Conformation and packing of unsaturated chains in cholesteryl linolelaidate at 123 K

At 123 K, crystals of cholesteryl trans-9-trans-12-octadecadienoate (cholesteryl linolelaidate, C47H76O2) are monoclinic, space group P2(1) with cell dimensions a = 13.03(3), b = 8.76(2), c = 17.90(4) A, beta = 89.7(2) degrees, having two molecules per unit cell. The crystal structure has been determined from 2041 X-ray intensities with sin theta/lambda less than 0.48 A-1, of which 922 gave I greater than 2 sigma(I). The hydrogen atoms were found in a difference Fourier synthesis. Block diagonal least squares refinement assuming isotropic thermal parameters has converged with Rw = 0.13. The molecule is fully extended (length 43.3 A), except for a symmetric bowing in the linolelaidate chain segment which contains the two unconjugated trans ethylenic bonds. The torsion angles at the four C--C bonds adjacent to the C=C bonds are all in the preferred (+/-)-skew range. Chain packing is efficient, without having a regular subcell structure. There is a similarity with the overall conformation of the oleate chains in crystals of cholesteryl oleate. Although chemically disparate, the oleate and linolelaidate chains have similar crystal environments.     

Transformation of macrophages into foam cells in vitro induced by cholesteryl oleate liquid crystals

Transformation of macrophages into foam cells after the uptake of cholesteryl oleate anisotropic liquid crystals was studied. A new technique to enhance the uptake of the liquid crystals by macrophages using an inverted petri dish was developed. Uptake of lipid droplets was found to increase in parallel with the amount of liquid crystals in the medium. A lysosomal enzyme was shown (by using lysosomotropic chloroquine) to be involved in the hydrolysis of the liquid crystals. About 47 and 72% of the [3H]cholesterol in liquid crystal-laden cells had disappeared after chase for 24 and 48 h, respectively. Thus the 50% clearance time of the liquid crystals by the macrophages was about 24 h, which was longer than that of denatured lipoprotein. A possible model of transformation of macrophages to foam cells is discussed.     

Properties of cholesteryl oleate and triolein in mixed monolayers at the air-water interface

The properties of cholesteryl oleate and triolein in mixed monolayers at the air-water interface have been measured between 24 and 37 degrees C. Analysis of force-area curves obtained as a function of the mol fraction of cholesteryl oleate indicates that at relatively low surface pressures these compounds are miscible in two dimensions up to a limit of about 0.5 mol fraction. At higher pressures either cholesteryl oleate or both lipids are expelled from the monolayer to form a bulk phase which is in rapid equilibrium with the surface phase. In the monolayer phase, orientation of the ester function of cholesteryl oleate is toward the aqueous phase, interaction with triolein is minimal, and packing is uniform over the solubility range. This together with the susceptibility of the cholesteryl oleate to enzymatic hydrolysis, suggests the applicability of monolayer systems to the study of cholesterol esterase activity. Comparison of our results with the bulk properties of these lipids suggests that the expelled cholesteryl oleate exists as a smectic mesophase and thus the system may provide a model for studying the transfer of molecules between the interior and surface of lipid deposits of the type found in atherosclerotic lesions.     

IR spectroscopic study of phospholipid emulsions containing cholesteryl oleate

As models for the lipid organization of low density lipoproteins (LDL), protein-free aqueous emulsions are prepared from dimyristoyl phosphatidyl choline (DMPC), dipalmitoyl phosphatidyl choline (DPPC), and cholesteryl oleate (CO). Aqueous dispersions containing these lipids are sonicated and yield stable particles with diameters varying between 20 and 40 nm as measured through electron microscopy. IR spectroscopy shows that emulsions consisting of DMPC, DPPC, and CO at 3/1/1 and 1/1/1 ratios undergo specific thermal transitions, depending on their composition, that can be assigned to the phospholipids forming the surface layer of the emulsion particles and to core-located CO. However, at the 1/3/1 DMPC/DPPC/CO ratio this lipid system exhibits an order-disorder transition of the mixed phospholipids with no significant transition associated with core-located CO. Observation of the methylene C&bond;H and C&bond;D stretching modes of nondeuterated and deuterated lipids enables the packing characteristics and conformational order of each lipid to be monitored separately. The transition temperature changes compared to the temperatures for the analogous transitions in neat CO and CO-free phospholipid vesicles suggest the existence of interactions between CO and the above phospholipids in the ternary emulsion particles; these interactions are stronger at the 1/3/1 DMPC/DPPC/CO ratio. The results show that interactions between core and surface phases are dependent on the emulsion lipid composition and that these findings may be extended to native lipoproteins.     

