6-acyl galactosyl ceramides of pig brain: structure and fatty acid composition

Two glycolipids were isolated from pig brain and were shown to be the fatty acid esters of kerasin and cerebron in which the second fatty acid moiety is attached to the 6-position of the galactose. The point of attachment was shown in two ways: by permethylation and by cleavage with periodate. Methanolysis of the permethylated cerebroside esters yielded O-methyl sphingosines, methyl esters of nonhydroxy or 2-methoxy acids, and methyl 2,3,4-trimethyl galactoside. Cleavage of the cerebron ester with periodate, followed by treatment with sodium borohydride and dilute HCl, yielded ceramide plus 1-monoglyceride. The ester-linked fatty acids were primarily 16:0, 18:0, and 18:1, while the amide-linked fatty acids showed the wide assortment of chain lengths typical of brain cerebrosides. The methylation step, with silver oxide and methyl iodide, yielded two derivatives with the cerebroside esters, but the structural explanation for the difference was not elucidated. The galactose in the cerebron ester was shown to exist in the beta-pyranoside form.     

Seeds of Trichosanthes kirilowii, an energy-rich diet

The kernels of Trichosanthes kirilowii seeds contain a green oil which makes up for 62% of their dry matter. This oil consists up to 95% of triglycerides, 2% of glycolipids, 1.3% of phospholipids and 1.8% of chlorophylls. As fatty acid components the triglycerides, glycolipids and phospholipids contain the unsaturated fatty acids linoleic and oleic acid and the saturated palmitic acid. In the triglycerides 19% of the C18:3 acid occur with the configuration delta9 cis, delta11 trans, delta13 cis. This acid is called trichosanic acid and is absent in glycolipids and phospholipids which contain instead another C18:3 fatty acid, which has conjugated double bounds and occurs with an amount of 21% and 3%, respectively. Typically, these oil seeds contain in addition up to 30% of their dry matter proteins and up to 2.5% mono- and oligosaccharides. The monosaccharides consist of rhamnose, galactose and glucose and the oligosaccharides represent a mixture of tri- and tetrasaccharides.     

Ornithine lipids and their structural modifications: from A to E and beyond

Ornithine lipids (OLs) are phosphorus-free membrane lipids that are widespread in eubacteria, but absent from archaea and eukaryotes. They contain a 3-hydroxy fatty acyl group attached in amide linkage to the α-amino group of the amino acid ornithine. A second fatty acyl group is ester-linked to the 3-hydroxy position of the first fatty acid. About 25% of the bacterial species whose genomes have been sequenced are predicted to have the capacity to form OLs. Distinct OL hydroxylations have been described in the ester-linked fatty acid, the amide-linked fatty acid, and the ornithine moiety. These modifications often seem to form part of a bacterial stress response to changing environmental conditions, allowing the bacteria to adjust membrane properties by simply modifying already existing membrane lipids without the need to synthesize new lipids.     

Structural characterization of the lipid A component of Sinorhizobium sp. NGR234 rough and smooth form lipopolysaccharide. Demonstration that the distal amide-linked acyloxyacyl residue containing the long chain fatty acid is conserved in rhizobium and Sinorhizobium sp

A broad-host-range endosymbiont, Sinorhizobium sp. NGR234 is a component of several legume-symbiont model systems; however, there is little structural information on the cell surface glycoconjugates. NGR234 cells in free-living culture produce a major rough lipopolysaccharide (LPS, lacking O-chain) and a minor smooth LPS (containing O-chain), and the structure of the lipid A components was investigated by chemical analyses, mass spectrometry, and NMR spectroscopy of the underivatized lipids A. The lipid A from rough LPS is heterogeneous and consists of six major bisphosphorylated species that differ in acylation. Pentaacyl species (52%) are acylated at positions 2, 3, 2', and 3', and tetraacyl species (46%) lack an acyl group at C-3 of the proximal glucosamine. In contrast to Rhizobium etli and Rhizobium leguminosarum, the NGR234 lipid A contains a bisphosphorylated beta-(1' --> 6)-glucosamine disaccharide, typical of enterobacterial lipid A. However, NGR234 lipid A retains the unusual acylation pattern of R. etli lipid A, including the presence of a distal, amide-linked acyloxyacyl residue containing a long chain fatty acid (LCFA) (e.g. 29-hydroxytriacontanoate) attached as the secondary fatty acid. As in R. etli, a 4-carbon fatty acid, beta-hydroxybutyrate, is esterified to (omega - 1) of the LCFA forming an acyloxyacyl residue at that location. The NGR234 lipid A lacks all other ester-linked acyloxyacyl residues and shows extensive heterogeneity of the amide-linked fatty acids. The N-acyl heterogeneity, including unsaturation, is localized mainly to the proximal glucosamine. The lipid A from smooth LPS contains unique triacyl species (20%) that lack ester-linked fatty acids but retain bisphosphorylation and the LCFA-acyloxyacyl moiety. The unusual structural features shared with R. etli/R. leguminosarum lipid A may be essential for symbiosis.     

