Metabolic interrelations of hydroxy-substituted ether-linked glycerolipids in the pink portion of the rabbit harderian gland.

The pink portion of the rabbit harderian gland is known to contain a preponderance of ether-linked glycerolipids consisting primarily of 1-(O-acyl)hydroxyalkyl-2,3-diacyl-sn-glycerols and smaller amounts of 1-alkyl-2,3-diacyl-sn-glycerols. In the present study, we have used a combination of chemical, enzymatic, and chromatographic techniques to identify two minor lipid components in the gland as 1-hydroxyalkyl-2-acyl-sn-glycerols and 1-hydroxyalkyl-2,3-diacyl-sn-glycerols. The long-chain acyl groups occurring in the 1-hydroxyalkyl-2-acyl-sn-glycerols and 1-hydroxyalkyl-2,3-diacyl-sn-glycerols are almost exclusively hexadecanoic acid, whereas the 1-(O-acyl)hydroxyalkyl-2,3-diacyl-sn-glycerols have a ratio of hexadecanoic acid to octadecanoic acid of $\text{2}\text{1}$. The 1-(O-acyl) hydroxyalkyl-2,3-diacyl-sn-glycerols and the 1-hydroxyalkyl-2,3-diacyl-sn-glycerols also contain a short-chain acyl moiety identified as 3-methylbutanoic acid (isovaleric acid). This acid was found to occupy the 3-position of the glycerol backbone in these lipid classes.Metabolic experiments demonstrate that 3-methylbutanoic acid in the lipids of the gland is derived from the catabolism of l-leucine. Pulse-chase data show a precursor-product relation between the 1-hydroxyalkyl-2,3-diacyl-sn-glycerols and 1-(O-acyl-hydroxyalkyl-2,3-diacyl-sn-glycerols and rule out direct hydroxylation of 1-alkyl-2,3-diacyl-sn-glycerols as a possible biosynthetic route to the 1-(O-acyl)hydroxyalkyl-2,3-diacyl-sn-glycerols.Characterization of the alkyl and acyl groups and the positional distributions of the acyl moieties in combination with the metabolic information indicated the acylation sequence involved in the formation of 1-(O-acyl)hydroxyalkyl-2,3-diacyl-sn-glycerol is 1-hydroxyalkyl-2-acyl-sn-glycerols → 1-hydroxyalkyl-2,3-diacyl-sn-glycerols → 1-(O-acyl)hydroxyalkyl-2,3-diacyl-sn-glycerols. The data also suggest that hydroxylation of the alkyl side-chain occurs before or at the alkylacylglycerol stage.     

Synthesis and chiroptical properties of neutral ether lipids

A variety of neutral ether lipids was synthesized. A method for the synthesis of 1,3-O-dialkyl-sn-glycerols was developed which involves selective alkylation of 3-O-alkyl-sn-glycerols. The ORD and CD curves of the various glyceryl ethers and their esters were analyzed. The correlation between the CD sign of the acyl residue and its position in the glycerol derivative was clarified.     

Characterization of the transacylase activity of rat liver 60-kDa lysophospholipase-transacylase. Acyl transfer from the sn-2 to the sn-1 position