Regulation of cholesteryl oleate and triolein miscibility in monolayers and bilayers

The miscibility of triolein and cholesteryl oleate with 1-palmitoyl-2-oleoyl phosphatidylcholine was studied at the argon-buffer interface. The surface phase behavior of the system was analogous to that for cholesteryl ester-phospholipid mixtures in that both monolayer and double layer surface phases were formed. By considering the bulk properties of cholesteryl oleatetriolein mixtures and the two-dimensional phase rule, the entire system could be described. Double layer properties suggest that it consists of mostly triolein and phospholipid in the layer adjacent to the aqueous phase. The monolayer phase shows the formation of complexes between the neutral lipids and the phospholipid with stoichiometries nearly identical with those reported for bilayers (Hamilton, J. A., Miller, K. W., and Small, D. M. (1983) J. Biol. Chem. 258, 12821-12826). A second complex with a 3:1 stoichiometry is formed between triolein and cholesteryl oleate independently of interactions with phospholipid. Upon interaction with phospholipid, the triolein-cholesteryl oleate complex loses proportionately more area than either lipid alone. Because the area of complexes with phospholipid is constant, overall neutral lipid miscibility in such complexes is enhanced by the cholesteryl oleate-triolein interaction. Thus, our data explain the apparently nonideal mixing of cholesteryl oleate, triolein, and phospholipid in monolayers and in bilayers.     

Conformation of aspartate aminotransferase in crystals

X-ray study of chicken cytosolic aspartate aminotransferase revealed conformational changes in the protein of two kinds: (1) a shift of the small domain adjacent to substrate-binding area due to interaction of the protein with two carboxyl groups of substrate and (2) a change in inclination of the coenzyme plane due to replacement of C = N bond of the coenzyme with Lys-258 by C = N bond with a substrate. An asymmetry in subunit behaviour is observed in both cases: the domain is shifted in one subunit and the coenzyme is rotated in other. Substrate-binding properties of each subunit are strictly dependent on the protein conformation in substrate-binding area.     

Intravascular metabolism of lipoprotein cholesteryl esters in African green monkeys: differential fate of doubly labeled cholesteryl oleate

High density lipoproteins (HDL), doubly labeled with [3H]cholesteryl oleate and cholesteryl [14C]oleate, were reinjected to study HDL cholesteryl ester metabolism in African green monkeys. The transfer of labeled HDL cholesteryl ester to low density lipoprotein (LDL) was rapid and equilibration of the [3H]cholesteryl oleate and cholesteryl [14C]oleate specific activities in LDL and HDL occurred within 90 min after reinjection. The apparent rates of disappearance from the circulation of the two moieties of the cholesteryl ester were different. In the same four animals, the residence time for the turnover of plasma [3H]cholesterol averaged 6.1 days while the residence time for the removal of cholesteryl [14C]oleate from plasma was approximately 2.1 days. These results suggest that for some lipoprotein cholesteryl esters removed from plasma, the cholesterol moiety subsequently reappeared in plasma. The difference between the rate of decay of the 14C-labeled fatty acid moiety, which represents all of the cholesteryl ester removed from plasma (0.48 pools/day) and the decay of the 3H-labeled cholesterol moiety, which represents the sum of cholesteryl ester removal and cholesterol reappearance (0.16 pools/day), is the fraction of the cholesteryl ester pool recycled per day (0.32 pools/day or 22.5 mg/kg per day). In other words, approximately 68% of the cholesterol moiety that was removed from plasma as cholesteryl oleate reappeared in the plasma cholesterol pool. These studies support the concept that an efficient reutilization cycle for plasma cholesterol occurs, i.e., the cholesteryl ester molecule can exit and the cholesterol moiety can re-enter plasma without effective equilibration of the cholesterol moiety with extravascular cholesterol pools.     

