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At least 433 records · Page 24Linked to original sources
PubMed1987

Calcium-activated phospholipase C associated with canine renal basolateral membranes.

To determine whether calcium-activated phospholipase C effects breakdown of phospholipids present in the basolateral membrane of the renal proximal tubular cell we incubated proximal tubular basolateral membranes isolated from dog kidney in the absence and presence of deoxycholate, and varied free calcium from 0 to 300 microM. Following incubations, lipids were extracted from membranes and separated using thin-layer chromatography. In the absence of deoxycholate, neither phosphatidylinositol nor diglyceride extractable from basolateral membranes changed significantly as calcium was increased. In the presence of deoxycholate, extractable phosphatidylinositol and diglyceride did not change significantly as free calcium was elevated from 0 to 0.03 microM, but phosphatidylinositol decreased and diglyceride increased progressively as 0.03-300 microM free calcium was included in incubations. Most of the increased extractable diglyceride could be accounted for by hydrolysis of phosphatidylinositol as reflected by decreased extractable phosphatidylinositol. Our findings suggest that calcium-activated phospholipase C effects breakdown of phospholipids present in the basolateral portion of the plasma membrane of the renal proximal tubular cell. It is possible that this enzyme plays a role in stimulating protein kinase c at this site.

Animals↗
PubMed1981

Regulation of hepatic triglyceride synthesis in diabetic rats.

The syntheses of triglyceride and its precursors were increased when liver homogenates of ketotic diabetic rats were incubated with [U-14C]-glycero 3-phosphate and cofactors. Triolein sonicates produced a concentration-dependent inhibition of the synthesis of both diglyceride and triglyceride, whereas monoolein sonicates had no effect. Rat serum very low density lipoproteins, like triolein sonicates, inhibited the synthesis of diglyceride and triglyceride. Furthermore, the intracellular form of very low density lipoproteins, namely nascent very low density lipoproteins, also inhibited the synthesis of diglyceride and triglyceride. A higher apparent I50 (concentration of inhibitor that produces 50% inhibition of activity) was observed in liver homogenates of ketotic diabetic rats for inhibition of triglyceride or diglyceride synthesis by triolein sonicates, serum very low density lipoproteins, high density lipoproteins, and nascent very low density lipoproteins. Insulin treatment of the diabetic rats reversed the I50 values to control. In studies on the site of inhibition of triglyceride synthesis in the overall biosynthetic pathway, serum very low density lipoproteins produced a concentration-dependent inhibition of liver cytosolic phosphatidate phosphohydrolase activity. A higher I50 value was obtained with the hepatic enzyme of the diabetic rats. This higher I50 value was reversed to control by insulin treatment of the diabetic rats. These results indicated that the activity of this enzyme was less sensitive to inhibition by very low density lipoproteins in the ketotic diabetic state. The reduced sensitivity of phosphatidate phosphohydrolase activity to triglyceride inhibition observed in the present studies could explain our previous observation of an increased rate of triglyceride synthesis in ketotic diabetic liver homogenates.

Animals↗
PubMed1978

A spectrophotometric method for the assay of cytidine 5'-diphospho-1,2-diacyl-sn-glycerol-dependent enzymes of phospholipid metabolism.

Cytidine 5'-diphospho-1,2-diacyl-sn-glycerol (CDP-diglyceride) hydrolase, CDP-diglyceride:L-serine O-phosphatidyltransferase, and CDP-diglyceride:sn-glycero-3-phosphate phosphatidyltransferase all release CMP from their liponucleotide substrate, CDP-diglyceride. We have developed a spectrophotometric assay for these enzymes using CMP kinase, pyruvate kinase, and lactate dehydrogenase to couple the release of CMP with the oxidation of NADH. The assay for each of the phospholipid-dependent enzymes was found to be linear both with time and with enzyme concentration. The assay should prove useful for continuous monitoring of enzymatic activity, determination of initial rates of reaction, and detailed kinetic analysis of these enzymes. Since several enzymes and substrates are used in the coupled assay system, the method is limited to analysis of partially purified preparations lacking competing activities.

Bacillus↗
PubMed1994

Pancreatic lipase assays with triglycerides as substrate: contribution of each sequential reaction to product formation.