Rat liver 60-kDa lysophospholipase-transacylase catalyzes not only the hydrolysis of 1-acyl-sn-glycero-3-phosphocholine, but also the transfer of its acyl chain to a second molecule of 1-acyl-sn-glycero-3-phosphocholine to form phosphatidylcholine (H. Sugimoto, S. Yamashita, J. Biol. Chem. 269 (1994) 6252–6258). Here we report the detailed characterization of the transacylase activity of the enzyme. The enzyme mediated three types of acyl transfer between donor and acceptor lipids, transferring acyl residues from: (1) the sn-1 to -1(3); (2) sn-1 to -2; and (3) sn-2 to -1 positions. In the sn-1 to -1(3) transfer, the sn-1 acyl residue of 1-acyl-sn-glycero-3-phosphocholine was transferred to the sn-1(3) positions of glycerol and 2-acyl-sn-glycerol, producing 1(3)-acyl-sn-glycerol and 1,2-diacyl-sn-glycerol, respectively. In the sn-1 to -2 transfer, the sn-1 acyl residue of 1-acyl-sn-glycero-3-phosphocholine was transferred to not only the sn-2 positions of 1-acyl-sn-glycero-3-phosphocholine, but also 1-acyl-sn-glycero-3-phosphoethanolamine, producing phosphatidylcholine and phosphatidylethanolamine, respectively. 1-Acyl-sn-glycero-3-phospho-myo-inositol and 1-acyl-sn-glycero-3-phosphoserine were much less effectively transacylated by the enzyme. In the sn-2 to -1 transfer, the sn-2 acyl residue of 2-acyl-sn-glycero-3-phosphocholine was transferred to the sn-1 position of 2-acyl-sn-glycero-3-phosphocholine and 2-acyl-sn-glycero-3-phosphoethanolamine, producing phosphatidylcholine and phosphatidylethanolamine, respectively. Consistently, the enzyme hydrolyzed the sn-2 acyl residue from 2-acyl-sn-glycero-3-phosphocholine. By the sn-2 to -1 transfer activity, arachidonic acid was transferred from the sn-2 position of donor lipids to the sn-1 position of acceptor lipids, thus producing 1-arachidonoyl phosphatidylcholine. When 2-arachidonoyl-sn-glycero-3-phosphocholine was used as the sole substrate, diarachidonoyl phosphatidylcholine was synthesized at a rate of 0.23 μmol/min/mg protein. Thus, 60-kDa lysophospholipase-transacylase may play a role in the synthesis of 1-arachidonoyl phosphatidylcholine needed for important cell functions, such as anandamide synthesis.     

The effect of cadmium ions and cadmium-metallothionein on the activities of phospholipid-synthesizing enzymes of rat liver microsomes in vitro

The effects of cadmium ions or cadmium-metallothionein on the activities of acyl-CoA:1acyl-sn-glycerol 3-phosphoric acid or 1-acyl-sn-glycero 3-phosphocholine acyltransferase of rat liver microsomes have been studied, in vitro. Cadmium ions were found to cause a noncompetitive type inhibition of these two acyltransferases. The K_i values were calculated, and found to be smallest (1.7 × 10^?5m) for palmitoyl-CoA and greatest (1.0 × 10^?4m) for linoleoyl-CoA, among the several fatty acyl-CoA's tested on the 1-acyl-sn-glycerol 3-phosphoric acid acyltransferases. With the 1-acyl-sn-glycero 3-phosphocholine acyltransferase, the K_i values were found to be smallest for the plamitoyl-CoA acyltransferase (3.8 × 10^?5m) and largest for thearachidonoyl-CoA acyltransferase (1.1 × 10^?4m). In contrast, mouse liver cadmium-metallothionein, including 4 mol of cadmium and 2 mol of zinc in one molecule of metallothionein, was not found to be inhibitory or rather stimulative on the above two acyltransferases at the same concentration of cadmium tested in the cadmium ion inhibitor experiments. The above results demonstrate that there is a strong and irreversible inhibition by cadmium ions on acyl-CoA acyltransferases, but that when cadmium acts on the enzyme in the form of a cadmium-metallothionein complex, the inhibition effect does not occur. These findings may reflect differing degrees of toxicity of these two types of cadmium compounds in mammalian tissues.     

Stereospecific distribution of plamitic acid in the triacylglycerols of rat adipocytes. Effects of varying the composition of the substrate fatty acid in vitro

