar ratios C12NO:diC22:1PC=0/1.2. In dilauroylphosphatidylcholines (diC12:0PC) and diC18:1PC, the effect of C12NO is twofold-the ATPase activity is stimulated at low and inhibited at high C12NO concentrations. In diC18:1PC, it is observed an increase of activity induced by C12NO in the range of molar ratios C12NO:diC18:1PC< or =1.3 in bilayers, where the bilayer thickness estimated by SANS decreases by 0.4+/-0.1 nm. In this range, the 31P-NMR chemical shift anisotropy increases indicating an effect of C12NO on the orientation of the phosphatidylcholine dipole N(+)-P- accompanied by a variation of the local membrane dipole potential. A decrease of the ATPase activity is observed in the range of molar ratios C12NO:diC18:1PC=1.3/2.5, where mixed tubular micelles are detected by SANS in C12NO+diC18:1PC mixtures. It is concluded that besides hydrophobic thickness changes, the changes in dipole potential and curvature frustration of the bilayer could contribute as well to C12NO effects on Ca(2+)-ATPase activity.     

Phospholipid and fatty acid enrichment of Phanerochaete chrysosporium INA-12 in relation to ligninase production

Summary Ligninase activity of Phanerochaete chrysosporium INA-12 was increased when vegetable oils emulsified with sorbitan polyoxyethylene monooleate (Tween 80) were added to growth medium. Maximal enzyme yield was 22.0 nkat·ml^-1 in olive oil cultures after 4 days incubation. P. chrysosporium INA-12 was also able to utilize tall oil fatty acids for ligninase synthesis. An extracellular lipase activity was detected during the primary phase of growth in culture containing vegetable oils. On the other hand, ligninase production was 1.5-fold enhanced when olive oil cultures were supplemented with soybean asolectin as a phospholipid source. In cultures supplied with olive oil plus asolectin, P. chrysosporium INA-12 mycelium exhibited a preferential enrichment of oleic acid (C18:1), phosphatidylcholine (PC) and lysophosphatidylcholine (LPC) as compared to lipid-free medium. PC and LPC enrichment was associated with an increased ratio of saturated versus unsaturated fatty acids of phospholipids.     

<Emphasis Type="Italic">C. trachomatis</Emphasis>-infection accelerates metabolism of phosphatidylcholine derived from low density lipoprotein but does not affect phosphatidylcholine secretion from hepatocytes

Background  

Chlamydia trachomatis is a prevalent sexually transmitted disease and the leading cause of infectious blindness in developing nations. It was not known if C. trachomatis-infection influenced metabolism of lipoprotein-derived phospholipids. Nor was it known if C. trachomatis-infection altered phosphatidylcholine (PC) secretion from hepatocytes. In the current study, low density lipoprotein (LDL)-derived [methyl-³H]PC metabolism was examined in L929 cells infected with C. trachomatis to deTermine if PC derived from LDL could serve as a potential source of PC trafficked to C. trachomatis. In addition, release of endogenously synthesized [methyl-³H]PC into the medium was examined in rat liver hepatocytes infected with C. trachomatis to deTermine if infection altered PC secretion.     

Physiologic 1,2-diacylglycerol levels induce protein kinase C-independent translocation of a regulatory enzyme

