Biomedical subjects

G G Shipley

Publications and source records attributed to G G Shipley.

At least 19 recordsLinked to original sources

PubMed1992

Circular dichroic studies of protein kinase C and its interactions with calcium and lipid vesicles.

Circular dichroism was used to study the secondary structure of protein kinase C (PKC) in aqueous solution and the conformational changes resulting due to the presence of its regulatory cofactors (e.g. Ca2+, phosphatidylserine (PS) and phorbol 12-myristate 13-acetate (PMA)). Computer analysis of the CD data for the estimates of secondary structure showed that PKC maintains a highly ordered structure containing 36% alpha-helix, 57% beta-sheet and 7% beta-turn. PKC displays a minor conformational change upon addition of Ca2+. However, a larger change is observed on adding phosphatidylserine vesicles in the presence of Ca2+. In this case, the alpha-helix content is decreased by approx. 35% and beta-sheet increased by approx. 16%. The protein does not experience further significant changes in conformation on adding PMA.

Adenosine Triphosphate↗

PubMed1991

Structure and interactions of ether- and ester-linked phosphatidylethanolamines.

The ether-linked phospholipid 1,2-dihexadecylphosphatidylethanolamine (DHPE) was studied as a function of hydration and in fully hydrated mixed phospholipid systems with its ester-linked analogue 1,2-dipalmitoylphosphatidylethanolamine (DPPE). A combination of differential scanning calorimetry (DSC) and X-ray diffraction was used to examine the phase behavior of these lipids. By DSC, from 0 to 10 wt % H2O, DHPE displayed a single reversible transition that decreased from 95.2 to 78.8 degrees C and which was shown by X-ray diffraction data to be a direct bilayer gel to inverted hexagonal conversion, L beta----HII. Above 15% H2O, two reversible transitions were observed which stabilized at 67.1 and 92.3 degrees C above 19% H2O. X-ray diffraction data of fully hydrated DHPE confirmed the lower temperature transition to be a bilayer gel to bilayer liquid-crystalline (L beta----L alpha) phase transition and the higher temperature transition to be a bilayer liquid-crystalline to inverted hexagonal (L alpha----HII) phase transition. The lamellar repeat distance of gel-state DHPE increased as a function of hydration to a limiting value of 62.5 A at 19% H2O (8.6 mol of water/mol of DHPE), which corresponds to the hydration at which the transition temperatures are seen to stabilize by DSC. Electron density profiles of DHPE, in addition to calculations of the lipid layer thickness, confirmed that DHPE in the gel state forms a noninterdigitated bilayer at all hydrations. Fully hydrated mixed phospholipid systems of DHPE and DPPE exhibited two reversible transitions by DSC.(ABSTRACT TRUNCATED AT 250 WORDS)

Drug Interactions↗

PubMed1991

Structure and thermotropic properties of hydrated N-stearoyl sphingomyelin bilayer membranes.

Hydrated multibilayers of N-stearoyl sphingomyelin were investigated as a function of hydration using differential scanning calorimetry (DSC) and X-ray diffraction. Anhydrous N-stearoyl sphingomyelin exhibits an endothermic transition at 75 degrees C (delta H = 3.8 kcal/mol); increasing hydration progressively lowers the transition temperature and increases the transition enthalpy, until limiting values (Tm = 45 degrees C, delta H = 6.7 kcal/mol) are observed for hydration values greater than 21.4% H2O. At low hydration levels, less than 20% H2O, an additional transition is observed at approx. 20 degrees C. X-ray diffraction studies at temperatures below (22 degrees C) and above (55 degrees C) the main endothermic transition confirm that the bilayer gel (sharp 4.2 A reflection)----bilayer liquid crystal (diffuse 4.5 A reflection) transition occurs at all hydration levels with limiting bilayer hydration occurring at approx. 31.5% H2O in the gel phase and at approx. 35% H2O in the liquid crystal phase. The thermotropic properties and metastability of this partial synthetic N-stearoyl sphingomyelin differ in some respects from that of the previously studied synthetic DL-erythro-N-stearoyl sphingomyelin (Estep, T.N., Calhoun, W.I., Barenholz, Y., Biltonen, R.L., Shipley, G.G. and Thompson, T.E. (1980) Biochemistry 19, 20-24), suggesting an influential role of the interfacial molecular conformation.