The effects of inclusion of different fatty acids in the medium on the rate of esterification of palmitic acid and its stereospecific distribution among the three positions of the triacyl-sn-glycerols by preparations of rat adipocytes in vitro have been determined. Myristic acid, stearic acid, oleic acid and linoleic acid were used as diluents and the concentration of the combined unesterified fatty acids in the medium was held constant; only the proportion of palmitic acid was varied. The amount of palmitic acid esterified was always linearly related to its relative concentration in the medium and was not significantly affected by the nature of the diluent fatty acid chosen. Constant relative proportions were recovered in triacylglycerols and in intermediates in each instance. The amount of palmitic acid esterified to each of the positions of the triacyl-sn-glycerols was linearly dependent on the relative proportion in the medium but the nature of the relationship was markedly influenced by which fatty acid was present. When stearic acid was present, simple relationships were found over the whole range tested. When either myristic acid, oleic acid or linoleic acid was present, abrupt changes in the manner of esterification of palmitic acid were observed in position sn-1 when the relative concentrations of palmitic acid and the diluent reached critical values, which differed with each fatty acid. In position sn-2 when oleic acid or linoleic acid was present, a similar change was observed, and in position sn-3 it was obtained with myristic acid as diluent. The results are discussed in terms of changes in the relative affinities of the acyltransferases for palmitic acid. Palmitic acid was esterified into various molecular species in proportions that indicated acylation with non-correlative specificity at higher relative concentrations but not at lower.     

Lysophospholipase and lysophosphatidylcholine:Lysophosphatidylcholine transacylase from rat lung: Evidence for a single enzyme and some aspects of its specificity

An enzyme preparation was isolated from rat lung cytosol with the capability to transfer the fatty acyl chain from 1-acyl-sn-glycero-3-phosphocholine to water and to another molecule of 1-acyl-sn-glycero-3-phosphocholine. The evidence presented to indicate that a single protein confers both activities includes: (a) both normal and sodium dodecyl sulfate polyacrylamide disc gel electrophoresis showed a single protein band, and (b) heat treatment and preincubation with increasing amounts of diisopropylfluorophosphate resulted in concomitant loss of fatty acid and phosphatidylcholine formation. The enzyme converted 1-[9,10-³H₂]stearoyl-sn-glycero-3-phospho[¹⁴C-methyl]choline into phosphatidylcholine with an isotopic ³H/¹⁴C ratio twice that of the substrate, even when an excess of unlabeled fatty acid was present. The acyl group from palmitoyl-propanediol (1,3)-phosphocholine and palmitoyl-propanediol (1,3)-phosphoethanolamine could be transferred to lysophosphatidylcholine acceptor to yield phosphatidylcholine. Neither acylglycerols and cholesterol nor glycero-3-phosphate and glycero-3-phosphocholine served as acyl acceptors. Lysophosphatidylethanolamine and lysophosphatidyglycerol were converted also into the corresponding diacylphospholipids. Palmitoyllysophosphatidylcholine is preferentially converted into phosphatidylcholine when compared with stearoyllysophosphatidylcholine. The possible involvement of the enzyme in the synthesis of dipalmitoylphosphatidylcholine for the production of lung surfactant is discussed.     

Biochemical studies of the excitable membrane of Paramecium tetraurelia. II. Phospholipids of ciliary and other membranes

The phospholipids of cilia and deciliated bodies of Paramecium tetraurelia were isolated and characterized. 1-alkyl-2-acyl-sn-glycero-3-(2′-aminoethyl) phosphonate (GAEPL), phosphatidylethanolamine, and 1-alkyl-2-acyl-sn-glycero-3-phosphorylcholine (GPC) were the major lipids of Paramecium, and the minor lipids included phosphatidylinositol, cardiolipin, ceramide-(2-aminoethyl) phosphonate (CAEP), ceramide phosphorylethanolamine (COPE) and four sphingolipids whose identity was not established. The deciliated bodies contained 4% cardiolipin, 15% GAEPL, 41% phosphatidylethanolamine, 30% GPC and 3% each of CAEP and phosphatidylinositol; the cilia contained no cardiolipin, 24% GAEPL, 37% phosphatidylethanolamine, 15% GPC, 15% CAEP, 3% phosphatidylinositol, 2% COPE and small amounts (approx. 1%) of the four uncharacterized sphingolipids. No alteration in phospholipid composition was found among cells harvested in the various stages of growth. The phospholipids of six Paramecium mutants of three distinct phenotypes (pawn, paranoiac and fast) were also examined. Only one significant difference was found on comparison of the whole cell, deciliated body and cilia fraction of the mutants with the analogous fractions from wild type cells: the fast mutant, fA 97, had two extra, minor phospholipids (approx. 2%) in the deciliated body fraction that were tentatively identified as 1,2-diacyl-sn-glycero-3-(2′-aminoethyl) phosphonate (AEPL) and 1-alkyl-2-acyl-sn-glycero-3-phosphorylethanolamine (GPE).     