Phorbol esters and 1,2-diacylglycerols have been used interchangeably to study protein kinase C action. This laboratory first suggested that 1,2-diacylglycerols may also act independent of protein kinase C using protein kinase C-"down-modulated" cells (Kolesnick, R. N., and Paley, A. E. (1987) J. Biol. Chem. 262, 9204-9210). Unfortunately, down-modulation was never complete. The present studies establish an in vitro system of enzyme translocation to resolve this issue. Choline phosphate cytidylyltransferase (EC 2.7.7.15), the regulatory enzyme for phosphatidylcholine biosynthesis, was utilized. Cytidylyltransferase translocation from cytosol to membranes mediates phorbol ester-induced phosphatidylcholine synthesis in GH3 pituitary cells. In the present studies, 1,2-diacylglycerols similarly induced phosphatidylcholine synthesis and cytidylyltransferase translocation. 1,2-Diacylglycerol-induced phosphatidylcholine synthesis, however, was not concentration-dependent but proportional to the moles of 1,2-diacylglycerol added per cell, i.e. subject to surface dilution. For instance, at constant cell number (1.67 x 10(6)/sample) and 1,2-dioctanoylglycerol concentration (diC8; 20 micrograms/ml), 32Pi incorporation into phosphatidylcholine varied from 150 to 350% above control as the incubation volume increased from 0.3 to 1.2 ml. Hence, the effective diC8 concentrations 0.5-30 micrograms/ml are preferably referred to as doses and reported as 0.25-15 nmol/10(6) cells. These doses increased cellular 1,2-diacylglycerol levels within a few fold of basal (374 pmol/10(6) cells). In vitro, diC8 also induced translocation of purified cytidylyltransferase devoid of protein kinase C to microsomes. Translocation was again subject to surface dilution. Translocation occurred with the same ratio of diC8 to microsomal membrane as phosphatidylcholine synthesis in intact cells (1-10 nmol of diC8/10(6) cell membranes). Despite stimulating cytidylyltransferase translocation in intact cells, phorbol esters failed to stimulate translocation in vitro. Hence, 1,2-diacylglycerols are not always interchangeable with phorbol esters and at physiologic levels may stimulate enzyme translocation by an alternative mechanism to protein kinase C.     

Altered intracellular calcium fluxes in pancreatic cancer induced diabetes mellitus: Relevance of the S100A8 N‐terminal peptide (NT‐S100A8)

After isolating NT‐S100A8 from pancreatic cancer (PC) tissue of diabetic patients, we verified whether this peptide alters PC cell growth and invasion and/or insulin release and [Ca²⁺]_i oscillations of insulin secreting cells and/or insulin signaling. BxPC3, Capan1, MiaPaCa2, Panc1 (PC cell lines) cell growth, and invasion were assessed in the absence or presence of 50, 200, and 500 nM NT‐S100A8. In NT‐S100A8 stimulated β‐TC6 (insulinoma cell line) culture medium, insulin and [Ca²⁺] were measured at 2, 3, 5, 10, 15, 30, and 60 min, and [Ca²⁺]_i oscillations were monitored (epifluorescence) for 3 min. Five hundred nanomolars NT‐S100A8 stimulated BxPC3 cell growth only and dose dependently reduced MiaPaCa2 and Panc1 invasion. Five hundred nanomolars NT‐S100A8 induced a rapid insulin release and enhanced β‐TC6 [Ca²⁺]_i oscillations after both one (F = 6.05, P < 0.01) and 2 min (F = 7.42, P < 0.01). In the presence of NT‐S100A8, [Ca²⁺] in β‐TC6 culture medium significantly decreased with respect to control cells (F = 6.3, P < 0.01). NT‐S100A8 did not counteract insulin induced phosphorylation of the insulin receptor, Akt and IκB‐α, but it independently activated Akt and NF‐κB signaling in PC cells. In conclusion, NT‐S100A8 exerts a mild effect on PC cell growth, while it reduces PC cell invasion, possibly by Akt and NF‐κB signaling, NT‐S100A8 enhances [Ca²⁺]_i oscillations and insulin release, probably by inducing Ca²⁺ influx from the extracellular space, but it does not interfere with insulin signaling. J. Cell. Physiol. 226: 456–468, 2011. © 2010 Wiley‐Liss, Inc.     