Calorimetry, Differential Scanning↗

PubMed1990

Structure and properties of mixed-chain phosphatidylcholine bilayers.

The structural and thermotropic properties of the hydrated mixed-chain phosphatidylcholines (PCs), C(8):C(18)-PC and C(10):C(18)-PC, have been studied by X-ray diffraction and differential scanning calorimetry. For fully hydrated C(8):C(18)-PC, the reversible chain melting transition is observed at 9.9 degrees C (delta H = 7.3 kcal/mol). X-ray diffraction at 0 degrees C (below the chain melting transition) shows a small bilayer repeat distance, d = 51.0 A, and a sharp, symmetric wide-angle reflection at 4.1 A, characteristic of a mixed interdigitated bilayer gel phase [see McIntosh, T. J., Simon, S. A., Ellington, J. C., Jr., & Porter, N. A. (1984) Biochemistry 23, 4038-4044; Hui, S. W., Mason, J. T., & Huang, C. (1984) Biochemistry 23, 5570-5577]. At 30 degrees C (above the chain melting transition), a diffuse band is observed at 4.5 A characteristic of an L alpha phase but with an increased bilayer periodicity, d = 61 A. Both the calculated lipid bilayer thickness (d1) and that determined directly from electron density profiles (dp-p) show unusual increases as a consequence of chain melting. In contrast, fully hydrated C(10):C(18)-PC shows an asymmetric endothermic transition at 11.8 degrees C. Below the chain melting transition, two lamellar phases are present, corresponding to coexisting interdigitated (d = 52.3 A) and noninterdigitated (d = 62.5 A) bilayer gel phases. The relative amounts of these phases depend upon the low-temperature incubation and/or hydration conditions, suggesting conversions, albeit kinetically complex, between metastable, and stable phases. The different behavior of C(8):C(18)-PC and C(10):C(18)-PC, as well as their positional isomers, is rationalized in terms of the molecular conformation of PC.

Calorimetry, Differential Scanning↗

PubMed1990

Crystallization and preliminary x-ray diffraction study of synthetic apolipoprotein E fragment (residues 129-169).

Apolipoprotein E is a plasma protein comprised of a lipid binding region (which together with other apoproteins maintains the structure of lipoprotein particles) and a receptor binding domain (which interacts with cellular receptors for control of triglyceride and cholesterol metabolism). A peptide, comprising residues 129-169 of human apolipoprotein E, which contains both a putative lipid-binding region and receptor binding domain, has been synthesized by solid phase techniques. Diffraction quality crystals of the synthetic apolipoprotein E fragment129-169 have been obtained at room temperature by vapor diffusion with polyethylene glycol in the presence of the nonionic detergent beta-octylglucoside. The crystals have been characterized with x-radiation as orthorhombic, space group I222 or I2(1)2(1)2(1), with unit cell dimensions a = 61.91, b = 30.84, and c = 42.79 A. There are eight molecules per unit cell, with one molecule (Mr = 4771) in each asymmetric unit. Precession photographs show that crystals diffract beyond 2.7-A resolution and are stable in the x-ray beam at room temperature for at least 200 h; thus, they can be used to collect three-dimensional data for a detailed crystallographic analysis.

Apolipoproteins E↗

PubMed1990

Hydrocarbon chain packing modes in lipids: effect of altered sub-cell dimensions and chain rotation.