Fatty acid acylation of proteins in Physarum polycephalum

We have investigated the occurrence of protein-fatty acid acylation by metabolic incorporation of [3H]myristic and [3H]palmitic acids in Physarum polycephalum. We show that this organism contains fatty acylated proteins with mainly myristic acid covalently attached in alkali-stable linkages, probably amides. We find no evidence for ester-linked fatty acids, in contrast to the situation in vertebrate cells.     

The testicular main ducts and the spermatic ducts in some cyprinid fishes—II. Composition of the seminal fluid

The composition of the organic compounds of the seminal fluid, pH values and osmolalities were investigated in three cyprinid species, the bleak ( Alburnus alburnus ), the chub ( Leuciscus cephalus ) and the zaehrte ( Vimba vimba ). The seminal fluid contains monosaccharides (glucose, fructose, galactose, xylose), lipids (cholesterol, fatty acids, phosphatidylcholine, glycolipids) and proteins, and exhibits activities of acid phosphatase, β-glucuronidase, proteases and to some extent of alkaline phosphatase. The composition of free amino acids reveals species specific differences.     

Isolation and characterization of novel neutral glycolipids from Thermoplasma acidophilum.

Several novel neutral glycolipids (GL-1a, GL-1b, GL-2a, GL-2b and GL-2c) were isolated from Thermoplasma acidophilum by high-performance liquid chromatography using phenylboronic acid-silica and preparative thin-layer chromatography. The tentative structures of these lipids were characterized by the combination of gas-liquid chromatography, the methylation procedure, and (1)H-NMR and FAB-mass spectrometries. The lipophilic portion of the neutral glycolipids was composed of a simple molecular species named caldarchaeol (dibiphytanyl-diglycerol tetraether). The sugar moieties of these glycolipids were composed of gulose and glucose which formed monosaccharide residues on one side or both sides of the core lipids. Gulose was attached to the terminal glycerol OH group of the core lipid with a beta-configuration and glucose being attached with an alpha-configuration. The proposed structure of GL-1a was gulosylcaldarchaeol and that of GL-1b was glucosylcaldarchaeol. The structures of GL-2a, GL-2b, and GL-2c were the analogs of the caldarchaeol derivatives attached by a variety of gulosyl residues or glucosyl residues on both sides of the terminal OH groups.     

Alterations in the molecular species of plasmalogen phospholipids and glycolipids due to peroxisomal dysfunction in Chinese hamster ovary-mutant Z65 cells by FABMS method

Changes in the molecular species of lipids associated with Pex2 gene-mutation were investigated to elucidate the pathogeneses of peroxisome biogenesis disorders. Although no differences were observed in the concentrations of cholesterol and phosphatidyl choline between mutated Z65 and control CHO-K1 cells, the amounts of cholesterol esters and glycolipids in Z65 cells were twice those in CHO-K1 cells, but phosphatidyl ethanolamine (PE), particularly 1-O-octadec-1'-enyl-2-oleoyl PE, was absent in Z65 cells by FABMS. Enhanced synthesis of glycolipids in Z65 cells was associated with an abundance of lignoceric acid-containing ones, suggesting a role of glycolipids in the retention of longer saturated fatty acids.     

Effect of electrical stimulation of the hypothalamus on plasma free fatty acid concentration in cats

The effect of electrical stimulation of various hypothalamic regions on levels of plasma free fatty acids, glucose, triglycerides, and cholesterol was studied in fasted cats. Appreciable changes were observed in plasma free fatty acids and glucose but not in plasma triglycerides or cholesterol. These changes appeared to be dependent upon small differences in the placement of electrodes and could not be related to a distinct hypothalamic locus. The results indicate that there is a dissociation between hypothalamic neurons that may affect plasma glucose concentration and those that may affect the plasma free fatty acids. It is suggested that the hypothalamus of the cat contains neurons that may influence autonomic discharge to adipose tissue and thus affect the plasma free fatty acid level and other neurons that may influence autonomic discharge to the liver and thus affect glucose output into the circulation. The distribution of both types of neurons is not limited to a distinct region of the hypothalamus in cats.     