Isolation and characterisation of a maize cDNA that complements a 1-acyl sn-glycerol-3-phosphate acyltransferase mutant of Escherichia coli and encodes a protein which has similarities to other acyltransferases

We selected cDNA plasmid clones that corrected the temperature-sensitive phenotype of Escherichia coli strain JC201, which is deficient in 1-acyl-sn-glycerol-3-phosphate acyltransferase activity. A plasmid-based maize endosperm cDNA library was used for complementation and a plasmid that enabled the cells to grow at 44°C on ampicillin was isolated. Addition of this plasmid (pMAT1) to JC201 restored 1-acyl-sn-glycerol-3-phosphate acyltransferase activity to the cells. Total phospholipid labelling showed that the substrate for the enzyme, lysophosphatidic acid, accumulated in JC201 and was further metabolised to phosphatidylethanolamine in complemented cells. Membranes isolated from such cells were able to convert lysophosphatidic acid to phosphatidic acid in acyltransferase assays. The cDNA insert of pMAT1 contains one long open reading frame of 374 amino acids which encodes a protein of relative molecular weight 42 543. The sequence of this protein is most similar to SLC1, which is thought to be able to acylate glycerol at the sn-2 position during synthesis of inositol-containing lipids. Homologies between the SLC1 protein, the 1-acyl-sn-glycerol-3-phosphate acyltransferase of E. coli (PlsC) and the maize ORF were found with blocks of conserved amino acids, whose spacing was conserved between the three proteins, identifiable.     

Characterization of the Escherichia coli gene for 1-acyl-sn-glycerol-3-phosphate acyltransferase (pIsC)

Summary An Escherichia coli strain deficient in 1-acyl-sn-glycerol-3-phosphate acyltransferase activity has previously been isolated, and the gene (plsC) has been shown to map near min 65 on the chromosome. I precisely mapped the location of plsC on the chromosome, and determined its DNA sequence. plsC is located between parC and sufI, and is separated from sufI by 74 bp. Upstream of plsC is parC, separated by 233 bp, which includes an active promoter. parC, plsC, and sufI are all transcribed in the counterclockwise direction on the chromosome, possibly in an operon with multiple promoters. The amino-terminal sequence of the partially purified protein, combined with the DNA sequence, reveal 1-acyl-sn-glycerol-3-phosphate acyltransferase to be a 27.5 kDa highly basic protein. The plsC gene product, 1-acyl-sn-glycerol-3-phosphate acyltransferase, is localized to the cytoplasmic membrane of the cell. The amino-terminal sequence of the purified protein reveals the first amino acid to be a blocked methionine residue, most probably a formyl-methionine. The amino acid sequence of 1-acyl-sn-glycerol-3-phosphate acyltransferase has a short region of homology to two other E. coli acyltransferases that utilize acyl-acyl carrier protein as the acyl donor, sn-glycerol-3-phosphate acyltransferase and UDP-N-acetyl-glucosamine acyltransferase (involved in lipid A biosynthesis).     

Towards the genetic engineering of triacylglycerols of defined fatty acid composition: major changes in erucic acid content at the sn-2 position affected by the introduction of a 1-acyl-sn-glycerol-3-phosphate acyltransferase from Limnanthes douglasii into oil seed rape

A cDNA encoding a 1-acyl-sn-glycerol-3-phosphate acyltransferase from Limnanthes douglasii was introduced into oil seed rape (Brassica napus) under the control of a napin promoter. Seed triacylglycerols from transgenic plants were analysed by reversed-phase HPLC and trierucin was detected at a level of 0.4% and 2.8% in two transgenic plants but was not found in untransformed rape seed. Total fatty acid composition analysis of seeds from these selected plants revealed that the erucic acid content was no higher than the maximum found in the starting population. Analysis of fatty acids at the sn-2 position showed no erucic acid in untransformed rape but in the selected transgenic plants 9% (mol/mol) and 28.3% (mol/mol) erucic acid was present. These results conclusively demonstrate that the gene from L. douglasii encodes a 1-acyl-sn-glycerol-3-phosphate acyltransferase which can function in rape and incorporate erucic acid at the sn-2 position of triacylglycerols in seed. Additional modifications may further increase levels of trierucin.     