The lateral hydrocarbon chain packing modes of lipids have been described in terms of specific hydrocarbon sub-cells as deduced from single crystal structural studies. To understand the changes in hydrocarbon chain packing in lipid bilayers induced by variations in temperature, hydration, ion-binding, etc., we have examined the effect on the calculated X-ray diffraction pattern of (a) systematic variations in the dimensions of the hydrocarbon sub-cell and (b) the effect of chain rotation at fixed lattice sites. For the O perpendicular (orthorhombic) sub-cell, the a and b sub-cell parameters were varied from as = 4.96 to 4.85 A and bs = 7.42 to 8.40 A in six steps and the positions (s = 2 sin theta/lambda) and intensities (Icalc = F2) of the strong sub-cell reflections calculated. In this way, the conversion of the O perpendicular sub-cell (with either fixed chain orientations or simulated chain rotation) to the hexagonal (H) sub-cell (with chain rotation) was followed. Notably, the two strong reflections characteristic of the O perpendicular sub-cell at 4.12 A (110) and 3.71 A (020) show progressive shifts in position and intensity, finally merging to give the strong (O1O) reflection at 4.2 A characteristic of the hexagonal sub-cell. Similar calculations were performed for the orthorhombic (O' perpendicular) and monoclinic (M parallel) sub-cells. This approach can be used to analyze changes in the X-ray diffraction data due to modifications of the hydrocarbon chain packing modes characteristic of simple and complex lipids.

Hydrocarbons↗

PubMed1990

Effect of chain-linkage on the structure of phosphatidyl choline bilayers. Hydration studies of 1-hexadecyl 2-palmitoyl-sn-glycero-3-phosphocholine.

While hydrated dipalmitoyl phosphatidylcholine (DPPC) forms tilted chain L beta' bilayers in the gel phase, the ether-linked analogue dihexadecyl phosphatidylcholine (DHPC) exhibits gel phase polymorphism. At low hydration DHPC forms L beta' phases but at greater than 30% H2O a chain-interdigitated gel phase is observed (Ruocco, M. J., D. S. Siminovitch, and R. G. Griffin. 1985. Biochemistry. 24:2406-2411; Kim, J.T., J. Mattai, and G.G. Shipley. 1987. Biochemistry. 26:6599-6603). In this study we report the behavior of a phosphatidylcholine (PC) with both types of chain linkage, 1-hexadecyl-2-palmitoyl-sn-glycero-3-phosphocholine (HPPC). HPPC has been investigated as a function of hydration using differential scanning calorimetry (DSC) and x-ray diffraction. By DSC, over the hydration range 5. 1-70.3 wt% H2O, HPPC exhibits two reversible transitions. The reversible main chain-melting transition decreases from 69 degrees C, reaching a limiting value of 40 degrees C at full hydration. X-ray diffraction patterns of hydrated HPPC have been recorded as a function of hydration at 20 degrees and 50 degrees C. At 50 degrees C, melted-chain L alpha bilayer phases are observed at all hydrations. At 20 degrees C, at low hydrations (less than 34 wt% H2O) HPPC exhibits diffraction patterns characteristic of bilayer gel phases similar to those of the gel phase of DPPC. In contrast, at greater than or equal to 34 wt% H2O, HPPC shows a much reduced bilayer periodicity, d = 47 A, and a single sharp reflection at 4.0 A in the wide angle region. This diffraction pattern is identical to that exhibited by the interdigitated phase of DHPC. Therefore, in the gel phase HPPC undergoes a hydration-dependent conversion from a regular bilayer structure to an interdigitated bilayer arrangement. Clearly, the presence of a single ether linkage (at the sn-i position) is sufficient to allow formation of the chain-interdigitated phase in a hydration-dependent way essentially identical to that of DHPC.

Calorimetry, Differential Scanning↗

PubMed1990

Structural studies of the lipid components of bile.