Water permeability and mechanical strength of polyunsaturated lipid bilayers

Micropipette aspiration was used to test mechanical strength and water permeability of giant-fluid bilayer vesicles composed of polyunsaturated phosphatidylcholine PC lipids. Eight synthetic-diacyl PCs were chosen with 18 carbon chains and degrees of unsaturation that ranged from one double bond (C18:0/1, C18:1/0) to six double bonds per PC molecule (diC18:3). Produced by increasing pipette pressurization, membrane tensions for lysis of single vesicles at 21 degrees C ranged from approximately 9 to 10 mN/m for mono- and dimono-unsaturated PCs (18:0/1, 18:1/0, and diC18:1) but dropped abruptly to approximately 5 mN/m when one or both PC chains contained two cis-double bonds (C18:0/2 and diC18:2) and even lower approximately 3 mN/m for diC18:3. Driven by osmotic filtration following transfer of individual vesicles to a hypertonic environment, the apparent coefficient for water permeability at 21 degrees C varied modestly in a range from approximately 30 to 40 microm/s for mono- and dimono-unsaturated PCs. However, with two or more cis-double bonds in a chain, the apparent permeability rose to approximately 50 microm/s for C18:0/2, then strikingly to approximately 90 microm/s for diC18:2 and approximately 150 microm/s for diC18:3. The measurements of water permeability were found to scale exponentially with the reduced temperatures reported for these lipids in the literature. The correlation supports the concept that increase in free volume acquired in thermal expansion above the main gel-liquid crystal transition of a bilayer is a major factor in water transport. Taken together, the prominent changes in lysis tension and water permeability indicate that major changes occur in chain packing and cohesive interactions when two or more cis-double bonds alternate with saturated bonds along a chain.     

The presence of odd-chain fatty acids in Drosophila phospholipids

ABSTRACT

Most fatty acids in phospholipids and other lipid species carry an even number of carbon atoms. Also odd-chain fatty acids (OCFAs), such as C15:0 and C17:0, are widespread throughout the living organism. However, the qualitative and quantitative profiles of OCFAs-containing lipids in living organisms remain unclear. Here, we show that OCFAs are present in Drosophila phosphatidylcholine (PC) and phosphatidylethanolamine (PE) and that their level increases in accordance with progression of growth. Furthermore, we found that food-derived propionic acid/propanoic acid (C3:0) is utilized for production of OCFA-containing PC and PE. This study provides the basis for understanding in vivo function of OCFA-containing phospholipids in development and lipid homeostasis.     

Biopolymer excreted by Pseudoalteromonas antarctica NF3, as a coating and protective agent of liposomes against dodecyl maltoside

The ability of an exopolymer of glycoproteic character (GP) excreted by a new gram‐negative species Pseudoalteromonas antarctica NF₃, to coat phosphatidylcholine (PC) liposomes and to protect these bilayers against the action of the nonionic surfactant dodecyl maltoside was investigated. Transmission electron microscopy (TEM) micrographs of freeze fractured liposome/GP aggregates reveal that the addition of the glycoprotein to liposomes led to the formation of a film (polymer adsorbed onto the bilayers) that tightly coated PC bilayers. The complete coating was already achieved at a PC : GP weight ratio of about 9:1. Image analysis profiles of digitalized TEM micrographs (PC : GP weight ratio 8:2) show that this film was formed by a multilayer structure. The periods of the average distance of the pattern ordering in layer structures (9–10 layers) were of about 2–3 nm and the thickness of the complete film was of about 25 nm. Higher amounts of glycoprotein resulted in a growth of this film, which exhibited at the highest proportion of this compound (50% in weight) a multifilm structure. An increasing resistance of liposomes to be affected by dodecyl maltoside both at subsolubilizing and solubilizing levels occurred as the proportion of the glycoprotein in the system rose, although this protective effect was more effective at low proportions of this compound (PC : GP weight ratios from 9:1 to 8:2). Thus, although a direct dependence was found between the growth of the enveloping structure and the resistance of the coated liposomes to be affected by the surfactant, the more effective protection occurred when this structure was a thin film formed by the assembly of various layers of GP of about 2–3 nm. © 1999 John Wiley & Sons, Inc. Biopoly 50: 579–588, 1999