Chemical composition of lipopolysaccharide from Azospirillum lipoferum

Abstract The lipopolysaccharide isolated from Azospirillum lipoferum strain SpBr17 (ATCC29709) was proven to be composed from 7 neutral sugars, two of which, rhamnose and glucose, were the major constituents. Two heptoses, l -glycero- d -mannoheptose and d -glycero- l -mannoheptose were identified. Among 8 fatty acids isolated from the lipopolysaccharide only 3-hydroxypalmitic acid was amide-bound. The approximate molar ratios of the constituents 3-deoxy- l -mannooctulosonic acid : glucosamine : amide-linked fatty acids : ester-linked fatty acids : phosphate were 0.8 : 4 : 2 : 4 : 2.5.     

The effect of variations in growth temperature, fatty acid composition and cholesterol content on the lipid polar head-group composition of Acholeplasma laidlawii B membranes

We have systematically investigated the effect of variations in growth temperature, fatty acid composition and cholesterol content on the membrane lipid polar headgroup composition of Acholeplasma laidlawii B. Two important lipid compositional parameters have been determined from such an analysis. The first parameter studied was the ratio of the two major neutral glycolipids of this organism, monoglucosyldiacylglycerol (MGDG) and diglucosyldiacylglycerol (DGDG). As the former lipid prefers to exist in a reversed hexagonal phase at higher temperatures, with unsaturated fatty acyl chains or in the presence of cholesterol, the ratio of these two lipids reflects the phase state preference of the total A. laidlawii membrane lipids. Although we find that the MGDG/DGDG ratio is reduced in response to an increase in fatty acid unsaturation, increases in growth temperature or cholesterol content reduce this ratio only in cells enriched in a saturated but not an unsaturated fatty acid. The second parameter studied was the ratio of these neutral glycolipids to the only phosphatide in the A. laidlawii membrane, phosphatidylglycerol (PG); this parameter reflects the relative balance of uncharged and charged lipids in the membrane of this organism. We find that the MGDG + DGDG/PG ratio is lowest in cells enriched in the saturated fatty acid even though these cells already have the highest lipid bilayer surface charge density. Moreover, this ratio is not consistently related to growth temperature or changes in cholesterol levels, as expected. We therefore conclude that A. laidlawii strain B, apparently unlike strain A, does not possess coherent regulatory mechanisms for maintaining either the phase preference or the surface charge density of its membrane lipid constant in response to variations in growth temperature, fatty acid composition or cholesterol content.     

Cyclic glycolipids from glandular trichome exudates of Cerastium glomeratum

Fourteen cyclic glycolipids, named glomerasides A–N, have been isolated from the glandular trichome exudate of Cerastium glomeratum (Caryophyllaceae). Their structures were determined by spectroscopic analysis of the glycolipids, as well as by application of the Ohrui–Akasaka method to the fatty acid methyl esters derived from the glycolipids and GCMS studies of trimethylsilyl ether derivatives of the methyl esters. The various glomerasides have a glycosidic linkage between the anomeric hydroxy group of the glucose and the C-11, C-10 or C-9 positions of the docosanoyl moiety. They also contained an ester linkage between the C-6 hydroxy group of the glucose ring and the carboxyl group of the oxygenated fatty acid to form their macrocyclic structures. The glucose moiety was optionally acetylated and/or malonylated at the C-2 or C-3 hydroxy groups. Among these compounds, the 1,6′-cyclic ester of 11(R)-(2-O-acetyl-β-d-glucopyranosyloxy)docosanoic acid (glomeraside D) was the most abundant (25%).     

The fatty acid and monosaccharide compositions of three neutral and three phosphorylated glycolipids isolated from Leishmania donovani promastigotes grown in a chemically defined medium

Several lipids and macromolecular lipoconjugates of Leishmania spp. have now been well characterized; however, the glycolipids of L. donovani have not been thoroughly examined. In the present study, 3 neutral and 3 phosphorylated glycolipids were detected in promastigote forms of the organism grown in a chemically defined medium. The fatty acid and sugar compositions of these glycolipids, isolated and purified by adsorption column chromatography and thin-layer chromatographic procedures, were identified and quantified by gas-liquid chromatography and mass spectrometry. Myristate (14:0), palmitate (16:0), palmitoleate (16:1), stearate (18:0), oleate (18:1), and linoleate (18:2) were the major fatty acids in all 6 glycolipids. Arabinose, mannose, glucose, and galactose were detected in the glycolipids. The biochemical nature of these lipids suggested that the major components in the isolated preparations of the 6 glycolipids are diacylglycerophospholipids, distinct from the major precursors of macromolecular lipoconjugates such as the lipid anchors of cell surface antigens that have been reported. These appear to be terminal products of lipid biosynthesis in this parasite.     