The lipid-rich molecular aggregates responsible for the transport of cholesterol in bile include mixed micelles and bilayer vesicles. In this review, the molecular conformations of the individual lipid components of biliary micelles and vesicles (phospholipids, bile salts and cholesterol) obtained by high-resolution x-ray crystallographical analysis are described. The key conformational/packing features relevant to lipid organization, molecular interactions and cholesterol solubility in bile are discussed. The structure of cholesterol monohydrate, the major component in cholesterol gallstones, is described and problems relevant to cholesterol crystal nucleation/growth processes are briefly discussed.

Bile↗

PubMed1989

Characterization of the L lambda phase in trehalose-stabilized dry membranes by solid-state NMR and X-ray diffraction.

Solid-state nuclear magnetic resonance (NMR) spectroscopy and X-ray powder diffraction were used to investigate the mechanism of trehalose (TRE) stabilization of lipid bilayers. Calorimetric investigation of dry TRE-stabilized bilayers reveals a first-order phase transition (L kappa----L lambda) at temperatures similar to the L beta'----(P beta')----L alpha transition of hydrated lipid bilayers. X-ray diffraction studies show that dry mixtures of TRE and 1,2-dipalmitoyl-sn-phosphatidylcholine (DPPC) have a lamellar structure with excess crystalline TRE being present. The L kappa phase shows typical gel-phase X-ray diffraction patterns. In contrast, the L lambda-phase diffraction patterns indicate disordered hydrocarbon chains. 2H NMR of specifically 2H chain-labeled DPPC confirmed that the acyl chains are disordered in the L lambda phase over their entire lengths. 2H spectra of the choline headgroup show hindered molecular motions as compared to dry DPPC alone, and 13C spectra of the sn-2-carbonyl show rigid lattice powder patterns indicating very little motion at the headgroup and interfacial regions. Thus, the sugar interacts extensively with the hydrophilic regions of the lipid, from the choline and the phosphate moieties in the headgroup to the glycerol and carbonyls in the interfacial region. We postulate that the sugar and the lipid form an extensive hydrogen-bonded network with the sugar acting as a spacer to expand the distance between lipids in the bilayer. The fluidity of the hydrophobic region in the L lambda phase together with the bilayer stabilization at the headgroup contributes to membrane viability in anhydrobiotic organisms.

Cell Survival↗

PubMed1989

Interactions of metal ions with phosphatidylserine bilayer membranes: effect of hydrocarbon chain unsaturation.

A combination of surface monolayer, scanning calorimetry, 31P NMR, and spin-label ESR techniques has been used to monitor the interactions of monovalent (NH4+, Na+, and Li+) and divalent (Ca2+) cations with phosphatidylserines (PS) differing in their levels of chain unsaturation. Comparisons are made between the disaturated dimyristoyl-, dipalmitoyl-, and dihexadecyl-PS (DMPS, DPPS, and DHPS), saturated cis-monounsaturated palmitoyloleoyl-PS (POPS) (and bovine brain PS), di-trans-monounsaturated dielaidoyl-PS (DEPS), and di-cis-monounsaturated dioleoyl-PS (DOPS). Na+ and NH4+ cations interact weakly with all PS monolayers and bilayers without significant changes in molecular conformation, chain packing, or headgroup dynamics and without dependence on chain composition. In contrast, considering these structural and dynamic parameters, Li+ shows a gradation in its interaction with PS (DMPS greater than POPS approximately bovine brain PS greater than DOPS), suggesting that Li+-PS interactions depend on the interfacial properties of the PS molecules (e.g., surface area). Finally, Ca2+ interacts strongly with all PS monolayers and bilayers, without obvious chain selectivity. Thus, ion binding to PS depends not only on the properties of the cation (Na+ vs Li+ vs Ca2+) but also on the molecular details of the PS membrane surface.

Calcium↗

PubMed1989

Effect of chain unsaturation on the structure and thermotropic properties of galactocerebrosides.