Human pancreatic lipase assays are usually performed in the presence of either emulsified triglycerides or diglycerides within the limits of their solubility. Two reactions are catalyzed in the presence of triglycerides: hydrolysis of triglycerides to diglycerides, and diglycerides to monoglycerides. The contribution of each reaction to the final result was determined after extensive kinetic studies on the appearance and/(or) accumulation of intermediates and/(or) products. Acylated glycerides were analyzed after extraction from the reaction mixture, separation of lipid classes by thin-layer chromatography, and quantification by capillary gas chromatography. The results show that after 10 min of reaction in the presence of high concentrations of triolein, 75% of the released fatty acids arise from the first reaction. Relative merits and disadvantages of each substrate (triglyceride or diglyceride) are discussed in terms of practicability.

Acylation↗
PubMed2005

Kinetic properties of turkey pancreatic lipase: a comparative study with emulsified tributyrin and monomolecular dicaprin.

Using the classical emulsified system and the monomolecular film technique, we compared several interfacial properties of turkey pancreatic lipase (TPL) and human pancreatic lipase (HPL). TPL, like HPL, presented the interfacial activation phenomenon when vinyl ester was used as substrate. In the absence of colipase and bile salts, using tributyrin emulsion or monomolecular films of dicaprin at low surface pressure, TPL, unlike HPL, hydrolyzes pure tributyrin emulsion as well as dicaprin films maintained at low surface pressures. TPL was also able to hydrolyze triolein emulsion in the absence of any additive and despite the accumulation of long-chain free fatty acids at the interface. The difference of behaviors between TPL and HPL can be explained by the penetration power of each enzyme. The enzyme that presents the maximal pi(c) (TPL) interacts more efficiently with interfaces, and it is not denaturated at high interfacial energy. Turkey pancreatic lipase is more active on rac-dicaprin than HPL; a maximal ratio of 9 was found between the catalytic activities of the two lipases measured at their surface pressure optima (20 mN m(-1)). A kinetic study on the surface pressure dependency, stereospecificity, and regioselectivity of TPL was performed using enantiopure diglyceride (1,2-sn-dicaprin and 2,3-sn-dicaprin) and a prochiral isomer (1,3-dicaprin) that were spread as monomolecular films at the air-water interface. At low surface pressure (15 mN m(-1)), TPL acts preferentially on primary carboxylic ester groups of the diglyceride isomers (1,3-dicaprin), but at high surface pressure (23 mN m(-1)), this enzyme prefers both adjacent ester groups of the diglyceride isomers (1,2-sn-dicaprin and 2,3-sn-dicaprin). HPL prefers adjacent ester groups of the diglyceride isomers (1,2-sn-dicaprin and 2,3-sn-dicaprin). Furthermore, TPL was found to be markedly stereospecific for the sn-1 position of the 1,2-sn-enantiomer of dicaprin at low surface pressure (15 mN m(-1)), while at high surface pressure (23 mN m(-1)), this lipase presents a stereopreference for the sn-3 position of the 2,3-sn-enantiomer of dicaprin. HPL is stereospecific for the sn-1 position of the 1,2-sn-enantiomer of dicaprin both at 15 and 23 mN m(-1).

Animals↗
PubMed1987

Phospholipid alterations in canine cardiac sarcoplasmic reticulum induced by an acid-active phospholipase C.

Phospholipid alterations and phospholipase activities were studied in a preparation of canine cardiac sarcoplasmic reticulum (SR) known to contain lysosomes. Incubation of SR at pH 5.0 (37 degrees C) resulted in a loss in total lipid phosphorus which was maximal (10%) by 30 min. whereas a modest increase in lipid phosphorus occurred during incubation at pH 7.0. The content of phospholipid phosphorus was decreased (8.6-19.0%) in sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine when SR was incubated at pH 5.0; however phosphatidic acid increased 14% relative to control. Lysophospholipids were not detected. Liposomes of 1-acyl 2-[1 14C]-linoleoyl-sn-glycero-3-phosphoryl-ethanolamine (14C-PE) were hydrolyzed by SR at pH 5.0 to yield radiolabelled diglyceride (18.1 nmol/hr mg protein) and free fatty acid (8.3 nmol/hr mg protein). SR-mediated diglyceride production at pH 5.0, but not free fatty acid release, was markedly potentiated when 14C-PE was peroxidized by preincubation at pH 5.0 for 3-24 hours at 37 degrees C. After 24 hours of preincubation at pH 5.0 the specific activity of SR-mediated diglyceride production was 188% of control. A comparable increase in diglyceride production occurred when 14C-PE was peroxidized at pH 7.0 and hydrolyzed by SR at pH 5.0. No increase in enzymatic activity occurred when liposomes were both preincubated and assayed at pH 7.0. When 14C-PE was exposed to air at pH 5.0 or 7.0 more polar radiolabelled derivatives were formed as determined by thin layer chromatography. Up to twice as much polar lipid was formed at pH 5.0 compared to pH 7.0.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1975

Phosphatidylglycerol in lung surfactant. II. Subcellular distribution and mechanism of biosynthesis in vitro.