Orientational order of cholesteryl oleate in low-density lipoprotein observed by 2H-NMR

Cholesteryl oleate, selectively deuterated at various positions along the acyl chain, has been incorporated into fresh human serum low-density lipoprotein (LDL2). Temperature-dependent 2H-NMR spectra were recorded between 15 and 45 degrees C. For deuterons at C-2' and C-5' of the acyl chain, two 2H-NMR spectral components, a broad and a narrow signal, are observed. This is interpreted as reflecting the coexistence of two cholesteryl ester regions in the LDL2 core which possess different degrees of order. The C-2H bond order parameters, SCD, are approx. 0.12-0.20 for the more ordered region and approx. 0.04-0.06 for the less ordered region. Longitudinal relaxation times, T1, of deuterated cholesteryl oleate are found to increase between C-8' and the terminal -C2H3 group, which is consistent with an increased rate of chain motion toward the free ends of the ester acyl chains.     

Microemulsions of cholesteryl oleate and dimyristoylphosphatidylcholine: a model for cholesteryl ester rich very low density lipoproteins

This study describes the preparation, purification, and characterization of a cholesteryl oleate/dimyristoylphosphatidylcholine microemulsion as a model for the interaction of lipid domains in cholesteryl ester rich very low density lipoproteins. These lipids were chosen specifically because their thermal transitions were distinct from each other, and their differences in chemical structure permitted the motion(s) of each lipid component to be monitored independently by 13C nuclear magnetic resonance (NMR). The model particles were formed by cosonication of cholesteryl oleate and dimyristoylphosphatidylcholine in a 4:1 molar ratio for 45 min at 55-60 degrees C (above both lipid phase transition temperatures). The crude microemulsion was fractionated by low-speed centrifugation and Sepharose CL-2B chromatography. Microemulsion particles which eluted from the column at a volume similar to that of cholesteryl ester rich very low density lipoproteins had high cholesteryl ester:phospholipid ratios (2.5:1----6:1). Electron micrographs of negatively stained particles showed them to be large spheres devoid of multilamellar or unilamellar vesicle structures. Particle size calculated from a simple compositional model correlated well with sizes determined by electron microscopy (500-1000 A) for various column fractions. Differential scanning calorimetry studies of the microemulsion revealed two thermal transitions for the model particles, at 31.0 and 46.6 degrees C, which were tentatively assigned to the surface phospholipid and core cholesteryl ester domains, respectively. These assignments were confirmed by 13C NMR which demonstrated that, at temperatures near the lower thermotropic transition, only resonances derived from carbon atoms of dimyristoylphosphatidylcholine (DMPC) were observable. As the temperature was raised to 38.6 degrees C, resonances from the olefinic carbons in the cholesteryl ester acyl chain appeared in the spectrum. At 46.6 degrees C, the center of the higher temperature endotherm, resonances from both the steroid ring and remaining acyl chain carbons of cholesteryl oleate became observable in the spectrum. Further increases in temperature did not result in the appearance of new resonances; however, those that were present narrowed and increased in intensity. The elevation in transition temperature for DMPC in these particles (31 degrees C) as compared to that for DMPC in small unilamellar (18 degrees C) and large multilamellar (23 degrees C) vesicles suggested a stabilization of the phospholipid monolayer, possibly by interaction with the nonpolar core lipids.(ABSTRACT TRUNCATED AT 400 WORDS)     

Crystal structure of form II of cholesteryl palmitelaidate at 295 K

Form II for cholesteryl palmitelaidate (trans-9-hexadecenoate) (C43H74O2) is monoclinic P2(1) with a = 12.745(3), b = 9.006(2), c = 18.153(4) A, beta = 96.63 (2) degrees, Z = 2. The X-ray crystal structure of form II has been determined from 2506 reflections of which 2126 gave (F greater than 2 sigma). The data up to sin theta/lambda = 0.44A-1 (Dmin = 1.14 A) were measured with CuK alpha radiation from a sealed tube. These were supplemented up to sin theta/lambda = 0.52 A-1 (Dmin = 0.96 A) by measurements on the same crystal using a rotating anode X-ray source. The electron density was diffuse in the ester chain and the atoms of the cholesteryl tail were found to be disordered. The tail and the chain atoms were refined by restrained least squares methods to give R = 0.087 and Rw = 0.10 for reflections with F greater than 2 sigma. Crystal forms I and II represent two standard structure types already characterized for fatty acid esters of cholesterol. In form II, the ester chain is almost fully extended as is also the case for one of the two independent molecules (A) in form I. In form II, the chains pack loosely together for most of their length. M.s. amplitudes of thermal vibration for the chain C-atoms are almost uniform along the entire chain (approximately 0.25 A2 at 295 K). In form I, the proximal part of the A chain is surrounded by rigid cholesteryl groups. In this region, C-atom m.s. amplitudes are much reduced (approximately 0.10 A2) but they increase to about 0.5 A2 at the distal end of the chain where packing is very loose.     