Differential scanning calorimetry (DSC) and x-ray diffraction have been used to study the effect of increasing chain-unsaturation on the structure and properties of the hydrated cerebrosides N-stearoyl, -oleoyl, and -linoleoyl galactosylsphingosine (NSGS, NOGS, and NLnGS, respectively). DSC of hydrated (70 wt% water) NSGS shows an endothermic transition at 85 degrees C (delta H = 18.0 kcal/mol NSGS) and a broad exothermic transition at 40-60 degrees C, the latter being dependent upon the previous cooling rate. X-Ray diffraction patterns recorded at 21, 61, and 86 degrees C provide evidence for interconversions between metastable and stable crystalline NSGS bilayer phases. The properties of the unsaturated-chain cerebrosides are more complex. Hydrated NOGS shows a single endothermic transition at 44.8 degrees C (delta H = 11.5 kcal/mol NOGS). However, incubation of NOGS at 49 degrees C for 24 h results in a second transition at 55.5 degrees C. By cycling NOGS between 0 and 49 degrees C complete conversion into this higher melting phase (delta H = 12.1 kcal/mol NOGS) is achieved. X-ray diffraction confirms a bilayer phase at all temperatures and delineates the conversions between a crystalline phase at 21 degrees C (bilayer period d = 56.5A), a second crystalline phase at 47 degrees C (d = 69.9A), and a liquid crystalline phase at 59 degrees C (d = 52.0A). The more unsaturated NLnGS shows two transitions, a sharp transition at 28 degrees C (delta H = 8.0 kcal/mol NLGS) and a broad, low-enthalpy transition at 42 degrees C (delta H = 0.4 kcal/mol NLGS). Again, incubation between the two transitions leads to a single transition at 44 degrees C (delta H = 9.3 kcal/mol NLGS). X-ray diffraction demonstrates conversions between two crystalline bilayer phases (d = 55.2A and d = 68.4A), and a liquid crystalline bilayer phase (d = 51.8A). Thus, increased unsaturation in the amide-linked fatty acyl chain of cerebrosides results in decreased chain-melting temperatures (NSGS greater than NOGS greater than NLnGS) and has marked effects on their structural properties.

Calorimetry, Differential Scanning↗

PubMed1988

Crystallization and preliminary x-ray diffraction study of cholera toxin B-subunit.

Cholera toxin binds to its ganglioside GM1 receptor via its B-subunit, a pentameric assembly of identical subunits (Mr = 11,600). Diffraction quality crystals of cholera toxin B-subunit have been obtained at room temperature by vapor diffusion with polyethylene glycol in the presence of the nonionic detergent beta-octyl glucoside. The crystals have been characterized with x-radiation as monoclinic, space group P21, with unit cell dimensions a = 39.0 A, b = 94.3 A, c = 67.5 A, beta = 96.0 degrees. There are two molecules per unit cell, with one molecule (Mr = 58,000) in each asymmetric unit. Precession photographs (micron = 13 degrees) show that crystals diffract beyond 3.3-A resolution and are stable in the x-ray beam at room temperature for at least 40 h; thus, they can be used to collect three-dimensional crystallographic data.

Cholera Toxin↗

PubMed1987

Gel phase polymorphism in ether-linked dihexadecylphosphatidylcholine bilayers.

The structure and properties of the ether-linked 1,2-dihexadecylphosphatidylcholine (DHPC) have been examined as a function of hydration. By differential scanning calorimetry, DHPC exhibits an endothermic (chain melting) transition with the transition temperature (limiting value, 44.2 degrees C) and enthalpy (limiting value, delta H = 8.0 kcal/mol) being hydration dependent. For hydration values greater than 30 wt % water, DHPC exhibits a pretransition at approximately 36 degrees C (delta H = 1.1 kcal/mol) and a subtransition at approximately 5 degrees C (delta H = 0.2 kcal/mol). By X-ray diffraction, at 22 degrees C DHPC exhibits a normal bilayer gel structure with the bilayer periodicity increasing from 58.0 to 62.5 A over the hydration range 9.5-25.4% water. At 30-32% water, two coexisting gel phases are observed with d = 63-64 A and d = 44-45 A; at higher hydration, only the latter phase is present, reaching a limiting d = 47.0 A at 37.5% water. Two different gel phases clearly exist at low and high hydrations. Electron density profiles at low hydration (9.5-25.4%) show a bilayer thickness dp-p = 46 A, whereas at greater than 32% water the bilayer thickness is markedly reduced, dp-p = 30 A. These and other structural parameters indicate a hydration-dependent gel----gel structural transition between a normal bilayer (two chains per polar group) and the chain-interdigitated bilayer (four chains per polar group) arrangement described previously for DHPC [Ruocco, M. J., Siminovitch, D. J., & Griffin, R. G. (1985) Biochemistry 24, 2406-2411].(ABSTRACT TRUNCATED AT 250 WORDS)