Glyceroglucolipids of the human saliva

Seven individual glycolipids (I--VII) have been isolated from the lipid extract of human saliva. All glycolipids contained glucose, glyceryl ethers and fatty acids, and differed from each other primarily with respect to the number of glucose residues. In addition, glycolipid V contained also the sulfate ester group. The structures of these glycolipids were identified by partial acid and alkaline hydrolysis, oxidation with periodate and chromium trioxide and methylation studies, as: Glc(alpha1 leads to 3)-diglyceride (glycolipid I), Glc(alpha1 leads to 6)Glc(alpha1 leads to 3)-diglyceride (glycolipids II and III), Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 3)-diglyceride (glycolipid IV), SO3H-6Glc(alpha1 leads to 6)Glc(alpha1 leads to 3)-diglyceride (glycolipid V), Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 3)-diglyceride (glycolipid VI) and Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 6)Glc(alpha1 lead to 6)Glc(alpha1 leads to 6)Glc(alpha1 leads to 3)-diglyceride (glycolipid VII). Diglyceride portion of these compounds consists of 1-O-alkyl-2-O-acyl-glycerol with the docosanoate and glyceryl-monodocosyl being the predominant acyl and alkyl components.     

Myristyl and palmityl acylation of the insulin receptor

The presence of covalently bound fatty acids in the insulin receptor has been explored in cultured human (IM-9) lymphocytes. Both alpha (Mr = 135,000) and beta (Mr = 95,000) subunits of the receptor incorporate [3H]myristic and [3H]palmitic acids in a covalent form. The effects of alkali and hydroxylamine on the labeled subunits indicate the existence of two different kinds of fatty acid linkage to the protein with chemical stabilities compatible with amide and ester bonds. The alpha subunit contains only amide-linked fatty acid while the beta subunit has both amide- and ester-linked fatty acids. Analysis by high performance liquid chromatography after acid hydrolysis of the [3H]myristate- and [3H]palmitate-labeled subunits demonstrates the fatty acid nature of the label. Furthermore, both [3H]myristic and [3H]palmitic acids are found attached to the receptor subunits regardless of which fatty acid was used for labeling. The incorporation of fatty acids into the insulin receptor is dependent on protein synthesis and is also detectable in the Mr = 190,000 proreceptor form. Fatty acylation is a newly identified post-translational modification of the insulin receptor which may have an important role in its interaction with the membrane and/or its biological function.     

Organization of skin stratum corneum extracellular lamellae: diffraction evidence for asymmetric distribution of cholesterol

Lipid suspensions containing 2:1:1 skin ceramides:palmitic acid:cholesterol, similar to the lipid composition found in the extracellular matrix of skin stratum corneum, were analyzed by X-ray diffraction methods. These suspensions gave a sharp wide-angle reflection at 4.1 A, indicating tight hydrocarbon chain packing that would function as a water barrier, and low-angle lamellar diffraction with a repeat period near 130 A, similar to that previously recorded from intact stratum corneum. The lamellar repeat increased from 121 A at pH 6 to 133 A at pH 8.5, allowing phase angles of the lamellar data to be obtained by a sampling theorem "swelling" analysis. Electron density profiles showed that each repeating unit contained two asymmetric bilayers, with a fluid space on one side of the bilayer that increased with increasing pH, due to electrostatic repulsion between bilayers because of ionization of the palmitic acid. Profiles obtained from lamellae with cholesterol sulfate partially substituted for cholesterol showed large density increases on that same side of the bilayer, indicating that cholesterol is asymmetrically distributed in each bilayer. A molecular model was developed postulating that this asymmetry is due to the exclusion of cholesterol from lipid monolayers containing the ester-linked unsaturated (linoleic) hydrocarbon chain of skin ceramide 1. This model can explain the altered organization of extracellular lamellae in epidermal cysts (P. W. Wertz, D. C. Swartzendruber, K. C. Madison, D. T. Downing. 1987. J. Invest. Dermatol. 89:419-425) where the ester-linked chains have a higher percentage of saturated fatty acids than found in normal epidermis.     

Partial characterization of glyceroglucolipids from human saliva

Glycolipids of human saliva have been isolated and partially characterized. The neutral glycolipids consisted of six compounds, composed of glucose, glyceryl ethers and fatty acids, and differed from each other primarily with respect to the number of glucose residues. The major acidic glycolipid contained glucose, glyceryl ethers, fatty acids and sulfate. Based on the data of chemical analyses we propose that the acidic glycolipid is a 1-0-alkyl-2-0-acylglycerol triglucoside sulfate and that the neutral glycolipids are mono-, di-, tri-, hexa- and octaglucoside derivatives of 1-0-alkyl-2-0-acylglycerol.     