Formation of optically active ether lipids from race-mic-1-O-tetradecylglycerol in plant cell cultures

Photomixotrophic rape cells in culture specifically incorporate 1-O-tetradecyl-sn-glycerol from a racemic mixture into complex alkyl glycerolipids. Thus, both neutral and ionic 1-O- alkyl-2-O-acyl-sn-glycerolipids with defined alkyl moieties can be prepared from racemic mixtures of alkylglycerols.     

Bradykinin-stimulated release of [3H]arachidonic acid from phospholipids of HSDM1C1 cells: Comparison of diacyl phospholipids and plasmalogens as sources of prostaglandin precursors

Ethanolamine plasmalogens (1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamines) of many tissues contain high levels of arachidonate at their 2-position, and in certain tissues have been implicated as possible donors of arachidonate required in the synthesis of prostaglandins and thromboxanes. In the present study, [³H]arachidonate-labeled phospholipids of HSDM₁C₁ cells, a cell line derived from a mouse fibrosarcoma, were examined to determine the donor of the arachidonic acid released upon bradykinin stimulation of the synthesis of PGE₂. HSDM₁C₁ cells labeled with [³H]arachidonic acid for 24 hr in serum-free medium were used in most of the experiments and had the following distribution of label among the cellular lipids; phosphatidylcholine (33%), phosphatidylinositol (20%), diacyl-sn-glycero-3-phosphoethanolamine (15%), ethanolamine plasmalogen (15%), and less polar lipids (16%). Bradykinin treatment stimulated a rapid hydrolysis of [³H]arachidonate from the cellular lipids and conversion of the released acid to PGE₂, which was secreted into the medium. The label was released predominantly from phosphatidylinositol and possibly from phosphatidylcholine with no detectable change in the labeling of diacyl- or 1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamine. The ethanolamine plasmalogens, therefore, do not appear to be involved in the stimulated release of arachidonate in the HSDM₁C₁ cells. Indomethacin blocked the bradykinin-stimulated synthesis of PGE₂ and to a lesser degree inhibited the release of [³H]-arachidonate from the cellular lipids into the medium.     

Determination of enantiomeric purity of glycerides with a chiral PMR shift reagent.

Tris(3-heptafluorobutyryl-d-camphorato)europium(III), Eu(hfbc)₃ was used to determine the optical purities of enantiomeric mixtures of tri-, di- and monoglycerides with various fatty acid chain lengths by proton magnetic resonance (PMR). Synthesized model enantiomers were used to assign PMR signals. Enantiomeric signal separation becomes more difficult if the chain length difference between the fatty acids in the 1- and 3-positions of glycerol becomes smaller. The sign of the enantiomeric shift difference (ΔΔδ) of the terminal acyl CH₃ group of 1-acyl-2,3-distearoyl-sn-glycerol vs its enantiometer remains the same in the series acyl is hexanoyl, butyryl, propionyl, but is reversed for acetyl.The absolute configuration of the main triglyceride of the seed oil of Euonymus alatus was determined to be 3-acetyl-1,2-distearoyl-sn-glycerol and that of a monobutyryl triglyceride fraction from hydrogenated bovine butterfat was confirmed to be mainly 1,2-diacyl-3-butyryl-sn-glycerol. The enantiotopic behaviour of the glycerol CH₂ groups in (nearly) symmetric di- and triglycerides is discussed.     

Ether lipids

The naturally occurring 1-O-alkyl-sn-glycerols and their methoxylated congeners, 1-O-(2′-methoxyalkyl)-sn-glycerols, are biologically active compounds, ubiquitously found in nature as diacyl glyceryl ether lipids and phosphoether lipids. The chief objective of this article is to provide a comprehensive and up to date review on such ether lipids. The occurrence and distribution of these compounds in nature are extensively reviewed, their chemical structure and molecular variety, their biosynthesis and chemical synthesis and, finally, their various biological effects are described and discussed. An unprecedented biosynthesis of the 2′-methoxylated alkylglycerols is proposed. The first synthesis of enantiopure (Z)-(2′R)-1-O-(2′-methoxyhexadec-4′-enyl)-sn-glycerol, the most prevalent 2′-methoxylated type alkylglycerol present in cartilaginous fish, is described. It was accomplished by a highly convergent five step process.     