Calorimetry, Differential Scanning↗

PubMed1987

Bilayer interactions of ether- and ester-linked phospholipids: dihexadecyl- and dipalmitoylphosphatidylcholines.

Mixed phospholipid systems of ether-linked 1,2-dihexadecylphosphatidylcholine (DHPC) and ester-linked 1,2-dipalmitoylphosphatidylcholine (DPPC) have been studied by differential scanning calorimetry and X-ray diffraction. At maximum hydration (60 wt % water), DHPC shows three reversible transitions: a main (chain melting) transition, TM = 44.2 degrees C; a pretransition, TP = 36.2 degrees C; and a subtransition, TS = 5.5 degrees C. DPPC shows two reversible transitions: TM = 41.3 degrees C and TP = 36.5 degrees C. TM decreases linearly from 44.2 to 41.3 degrees C as DPPC is incorporated into DHPC bilayers; TP exhibits eutectic behavior, decreasing sharply to reach 23.3 degrees C at 40.4 mol % DPPC and then increasing over the range 40-100 mol % DPPC; TS remains constant at 4-5 degrees C and is not observed at greater than 20 mol % DPPC. At 50 degrees C, X-ray diffraction shows a liquid-crystalline bilayer L alpha phase at all DHPC:DPPC mole ratios. At 22 degrees C, DHPC shows an interdigitated bilayer gel L beta phase (bilayer periodicity d = 47.0 A) into which approximately 30 mol % DPPC can be incorporated. Above 30 mol % DPPC, a noninterdigitated gel L beta' phase (d = 64-66 A) is observed. Thus, at T greater than TM, DHPC and DPPC are miscible in all proportions in an L alpha bilayer phase. In contrast, a composition-dependent gel----gel transition between interdigitated and noninterdigitated bilayers is observed at T less than TP, and this leads to eutectic behavior of the DHPC/DPPC system.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1987

Mixed-chain phosphatidylcholine bilayers: structure and properties.

Calorimetric and X-ray diffraction data are reported for two series of saturated mixed-chain phosphatidylcholines (PCs), 18:0/n:0-PC and n:0/18:0-PC, where the sn-1 and sn-2 fatty acyl chains on the glycerol backbone are systematically varied by two methylene groups from 18:0 to 10:0 (n = 18, 16, 14, 12, or 10). Fully hydrated PCs were annealed at -4 degrees C and their multilamellar dispersions characterized by differential scanning calorimetry and X-ray diffraction. All mixed-chain PCs form low-temperature "crystalline" bilayer phases following low-temperature incubation, except 18:0/10:0-PC. The subtransition temperature (Ts) shifts toward the main (chain melting) transition temperature (Tm) as the sn-1 or sn-2 fatty acyl chain is reduced in length; for the shorter chain PCs (18:0/12:0-PC, 12:0/18:0-PC, and 10:0/18:0-PC), Ts is 1-2 degrees C greater than Tm, and the subtransition enthalpy (delta Hs) is much greater than for the longer acyl chain PCs. Tm decreases with acyl chain length for both series of PCs except 18:0/10:0-PC, while for the positional isomers, n:0/18:0-PC and 18:0/n:0-PC, Tm is higher for the isomer with the longer acyl chain in the sn-2 position of the glycerol backbone. The conversion from the crystalline bilayer Lc phase to the liquid-crystalline L alpha phase with melted hydrocarbon chains occurs through a series of phase changes which are chain length dependent. For example, 18:0/18:0-PC undergoes the phase changes Lc----L beta'----P beta'----L alpha, while the shorter chain PC, 10:0/18:0-PC, is directly transformed from the Lc phase to the L alpha phase. However, normalized enthalpy and entropy data suggest that the overall thermodynamic change, Lc----L alpha, is essentially chain length independent. On cooling, the conversion to the Lc phases occurs via bilayer gel phases, L beta', for the longer chain PCs or through triple-chain interdigitated bilayer gel phases, L beta, for the shorter chain PC 18:0/12:0-PC and possibly 10:0/18:0-PC. Molecular models indicate that the bilayer gel phases for the more asymmetric PC series, 18:0/n:0-PC, must undergo progressive interdigitation with chain length reduction to maintain maximum chain-chain interaction. The L beta phase of 18:0/10:0-PC is the most stable structure for this PC below Tm. The formation and stability of the triple-chain structures can be rationalized from molecular models.