Evidence that trans-bilayer interdigitation of glycosphingolipid long chain fatty acids may be a general phenomenon

'Interdigitation' is a term coined to describe the phenomenon whereby pure phosphatidylcholines with intramolecular fatty acid chain length heterogeneity when hydrated to form bilayers may insert the methyl ends of long fatty acids from one side across more than half of the membrane thickness to protrude amongst the acyl chains of the opposite side of the bilayer (Keough, K.M.W. and Davis, P.J. (1979) Biochemistry 18, 1453-1459; Huang, C. and Mason, J.T. (1986) Biochim. Biophys. Acta 864, 423-470). In this article we address the fate of long fatty acid chains of glycosphingolipids present as minor components in membranes of non-interdigitating phosphatidylcholines. In this pursuit, derivatives of galactosyl ceramide, lactosyl ceramide, globoside and GM1 were synthesized having either 18-carbon or 24-carbon fatty acid with a spin label covalently attached at C-16. Labelled glycolipids were incorporated at 1-2 mol% into bilayers of synthetic phosphatidylcholines, their mixtures with cholesterol, or natural egg phosphatidylcholine. In each case the C-16 carbon of the glycolipid long chain fatty acid showed considerably greater 'order' and immobility than did C-16 of the fatty acid which was similar in length to the host matrix phospholipids. We interpret this as strong evidence that the long chain fatty acid interdigitates across the mid point of the bilayer in the systems studied. Clearly this phenomenon did not require that the phospholipid host matrix have mixed chain lengths. Furthermore it was totally independent of glycolipid family: for a given host matrix and (glycolipid) fatty acid chain length the order parameter values found were the same amongst all four glycolipid families tested.     

Cholesterol lipids of Borrelia burgdorferi form lipid rafts and are required for the bactericidal activity of a complement-independent antibody

Borrelia burgdorferi, the agent of Lyme disease, is unusual as it contains free cholesterol and cholesterol glycolipids. It is also susceptible to complement-independent bactericidal antibodies, such as CB2, a monoclonal IgG1 against outer surface protein B (OspB). We find that the bactericidal action?of CB2 requires the presence of cholesterol glycolipids and cholesterol. Ultrastructural, biochemical, and biophysical analysis revealed that the bacterial?cholesterol glycolipids exist as lipid raft-like microdomains in the outer membrane of cultured and mouse-derived B. burgdorferi and in model membranes from B. burgdorferi lipids. The order and size of the microdomains are temperature sensitive and correlate with the bactericidal activity of CB2. This study demonstrates the existence of cholesterol-containing lipid raft-like microdomains in a prokaryote, and we suggest that the temperature dependence of B. burgdorferi lipid raft organization may have significant implications in the transmission cycle of the spirochetes which are exposed to a range of temperatures.     

Characterization of membrane components of the flask-shaped mycoplasma Mycoplasma mobile

The cell membrane of Mycoplasma mobile was isolated by either ultrasonic or French press treatment of intact cells. The membrane fraction contained all of the cellular lipids, but only one-third of cellular proteins and had a density of 1.14 g ml-1. The soluble fraction contained the NADH dehydrogenase activity of the cells, as well as a protein with an apparent molecular mass of 55 kDa that was phosphorylated in the presence of ATP. Lipid analyses of M. mobile membranes revealed that membrane lipid could be labelled by radioactive glycerol, oleate and to a much higher extent by palmitate but not by acetic acid. The membrane lipid fraction was composed of 54% neutral and 46% polar lipid. The major constituents of the neutral lipid fraction were free fatty acid, free cholesterol and cholesterol esters (45, 25 and 20%, respectively, of total neutral lipid fraction). The free cholesterol count was 13% (w/w) of total membrane lipids with a cholesterol:phospholipid molar ratio of about 0.9. Among the polar lipids, both phospho- and glycolipids were detected. The phospholipid fraction consisted of a major de novo-synthesized phosphatidylglycerol (approximately 63% of total phospholipids), plus exogenous phosphatidylcholine and sphingomyelin incorporated in an unchanged form from the growth medium. The glycolipid fraction was dominated by a single glycolipid (approximately 90% of total glycolipids) that was preferentially labelled by palmitic acid and showed a very high saturated:unsaturated fatty acids ratio.