Conformation analysis of carbohydrate chains of glycoconjugates

A problem of conformations of carbohydrate chains of glycoconjugates-glycoproteins and glycolipids--is reviewed. Experimental data (NMR, X-Ray) and theoretical conformational analysis data are discussed. Spatial structures of O-linked oligosaccharides from blood-group glycoproteins, N-linked oligosaccharides of different types (oligomannosidic, complex, hybrid, bisect) and carbohydrate chains of glycosphingolipids are considered.     

LDL particle core enrichment in cholesteryl oleate increases proteoglycan binding and promotes atherosclerosis

Several studies in humans and animals suggest that LDL particle core enrichment in cholesteryl oleate (CO) is associated with increased atherosclerosis. Diet enrichment with MUFAs enhances LDL CO content. Steroyl O-acyltransferase 2 (SOAT2) is the enzyme that catalyzes the synthesis of much of the CO found in LDL, and gene deletion of SOAT2 minimizes CO in LDL and protects against atherosclerosis. The purpose of this study was to test the hypothesis that the increased atherosclerosis associated with LDL core enrichment in CO results from an increased affinity of the LDL particle for arterial proteoglycans. ApoB-100-only Ldlr^−/− mice with and without Soat2 gene deletions were fed diets enriched in either cis-MUFA or n-3 PUFA, and LDL particles were isolated. LDL:proteogylcan binding was measured using surface plasmon resonance. Particles with higher CO content consistently bound with higher affinity to human biglycan and the amount of binding was shown to be proportional to the extent of atherosclerosis of the LDL donor mice. The data strongly support the thesis that atherosclerosis was induced through enhanced proteoglycan binding of LDL resulting from LDL core CO enrichment.     

Conformation and intermolecular interactions of carbohydrate chains

For consideration of their conformations and interactions, carbohydrate chains can conveniently be divided into 3 classes on the basis of their covalent structure; namely periodic (a), interrupted periodic (b), and aperiodic (c) types. In aqueous solution carbohydrate chains often exist as highly disordered random coils. Under appropriate conditions, however, polysaccharides of types (a) and (b) can adopt a variety of ordered conformations. Physical methods, and in particular optical rotation, circular dichroism, and nuclear magnetic resonance, provide sensitive probes for the study of the mechanism and specificity of these disorder-order transitions in aqueous solution. Intermolecular interactions between such polysaccharide chains arise from co-operative associations of long structurally regular regions which adopt the ordered conformations. For acidic polysaccharides these cooperative associations may involve alignment of extended ribbons with cations sandwhiched between them. In other systems the interactions involve double belices which may then aggregate further, and geometric “matching” of different polysaccharide chains can also occur. These ordered, associated regions are generally terminated by deviations from structural regularity or by “kinks” which prevent complete aggregation of the molecules. The complex carbohydrate chains which occur at the periphery of animal cells have very different, aperiodic structures and although their conformations are as yet poorly understood, preliminary indications are considered.     

Conformation of polydispersed chains grafted on nanoparticles

The conformations of polydispersed polymer chains grafted on nanoparticles (NPs) are investigated by coarse-grained molecular dynamics simulations. Combined with the geometric and steric features, we find that most results can be understood by analysing the excluded volume interaction condition of the accommodated chains. By controlling suitable NP size and grafting density, as well as designing a suitable route for preparing the grafted chains with controllable polydispersity and length, it is possible to obtain polymer chains with various conformations, which may be greatly helpful for improving the dispersion of the grafted NPs in the polymer matrix. These results shed light on improved designs of grafted NPs and a better control of dispersion in polymer matrices for promoting the performance of polymer nanocomposite materials.     

Conformation analysis of carbohydrate chains of group-specific substances

Theoretical conformational analysis of bi- and three-antennary carbohydrate chains of H-specific group substances has been carried out. It has been shown that O-glycosylating oligosaccharides can form compact Y-shaped structures with effective non-bonded interactions between the antennae residues.     

