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FEBS Letters Jan 1993Low density lipoproteins (LDL) as well as isolated apolipoprotein B (ApoB) have been shown to exhibit phospholipase A2 (PLA2) activity toward phospholipids containing an...
Low density lipoproteins (LDL) as well as isolated apolipoprotein B (ApoB) have been shown to exhibit phospholipase A2 (PLA2) activity toward phospholipids containing an oxidized or short fatty acyl chain at position 2. Some of these studies employed the fluorescent analogue of phosphatidyl choline (PC), C6-NBD-PC, containing NBD-caproic acid (C6-NBD-FA) at position 2 as a substrate, representative of short fatty acyl chains. The release of NBD-caproic acid from position 2 is attributed to PLA2-catalysed hydrolysis. However, this fatty acid can be released also by other enzymatic pathways. In the present study we examined, and ruled out, other enzymatic pathways which may be responsible for the hydrolysis of fatty acids from position 2 of phospholipids. On the other hand, we found that LDL as well as isolated ApoB hydrolyse C6-NBD-FA from both carbon 1 and carbon 2 of these phospholipids, thus exhibiting independent and simultaneous activities of phospholipase A1 and phospholipase A2.
Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Apolipoproteins B; Brain; Chromatography, Thin Layer; Humans; Hydrolysis; Phosphatidylcholines; Phospholipases A; Phospholipases A1; Phospholipases A2; Rats
PubMed: 8422916
DOI: 10.1016/0014-5793(93)81176-z -
Hepatology (Baltimore, Md.) Jun 2019
Topics: Animals; Hydrolysis; Lipase; Lipid Droplets; Mice; Phospholipases; Phospholipases A2, Calcium-Independent; Triglycerides
PubMed: 30901101
DOI: 10.1002/hep.30620 -
The Journal of Biological Chemistry Jun 2003Oxidant stress and phospholipase A2 (PLA2) activation have been implicated in numerous proinflammatory responses of the mesangial cell (MC). We investigated the...
Cross-talk between cytosolic phospholipase A2 alpha (cPLA2 alpha) and secretory phospholipase A2 (sPLA2) in hydrogen peroxide-induced arachidonic acid release in murine mesangial cells: sPLA2 regulates cPLA2 alpha activity that is responsible for arachidonic acid release.
Oxidant stress and phospholipase A2 (PLA2) activation have been implicated in numerous proinflammatory responses of the mesangial cell (MC). We investigated the cross-talk between group IValpha cytosolic PLA2 (cPLA2alpha) and secretory PLA2s (sPLA2s) during H2O2-induced arachidonic acid (AA) release using two types of murine MC: (i). MC+/+, which lack group IIa and V PLA2s, and (ii). MC-/-, which lack groups IIa, V, and IValpha PLA2s. H2O2-induced AA release was greater in MC+/+ compared with MC-/-. It has been argued that cPLA2alpha plays a regulatory role enhancing the activity of sPLA2s, which act on phospholipids to release fatty acid. Group IIa, V, or IValpha PLA2s were expressed in MC-/- or MC+/+ using recombinant adenovirus vectors. Expression of cPLA2alpha in H2O2-treated MC-/- increased AA release to a level approaching that of H2O2-treated MC+/+. Expression of either group IIa PLA2 or V PLA2 enhanced AA release in MC+/+ but had no effect on AA release in MC-/-. When sPLA2 and cPLA2alpha are both present, the effect of H2O2 is manifested by preferential release of AA compared with oleic acid. Inhibition of the ERK and protein kinase C signaling pathways with the MEK-1 inhibitor, U0126, and protein kinase C inhibitor, GF 1092030x, respectively, and chelating intracellular free calcium with 1,2-bis(2-aminophenoyl)ethane-N,N,N',N'-tetraacetic acid-AM, which also reduced ERK1/2 activation, significantly reduced H2O2-induced AA release in MC+/+ expressing either group IIa or V PLA2s. By contrast, H2O2-induced AA release was not enhanced when ERK1/2 was activated by infection of MC+/+ with constitutively active MEK1-DD. We conclude that the effect of group IIa and V PLA2s on H2O2-induced AA release is dependent upon the presence of cPLA2alpha and the activation of PKC and ERK1/2. Group IIa and V PLA2s are regulatory and cPLA2alpha is responsible for AA release.
Topics: Animals; Arachidonic Acid; Glomerular Mesangium; Group II Phospholipases A2; Group IV Phospholipases A2; Group V Phospholipases A2; Hydrogen Peroxide; Mice; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Oxidative Stress; Phospholipases A; Phospholipases A2; Receptor Cross-Talk
PubMed: 12676927
DOI: 10.1074/jbc.M300424200 -
Cell Reports Jan 2018Variants in the phospholipase D3 (PLD3) gene have genetically been linked to late-onset Alzheimer's disease. We present a detailed biochemical analysis of PLD3 and...