Calorimetry↗

PubMed1987

Regulation of bilayer stability in Clostridium butyricum: studies on the polymorphic phase behavior of the ether lipids.

Three of the major phospholipids of the cell membrane of Clostridium butyricum are phosphatidylethanolamine (PE), plasmenylethanolamine (PlaE), and the glycerol acetal of plasmenylethanolamine. When cultured in the absence of biotin in media supplemented with a cis-unsaturated fatty acid, the cellular lipids become highly enriched with the fed fatty acid. Under these conditions, the ratio of the glycerol acetal of PlaE to the sum of PE plus PlaE increases markedly over that seen in cells containing mixtures of saturated and unsaturated fatty acids [Johnston, N.C., & Goldfine, H. (1985) Biochim. Biophys. Acta 813, 10-18]. We have studied the polymorphic phase behavior of the phospholipids from C. butyricum grown on oleic acid using differential scanning calorimetry, 31P nuclear magnetic resonance, and X-ray diffraction. The mixed PE plus PlaE fraction undergoes a transition from the gel to liquid-crystalline state at -1.9 degrees C and a lamellar to reversed hexagonal (L----H) transition at or near 0 degrees C. The glycerol acetal of PlaE melts at 16.1 degrees C, and as predicted from lipid packing theory, the lamellar phase is stabilized, up to 50 degrees C. Addition of the oleate-enriched glycerol acetal of PlaE to dioleoylphosphatidylethanolamine, or the PE plus PlaE fraction from oleate-grown cells, stabilized the lamellar arrangement of the mixtures. A ratio of glycerol acetal of PlaE to total PE (PE plus PlaE) of 0.5, which is close to that found in cells grown on palmitic plus oleic acid, 0.6-0.7, did not produce a lamellar phase at 37 degrees C when the lipids enriched with oleic acid were tested,(ABSTRACT TRUNCATED AT 250 WORDS)

Calorimetry, Differential Scanning↗

PubMed1987

Interaction of cholera toxin with ganglioside GM1 receptors in supported lipid monolayers.