Lamellar inclusion bodies, apparent precursors for alveolar surfactant lining, have remarkably similar phospholipid composition to surfactant from alveolar lavage, but distinctly different from other fractions studied: mitochondria, microsomal fraction containing endoplasmic reticulum membranes, plasma membranes and nuclei. Surfactant contained (as % of total phospholipid phosphate): 75.5-77.0% lecithin, 11.0-11.2% phosphatidylglycerol, 4.2-4.6% phosphatidylethanolamine, 3.0-3.2% phosphatidylinositol, 1.5-1.7% bis-(monoacylglycerol) phosphate, 1.2-1.9% phosphatidylserine, and 0.7-1.5% sphingomyelin. Fatty acids of phosphatidylglycerol from lamellar bodies were similar to those from microsomes but different from those in mitochondria. Lung homogenate in continuous sucrose density gradient displayed two major activity peaks of phosphatidylglycerol synthesis: the heavier from mitochondria; the lighter from endoplasmic reticulum. Studies on mechanism of phosphatidylglycerol synthesis in vitro revealed (in these two fractions) CDP-diglyceride and sn-glycerol phosphate precursors to phosphatidylglycerol phosphate, that hydrolysed to phosphatidylglycerol. In microsomes disaturated CDP-diglycerides were 1.6-1.9 times more active substrates than in mitochondria, whereas CDP-diglycerides from egg lecithin were almost equally active. In contrast to lung mitochondria no cardiolipin synthesis was detected in microsomes. The highest specific activities for phosphatidate cytidyltransferase, CDP-diglyceride-inositol phosphatidyltransferase, choline phosphotransferase, and phosphatidylethanolamine methyltransferase were all found in microsomes. The present in vitro studies and additional evidence (M. Hallman and L. Gluck, (1975) Fed. Proc. 34, 274) support the hypothesis that de novo synthesis of surfactant lecithin phosphatidylinositol and phosphatidylglycerol takes place in the endoplasmic reticulum of alveolar cells.

Animals↗
PubMed1978

The lipid composition of the non-photosynthetic diatom Nitzschia alba.

The lipid composition of the non-photosynthetic marine diatom, Nitzschia alba, has been quantitatively determined. Triglycerides accounted for 20% of the cell dry weight and 87% of the total lipids. Smaller amounts of 1,2- and 1,3-diglycerides, free sterol (24-methylene cholesterol), hydrocarbons and an unknown component were the remaining neutral lipids detected. Phosphatidylsulfocholine (phosphatidyl S,S-dimethylmercaptoethanol), present in amounts of 0.8% of cell dry weight (35% of total polar lipids), was the major polar lipid component. Other phospholipids were lysophosphatidylsulfocholine, phosphatidylglycerol, phosphatidylinositol and cardiolipin, but both phosphatidylcholine and phosphatidylethanolamine were completely absent. Another novel sulfolipid, deoxyceramide sulfonic acid, as well as the sulfate ester of the free sterol, were also present. Considerable amounts of the four lipids often associated with photosynthetic organisms, mono- and di-galactosyl diglycerides, sulfoquinovosyl diglyceride and phosphatidylglycerol, were identified in N. alba. However, the fatty acid components of the glycosyl diglycerides did not show the high amounts of polyunsaturated acids (18 : 2, 18 : 3) normally found in photosynthesizing organisms. All polar lipids were found to be associated with various cell membrane fractions in N. alba.

Cell Membrane↗
PubMed1976

Novel lipids of Butyrivibrio spp.