Study of the miscibility of cholesteryl oleate in a matrix of ceramide, cholesterol and fatty acid

Cholesteryl esters (CE) are not generally abundant but are ubiquitous in living organisms and have markedly different properties from cholesterol because of their acyl chain. The miscibility/immiscibility of CE with biological lipid structures is a key property for their functions. In this work we study the solubility of cholesteryl oleate (ChO) in a model of the stratum corneum lipid matrix composed of ceramide C16, cholesterol and palmitic acid in excess water. Experiments were done in conditions of fully ionized (pH = 9.0) and fully neutralized fatty acid (pH = 4.0), and differential scanning calorimetry of the ternary mixtures with added ChO at pH = 9.0 clearly displayed a main transition with the same maximum temperature, peak shape, and enthalpy, suggesting that ChO was excluded from the remaining lipids. This technique is not conclusive at pH = 4.0 because the transitions of the lipid matrix and ChO overlap. The insolubility of ChO at both pH values is supported by X-ray diffraction. Adding the ceramide:cholesterol:fatty acid lipid mixture to ChO did not change the X-ray pattern of the mixture nor that of the ChO. To supplement the above physical techniques, we applied ¹³C MAS NMR spectroscopy with C-13 carbonyl-labeled ChO. A single ¹³C carbonyl peak from the ChO at 171.5 ppm was observed, indicating exposure to only one environment. The chemical shift was identical to pure ChO below and above the temperature of isotropic liquid formation. Taken together, our results lead to the conclusion that the solubility of ChO is negligible in the ceramide:cholesterol:fatty acid lipid mixture.     

The crystal structure of isopropyl 1-thio-beta-D-galactopyranoside monohydrate at 123 K

The crystal structure of isopropyl 1-thio-beta-D-galactopyranoside monohydrate is orthorhombic, P2(1)2(1)2(1), Z = 4, with cell dimensions at 123 K [293 K] of a = 7.983(1) [8.037(1)], b = 24.574(5) [24.709(4)], c = 6.329(1) [6.3736(8)] A, V = 1241.84 [1265.71] A3. The calculated and measured density is Dx = 1.371 [1.345] g cm-3, Dm = [1.340] g cm-3. Diffraction data were obtained with CuK alpha radiation and a Nonius CAD-4 diffractometer. The structure was solved by using MULTAN, and refined to R(F2) = 0.051, RW(F2) = 0.078, R(F) = 0.029, S = 1.16 for 1502 reflections. The molecule has the 4C1(D) conformation. The orientation of the primary alcohol group is gauche/trans, and that about the glycosidic C-S bond is (-)synclinal relative to the ring C-O bond. Although this compound does not form thermotropic liquid crystals, it has two crystal-to-crystal phase-transitions, at 70 and 104 degrees, prior to melting at 126 degrees. The crystal structure has a characteristic, amphiphilic, head-to-head bilayer molecular packing, with intercalated alkyl groups. The water molecule is included in the hydrogen-bond structure that links the galactoside moieties.     

Lipoprotein lipase hydrolysis of trioleoylglycerol in a phospholipid interface. Effect of cholesteryl oleate on catalysis

The effect of cholesteryl oleate on the lipoprotein lipase-catalyzed hydrolysis of trioleoylglycerol was determined in monolayers of egg phosphatidylcholine at a constant surface pressure of 24 mN m-1. The phospholipid monolayers contained 1.0 to 7.5 mol % trioleoylglycerol and various amounts (0 to 20 mol %) of cholesteryl oleate. The initial rates of trioleoylglycerol hydrolysis were determined with lipoprotein lipase purified from bovine milk. In phospholipid monolayers containing 5.0 or 7.5 mol % trioleoylglycerol, the further addition of cholesteryl oleate caused a decrease in lipoprotein lipase activity. In contrast, addition of cholesteryl oleate to phospholipid monolayers containing 1.0 or 2.5 mol % trioleoylglycerol enhanced enzyme activity; a 3-fold enhancement was observed with 5.0-7.5 mol % cholesteryl oleate. Based on force-area measurements, the cholesteryl ester-mediated decrease in lipoprotein lipase activity observed at high substrate concentrations may be explained by displacement of trioleoylglycerol from the interface, thereby reducing the interfacial trioleoylglycerol concentration available for enzyme catalysis. One explanation for the cholesteryl oleate-mediated enhancement of lipoprotein lipase activity at low trioleoylglycerol concentrations is that the additional spreading of cholesteryl oleate disrupts microemulsions of trioleoylglycerol, thereby increasing the effective monomer substrate concentration available for enzyme catalysis. Based on these monolayer studies with model systems, we suggest that the relative amount of cholesteryl esters in plasma triacylglycerol-rich lipoproteins plays a regulatory role in determining the rate at which triacylglycerols are cleared from the circulation.