Syntheses and chiroptical properties of some derivatives of 1-thioglycerol

A modified synthesis of 3-thio-sn-glycerol which leads to a product of high optical purity is described. The purity was demonstrated by the use of a chiral shift reagent. 3-S-Acetyl- and 3-S-oleyl-3-thio-sn-glycerol as well as 3-acyl derivatives of 3-thio-sn-glycerol have been synthesized. The ORD and CD curves of these compounds as well as of some other derivatives of 3-thio-sn-glycerol were analyzed and discussed. The CD curves of the triacyl derivatives show a positive effect at 260 nm and a negative effect at 230–250 nm.     

High myristic acid content in the cyanobacterium Cyanothece sp. PCC 8801 results from substrate specificity of lysophosphatidic acid acyltransferase

Analysis of fatty acids from the cyanobacterium Cyanothece sp. PCC 8801 revealed that this species contained high levels of myristic acid (14:0) and linoleic acid in its glycerolipids, with minor contributions from palmitic acid (16:0), stearic acid, and oleic acid. The level of 14:0 relative to total fatty acids reached nearly 50%. This 14:0 fatty acid was esterified primarily to the sn-2 position of the glycerol moiety of glycerolipids. This characteristic is unique because, in most of the cyanobacterial strains, the sn-2 position is esterified exclusively with C16 fatty acids, generally 16:0. Transformation of Synechocystis sp. PCC 6803 with the PCC8801_1274 gene for lysophosphatidic acid acyltransferase (1-acyl-sn-glycerol-3-phosphate acyltransferase) from Cyanothece sp. PCC 8801 increased the level of 14:0 from 2% to 17% in total lipids and the increase in the 14:0 content was observed in all lipid classes. These findings suggest that the high content of 14:0 in Cyanothece sp. PCC 8801 might be a result of the high specificity of this acyltransferase toward the 14:0-acyl-carrier protein.     

Accumulation of neutral acylglycerols during the formation of morphologo-anatomical structure of euonymus Fruits

At three stages of fruit ripening of three euonymus species (Euonymus sp.), growth parameters, the development of morphologo-anatomical structure, and the accumulation of neutral acylglycerols (NAG) of fatty acids were studied. It was established that, in all species studied, in the cells of endosperm and cotyledons, as well as in arillus cells, small oleosomes of similar size were formed, whereas in Euonymus europaeus and E. maximowiczianus large oleosomes differing in their sizes also appeared. Independently of fruit age, dry weight of seeds exceeded that of arils by several times. At the determination of separate NAG classes: TAG (1,2,3-triacyl-sn-glycerols) and acDAG (3-acetyl-1,2-diacyl-sn-glycerols), it was shown that the absolute content of acDAG in seeds was 1.5 orders of magnitude higher than that of TAG; in contrast, in arils TAG were much more abundant than acDAG. Euonymus species differed markedly in the growth pattern and NAG accumulation in seeds and arils.     

Maintenance of respiratory control by beef heart mitochondria incubated at 25 degrees C: response to protective agents and to protective agents and to prior stress.