(1) An analysis has been conducted of the lipids present in three obligately anaerobic bacteria isolated from the ovine rumen belonging to the genus Butyrivibrio. Two of these organisms are rich in phospholipase (A1 + A2) activity, and appear to be different strains of the species fibrisolvens. (2) The only N-containing lipids comprise N-acyl-phosphatidylethanolamine occurring as a minor component in all organisms and a new lipid, diglyceride galactosylphosphorylethanolamine in one of these. (3) All three organisms contained the n-butyryl ester of phosphatidyl-glycerol and in one this represented the major phospholipid present. Valeryl, iso-valeryl, propionyl and myristoyl esters of phosphatidylglycerol were also detected. (4) Two organisms contained glycerylphosphorylgalactosyldiglyceride and one of these also contained a large proportion of a less polar galactophospholipid which is probably a diacyl derivative of the former lipid. (5) All three organisms contained monogalactofuranosyl diglyceride and from one a n-butyryl ester of this galactolipid was isolated. (6) In all of the lipids examined the "diglyceride' moiety consisted almost entirely of plasmalogenic diglyceride (alk-1-enyl, acyl, glycerol).

Animals↗
PubMed1986

Transmembrane electrical potential affects the lipid composition of Acholeplasma laidlawii.

In membranes of Acholeplasma laidlawii, lipid composition is regulated as a function of several stimuli affecting the volume and length of the hydrocarbon chains and the hydrocarbon-water interfacial area. This regulation is vizualized as changes in the relative amounts of the major polar lipids monoglucosyl diglyceride and diglucosyl diglyceride. These lipids form reversed hexagonal and lamellar phases with water, respectively. However, mixtures of the two lipids, in the molar proportions found in the A. laidlawii membrane, form a lamellar phase. By adjustment of the glycolipid ratio as a response to environmental stimuli, a certain stability of the lamellar membrane is maintained. In growing cells with oleoyl membrane lipids, a transmembrane electrical potential of approximately -50 mV (inside negative), but no transmembrane pH difference, was found. Addition of the K+ ionophore valinomycin caused a rapid and dose-dependent hyperpolarization remaining for at least 7 h. Simultaneously, a rapid and lasting metabolic decrease in the ratio monoglucosyl diglyceride/diglucosyl diglyceride occurred. The increase in potential and the decrease in the lipid ratio were both reversed in a dose-dependent manner by extracellular KCl. Likewise, the lipophilic cation tetraphenylphosphonium caused a dose-dependent decrease in membrane potential and an increase in the monoglucosyl diglyceride/diglucosyl diglyceride ratio, respectively. The ionophores monensin and particularly nigericin had similar but less pronounced effects on the potential and lipid ratios as valinomycin. The uncoupler carbonyl cyanide m-chlorophenylhydrazone had no effect on cell growth, membrane potential, or lipid regulation at 10 microM. These dissimilar structures and the low concentrations used make a direct disturbance of drug molecules on lipid packing in membranes less likely.(ABSTRACT TRUNCATED AT 250 WORDS)

Acholeplasma laidlawii↗
PubMed1968

The mechanism of thyrotrophin action in relation to lipid metabolism in thyroid tissue.

1. The effects of thyrotrophin in vitro on the incorporation of [(14)C]-glucose, -glycerol, -palmitate and -oleate into the lipids of thyroid tissue were examined. 2. Thyrotrophin increased the incorporation of these (14)C-labelled precursors into phosphatidylinositol specifically. 3. Thyrotrophin also increased the proportion of (14)C radioactivity from labelled glucose, glycerol, palmitate and oleate incorporated into the 1,2-diglycerides. 4. The addition of thyrotrophin to thyroid slices for 10min., after 2hr. of prelabelling with [(14)C]glycerol, also increased the proportion of (14)C radioactivity incorporated into the 1,2-diglyceride fraction. 5. After incubation of thyroid tissue with [1-(14)C]palmitate, thyrotrophin caused a two- to three-fold increase in the specific radioactivity of palmitate isolated from phosphatidylinositol and 1,2-diglycerides. In contrast, the specific radioactivity of palmitate isolated from the choline and ethanolamine phosphoglycerides, 1,3-diglycerides and triglycerides was not increased by thyrotrophin.

Animals↗
PubMed1976

Role of Galactolipids in Spinach Chloroplast Lamellar Membranes: II. Effects of Galactolipid Depletion on Phosphorylation and Electron Flow.