Lipid monolayers formed at the air-water interface containing the ganglioside GM1 in egg yolk phosphatidylcholine have been transferred according to the Langmuir-Blodgett technique to glass cover slips coated with octadecyl- or hexadecyltrichlorosilane and carbon-coated electron microscope grids. Monolayer transfer has been demonstrated with fluorescence microscopy, by the transfer of a fluorescent phospholipid analogue, N-(7-nitro-2,1,3-benzoxadiazol-4-yl)phosphatidylethanolamine or Lucifer yellow labeled GM1 (LY-GM1), incorporated into the lipid monolayer. Incubation of supported monolayers with solutions of fluorescein-labeled cholera toxin (FITC cholera toxin) resulted in specific binding of the toxin to monolayers containing GM1, as revealed by fluorescence microscopy. Lateral diffusion coefficients were measured for both the receptor (LY-GM1) [(3.9 +/- 2.1) X 10(-8) cm2/s] and the receptor-ligand complex (GM1-FITC cholera toxin) [(8.9 +/- 3.2) X 10(-9) cm2/s] according to the technique of fluorescence recovery after photobleaching. In separate studies, GM1-containing monolayers transferred to electron microscope grids were incubated with solutions containing unlabeled cholera toxin, followed by negative staining with uranyl acetate. Electron microscopy revealed patches of stained cholera toxin molecules (diameter approximately 70 A) in crystalline, two-dimensional hexagonal arrays. Optical diffraction and image reconstruction showed the arrangement of the cholera toxin molecules in a planar hexagonal cell, a = 81 A. These initial reconstructions give structural information to a resolution of approximately 30 A and indicate a doughnut-shaped molecule with a central aqueous channel.

Animals↗

PubMed1987

Structure and thermotropic properties of hydrated 1-eicosyl-2-dodecyl-rac-glycero-3-phosphocholine and 1-dodecyl-2-eicosyl-rac-glycero-3-phosphocholine bilayer membranes.

The ether-linked phosphatidylcholines 1-eicosyl-2-dodecyl-rac-glycero-3-phosphocholine (EDPC) and 1-dodecyl-2-eicosyl-rac-glycero-3-phosphocholine (DEPC) have been investigated by differential scanning calorimetry (DSC) and X-ray diffraction. DSC of hydrated EDPC shows a single endothermic transition at 34.8 degrees C (delta H = 11.2 kcal/mol) after storage at -4 degrees C while DEPC shows three endothermic transitions at 7.7 and approximately 9.0 degrees C (combined delta H approximately 0.4 kcal/mol) and at 25.2 degrees C (delta H = 4.7 kcal/mol). Both the single transition of EDPC and the two higher temperature transitions of DEPC are reversible, while the approximately 7.7 degrees C transition of DEPC increases in enthalpy on low-temperature incubation. At 23 degrees C, X-ray diffraction of hydrated EDPC shows a sharp reflection at 4.2 A together with lamellar reflections corresponding to a bilayer periodicity, d = 56.2 A. Electron density profiles derived from swelling experiments show a phosphate-phosphate intrabilayer distance, dp-p, of 36 A at all hydrations. This, together with calculated lipid thickness and molecular area considerations, suggests an interdigitated, three chains per head group, bilayer gel phase, L beta*, with no hydrocarbon chain tilt. This is structurally analogous to the bilayer gel phase of hydrated 18:0/10:0 ester PC [McIntosh, T. J., Simon, S. A., Ellington, J. C., Jr., & Porter, N. A. (1984) Biochemistry 23, 4038]. In contrast, DEPC at -4 degrees C shows an L beta' bilayer gel phase with tilted hydrocarbon chains (d = 61.1 A). However, this transforms above 9 degrees C to an interdigitated, triple-chain, L beta* bilayer gel phase (identical with that of EDPC) with d = 56.6 A and a phosphate-phosphate distance of 36 A. Above their respective chain melting transitions, Tm, EDPC and DEPC exhibit liquid-crystalline L alpha bilayer phases with d = 64.5 and 65.0 A at 55 and 45 degrees C, respectively. The ability of both EDPC and DEPC to form triple-chain interdigitated gel-state bilayers suggests that the conformational inequivalence at the sn-1 and sn-2 positions is less pronounced in the ether-linked PCs compared to the ester-linked PCs, where only one of the positional isomers, e.g., 18:0/10:0 PC but not 10:0/18:0 PC, forms the triple-chain structure (J. Mattai, unpublished results). Thus, a different conformation around the glycerol is predicted for ether-linked PC compared to ester-linked PC.

Calorimetry, Differential Scanning↗