Lysophospholipase D (EC 3.1.4.-) activity was demonstrated in rat kidneys, intestines, lungs, testes, and liver. The liver enzyme was studied in greatest detail and its labeled products were identified by chemical and Chromatographic techniques. This enzyme hydrolyzes 1-[1-¹⁴C]hexadecyl-sn-glycero-3-phosphoethanolamine and 1-[1-¹⁴C]hexadecyl-sn-glycero-3-phosphocholine to yield 1-[1-¹⁴C]hexadecyl-sn-glycero-3-phosphate; the initial product is subsequently dephosphorylated by a phosphohydrolase in microsomes to form 1-[1-¹⁴C]hexadecyl-sn-glycerol. The possibility that phospholipase C and a phosphotransferase were responsible for the formation of 1-[1-¹⁴C]hexadecyl-sn-glycero-3-phosphate was ruled out. Neither 1-[1-¹⁴C]hexadecyl-2-acyl-sn-glycero-3-phosphoethanolamine nor 1-[1-¹⁴C]hexadecyl-2-acyl-sn-glycero-3-phosphocholine was hydrolyzed. The enzyme requires Mg²⁺, is inhibited by Ca²⁺, and is stimulated by high salt concentrations; it is localized in the microsomal fraction and has a pH optimum between 7.0 and 7.6. Inhibition by sulfhydryl reagents and protection by glutathione and dithiothreitol suggest that a sulfhydryl group is required for activity. The enzyme is inhibited by detergents and by organic solvent extraction. It appears to be tightly bound to the microsomes, since repeated freeze-thawing or sonication did not release the activity, and trypsin digestion (either in the presence or in the absence of 0.04% deoxycholate) did not destroy the activity. Lysophospholipase D was previously known to occur only in brain (R. L. Wykle and J. M. Schremmer, 1974, J. Biol. Chem., 249, 1742–1746).     

Alterations in cerebrospinal fluid glycerophospholipids and phospholipase A2 activity in Alzheimer's disease

Our aim is to study selected cerebrospinal fluid (CSF) glycerophospholipids (GP) that are important in brain pathophysiology. We recruited cognitively healthy (CH), minimally cognitively impaired (MCI), and late onset Alzheimer''s disease (LOAD) study participants and collected their CSF. After fractionation into nanometer particles (NP) and supernatant fluids (SF), we studied the lipid composition of these compartments. LC-MS/MS studies reveal that both CSF fractions from CH subjects have N-acyl phosphatidylethanolamine, 1-radyl-2-acyl-sn-glycerophosphoethanolamine (PE), 1-radyl-2-acyl-sn-glycerophosphocholine (PC), 1,2-diacyl-sn-glycerophosphoserine (PS), platelet-activating factor-like lipids, and lysophosphatidylcholine (LPC). In the NP fraction, GPs are enriched with a mixture of saturated, monounsaturated, and polyunsaturated fatty acid species, while PE and PS in the SF fractions are enriched with PUFA-containing molecular species. PC, PE, and PS levels in CSF fractions decrease progressively in participants from CH to MCI, and then to LOAD. Whereas most PC species decrease equally in LOAD, plasmalogen species account for most of the decrease in PE. A significant increase in the LPC-to-PC ratio and PLA₂ activity accompanies the GP decrease in LOAD. These studies reveal that CSF supernatant fluid and nanometer particles have different GP composition, and that PLA₂ activity accounts for altered GPs in these fractions as neurodegeneration progresses.     

Acyltransferases and the biosynthesis of pulmonary surfactant lipid in adenoma alveolar type II cells.

Acyltransferases are present in microsomes from alveolar type II cell adenomas (produced by urethan injections) that transfer palmitic acid in the presence of CoA, ATP, and Mg⁺⁺ to $\text{sn}$-glycerol-3-P to form phosphatidic acid, to dihydroxyacetone-P to form acyldihydroxyacetone-P, and to 1-acyl-$\text{sn}$-glycero-3-phosphocholine to form 3-$\text{sn}$-phosphatidylcholine. The data clearly demonstrate that the microsomal preparations can catalyze significant incorporation of palmitic acid into the 2-position of the disaturated species of 3-sn-phosphatidylcholine independently of phosphatidic acid formation as evidenced by the fact that $\text{sn}$-glycerol-3-P and calcium ions (which inhibit choline phosphotransferase) did not influence the incorporation of palmitic acid into the main surfactant lipid. Thus, a deacylation-acylation reaction involving 2-lysophosphatidylcholine appears to be an important pathway for the synthesis of surfactant lipid in alveolar type II cells; the control of acyl specificity at the 2-position is determined by the relative concentrations of the coparticipating substrates, l-palmitoyl-$\text{sn}$-glycero-3-phosphocholine and palmitoyl-CoA.