A galactolipid lipase from primary bean (Phaseolus vulgaris) leaves has been used to partially deplete spinach chloroplast inner membranes of their galactolipids. Chloroplasts treated with the lipase in the absence of bovine serum albumin lost 91% of their monogalactosyl diglyceride, 83% of their digalactosyl diglyceride, all of their phosphatidyl choline, but none of their sulfolipid. Electron microscopy of this sections revealed that the treated chloroplasts were greatly enlarged and lacked membrane stacking. Linolenic acid had similar effects on the structure of the chloroplasts. Chlorophyll, carotenoids, and coupling factor 1 remained bound to the treated membranes.To minimize the inhibition of phosphorylation and electron flow by fatty acids released by the lipase, bovine serum albumin (15-24 mg/ml) was added to the lipase incubation mixtures. Bovine serum albumin inhibited the extent, but not the initial rate, of fatty acid release by the lipase. Electron microscopy of chloroplasts treated with the lipase in the presence of bovine serum albumin showed that membrane stacking was partially maintained. Chloroplasts treated with lipase under these conditions retained about 30% of their monogalactosyl diglyceride, 50% of their digalactosyl diglyceride and phosphatidyl choline. The sulfolipid and phosphatidyl glycerol contents were unchanged. Electron flow through photosystems I and II with artificial electron donors and acceptors was not affected by lipase treatment in the presence of bovine serum albumin. In contrast, oxygen evolution and phosphorylation were partially inhibited. These reactions are also very sensitive to fatty acids and it is possible that the inhibition is the result of interaction of fatty acids with the membrane prior to their binding to bovine serum albumin.In view of the irreversible inactivation of electron flow and phosphorylation by fatty acids, it is difficult to assess the role of galactolipids in these processes when a specfic lipase is used to deplete the membrane.

Journal Article↗
PubMed1981

Lipid composition of chloroplast membranes from weed biotypes differentially sensitive to triazine herbicides.

Chloroplasts were isolated from triazine-sensitive and triazine-resistant biotypes of common groundsel (Senecio vulgaris L.), common lambsquarter (Chenopodium album L.), and redroot pigweed (Amaranthus retroflexus L.). Chloroplast lipids were extracted and analyzed for differences among sensitive and resistant biotypes. The distribution of lipid between major lipid classes differed in chloroplasts from resistant and susceptible biotypes. Chloroplasts from resistant biotypes contained higher proportions of monogalactosyl diglyceride and phosphatidyl ethanolamine and lower proportions of digalactosyl diglyceride and phosphatidyl choline than did chloroplasts from susceptible biotypes. Monogalactosyl diglyceride and phosphatidyl ethanolamine were also quantitatively higher in membranes of resistant versus susceptible biotypes. The major lipid classes of resistant chloroplast membranes contained lipids comparatively richer in unsaturated fatty acids with the exceptions of digalactosyl diglyceride from all three biotypes and phosphatidyl ethanolamine from common groundsel. Results correlated changes in triazine sensitivity with qualitative and quantitative differences in the lipid composition of chloroplast membranes.

Journal Article↗
PubMed1971

Lipid composition of some species of Arthrobacter.

The lipids from Arthrobacter crystallopoietes, A. pascens, and A. globiformis were investigated. Each strain contained three glycolipids, a monogalactosyl diglyceride, a digalactosyl diglyceride, and a dimannosyl diglyceride, and traces of triand tetraglycosyl diglycerides. The phospholipids in all three strains consisted of bisphosphatidylglycerol, phosphatidylglycerol, and phosphatidylmyoinositol. No evidence could be obtained for the occurrence of mannophosphoinositides. Analysis of the fatty acids by gas-liquid chromatography showed that they are predominantly C(15:0)anteiso and C(17:0)anteiso compounds. No significant differences were observed in the composition of lipids extracted from homogeneous cell preparations of the rod and sphere forms of A. crystallopoietes.

Arthrobacter↗
PubMed1972

Lipid composition of Mycoplasma neurolyticum.

The total lipid content of Mycoplasma neurolyticum comprises about 14% of the dry weight of the organisms and is about equally distributed between the phospholipid and the neutral-glycolipid fractions. The neutral lipids were identified as triglycerides, diglycerides, and cholesterol. The glycolipid fraction contained 1-O-beta-glucopyranosyl-d-2,3-diglyceride and 1-[O-beta-d-glycopyranosyl-(1-->6)-O-beta-d-glucopyranosyl]-d-2,3-diglyceride. The latter lipid is structurally identical to the diglucosyl diglyceride which occurs in Staphylococcus aureus. The phospholipids of the organism consist of a fully acylated glycerophosphoryl-glycerophosphoryl glycerol, phosphatidic acid, diphosphatidyl glycerol, phosphatidyl glycerol, and amino acyl esters of phosphatidyl glycerol. Phosphatidic acid and phosphatidyl glycerol account for greater than 90% of the phospholipids of organisms in the exponential phase of growth. The predominant fatty acids found in all of the acyl lipids were palmitic, stearic, and oleic acids.

Carbon Isotopes↗
PubMed1977

Comparative lipid composition of heterotrophically and autotrophically grown Sulfolobus acidocaldarius.

Complex lipids from the thermoacidophilic facultative autotroph Sulfolobus acidocaldarius, as well as a strictly autotrophic isolate, were compared between cells grown on yeast extract and elemental sulfur. Lipids from both organisms grown autotrophically were nearly identical. Each contained about 15% neutral lipids, 35% glycolipids, and 50% acidic lipids. Glycolipids and acidic lipids contained C40H82-76-derived glycerol ether residues. Major glycolipids included the glycerol ether analogues of glucosyl galactosyl diglyceride (5%) and glucosyl polyol diglyceride (75%). Acidic lipids were comprised mainly of the glycerol ether analogues of phosphatidyl inositol (7%), inositolphosphoryl glucosyl polyol diglyceride (72%), and a partially characterized sulfate- and phosphate-containing derivative of glucosyl polyol diglyceride (13%). The lipids from cells grown heterotrophically were similar to those from autotrophically grown cells, except that the partially characterized acidic lipid was absent. In addition, the two glycolipids as well as the respective inositolphosphoryl derivatives were each present in nearly equal proportions.

Chromatography, Gas↗
PubMedDate not supplied

[Comparative study of the lipid composition of seven species of "Micromonospora"].

Analysis of the lipids of seven species of Micromonospora showed the constant presence of five phospholipids and three glycolipids. The phospholipids were identified as cardiolipids, phosphatidylethanolamine, phosphatidyl-inositol, mannosides of phosphatidyl-inositol, and possibly lysocardiolipids. The glycolipids were monoglucosyl-diglycerides, diglucosyl-diglycerides (with a small amount of galactosyl-glucosyl-diglycerides) and esters of fatty acids and trehalose. The simultaneous presence of glucosyl-diglycerides and trehalose esters might be specific of the genus Micromonospora. In the seven species studied, a large amount of the total fatty acids (obtained by direct saponification of the cells) consisted of iso and anteiso fatty acids. Three groups could be distinguished, having respectively iso-C15, iso-C16 and n-C17 fatty acids as the main members. A monounsaturated C17 acid has been identified as heptadec-9,10-enoic acid.

Chemical Phenomena↗
PubMed1993

Phospholipase D hydrolysis of choline phosphoglycerides is selective for the alkyl-linked subclass of Madin-Darby canine kidney cells.

Madin-Darby canine kidney (MDCK) cells were used to study the synthesis of diglycerides from choline phospholipids (PC) in response to 12-O-tetradecanoylphorbol-13-acetate (TPA). In this system, diglyceride formation was blocked in the presence of ethanol (0.5%), and a corresponding amount of phosphatidylethanol (PEt) was formed, indicating that phospholipase D is responsible for the diglyceride production. Analysis of the subclasses of phosphatidylethanol revealed 1-O-alkyl-(alkyl), 1-O-alk-1'-enyl-(alkenyl), and 1-acyl species of PEt (38.0, 8.3, and 53.7%, respectively). The molecular species of the alkyl-PEt most closely matched the alkyl-PC. No change in the relative amounts of alkyl- versus acyl-PEt was observed with time after stimulation. Comparison of the alkyl content of PEt (38.0%) and the parent PC (15.2%) indicated a marked selectivity for the alkyl subclass of PC. A cell-free assay (Huang, C., Wykle, R. L., Daniel, L. W., and Cabot, M. C. (1992) J. Biol. Chem. 267, 16859-16865) for phospholipase D was also used to confirm the selectivity of the enzyme for alkyl-PC versus acyl-PC. The predominant molecular species of PEt contained saturated acyl or alkyl chains in position-1 and monounsaturated residues in position-2 accounting for approximately 50% of the total PEt. 1-O-Octadecyl-2-oleoyl-sn-glycerol, a representative alkyl molecular species, was synthesized and tested for its effect upon protein kinase C derived from MDCK cells. This alkyl-diglyceride (DG) neither stimulated protein kinase C nor inhibited its activation by diacylglycerol. In summary, TPA-stimulated phospholipase D is selective for the alkyl-PC subclass in MDCK cells. The alkyl-DG subsequently formed does not appear to function as a second-messenger in activating protein kinase C.

Alkylation↗