Variants in the phospholipase D3 (PLD3) gene have genetically been linked to late-onset Alzheimer's disease. We present a detailed biochemical analysis of PLD3 and reveal its endogenous localization in endosomes and lysosomes. PLD3 reaches lysosomes as a type II transmembrane protein via a (for mammalian cells) uncommon intracellular biosynthetic route that depends on the ESCRT (endosomal sorting complex required for transport) machinery. PLD3 is sorted into intraluminal vesicles of multivesicular endosomes, and ESCRT-dependent sorting correlates with ubiquitination. In multivesicular endosomes, PLD3 is subjected to proteolytic cleavage, yielding a stable glycosylated luminal polypeptide and a rapidly degraded N-terminal membrane-bound fragment. This pathway closely resembles the delivery route of carboxypeptidase S to the yeast vacuole. Our experiments reveal a biosynthetic route of PLD3 involving proteolytic processing and ESCRT-dependent sorting for its delivery to lysosomes in mammalian cells.
Topics: Humans; Lysosomes; Phospholipases
PubMed: 29386126
DOI: 10.1016/j.celrep.2017.12.100 -
Nature Microbiology Jan 2019Hydrolysis of phosphatidylcholine to choline was found to be catalysed by phospholipase D enzymes from diverse members of the gut microbiota, revealing a mechanism by...
Hydrolysis of phosphatidylcholine to choline was found to be catalysed by phospholipase D enzymes from diverse members of the gut microbiota, revealing a mechanism by which commensals obtain choline for subsequent production of disease-associated trimethylamine.
Topics: Choline; Gastrointestinal Microbiome; Phospholipase D; Phospholipases; Symbiosis
PubMed: 30546098
DOI: 10.1038/s41564-018-0325-1 -
The Journal of Biological Chemistry Nov 1980Deoxycholate treatment of horse platelets previously labeled in their phospholipids with [14C]arachidonate produces selective conversion of [14C]phosphatidylinositol...
Deoxycholate treatment of horse platelets previously labeled in their phospholipids with [14C]arachidonate produces selective conversion of [14C]phosphatidylinositol (PI) to [14C]1,2-diacylglycerol. This phospholipase C activity, which has a pH optimum of 7.5, is specific for phosphatidylinositol since other phospholipids or neutral lipids are not affected. Although exogenous Ca2+ is not required for activity, ethylene glycol bis(beta-aminoethyl ether)N,N,N',N-tetraacetic acid or EDTA abolishes phosphatidylinositol degradation. However, in the presence of added Ca2+, other phospholipids such as phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylserine (PS) are also degraded but by a phospholipase A2 activity. This activity generates the respective lyso-derivatives as well as various [14C]arachidonate metabolites. The phospholipase A2 activity is further enhanced by increasing the pH (7.5 to 9.5), a condition which severely suppresses the phospholipase C activity. Most of the platelet phospholipase A2 activity is associated with the particulate fractions of the cell, while the phospholipase C activity appears to be almost completely restricted to the soluble fraction. Deoxycholate treatment of the particulate fractions results in cleavage by phospholipase A2 of phosphatidylcholine and phosphatidylethanolamine but not of phosphatidylinositol. The preferred substrates for platelet phospholipase A2 appear to be phosphatidylethanolamine, phosphatidylcholine, and phosphatidylserine, while phosphatidylinositol seems to be degraded nearly exclusively by phospholipase C.
Topics: Animals; Arachidonic Acids; Blood Platelets; Calcium; Diglycerides; Horses; Hydrogen-Ion Concentration; Kinetics; Phospholipases; Phospholipases A; Phospholipases A2; Substrate Specificity; Type C Phospholipases
PubMed: 7430120
DOI: No ID Found -
Journal of Lipid Research Dec 1984Spectrophotometric techniques for determining the activities of lipases, lysophospholipases, and phospholipases are reviewed. These methods involve the use of thioester... (Comparative Study)
Comparative Study Review
Spectrophotometric techniques for determining the activities of lipases, lysophospholipases, and phospholipases are reviewed. These methods involve the use of thioester substrate analogs as well as omega-nitrophenyl derivatives of the corresponding lipids. The most promising results are obtained with the thioester substrate analogs. Mono- and diacylglycerol lipases are assayed by using rac-1-S-decanoyl-1-mercapto-2,3-propanediol and rac-1,2-S,O-didecanoyl-1-mercapto-2,3-propanediol, respectively. Phospholipases A1 and A2 are determined by using rac-1,2-S,O-didecanoyl-3-phosphocholine-1-mercapto-2,3-propanediol and 2-hexadecanoylthio-1-ethyl-phosphocholine, respectively. Lysophospholipases are measured by using 2-hexadecanoylthio-1-ethyl-phosphocholine. Phospholipase C is assayed with rac-1-S-phosphocholine-2,3-O-didecanoyl-1-mercapto-2,3-propanediol. Thioester substrate analog assay procedures are more rapid, sensitive, convenient, continuous, and less expensive than the classical radiochemical techniques.
Topics: Lipase; Lipoprotein Lipase; Lysophospholipase; Monoacylglycerol Lipases; Phospholipases; Phospholipases A; Spectrophotometry; Sulfides; Type C Phospholipases
PubMed: 6397560
DOI: No ID Found -
The Biochemical Journal Aug 1988A phospholipase A2, Laticauda colubrina phospholipase A2 II (LcPLA-II), and a phospholipase A2 homologue, Laticauda colubrina phospholipase A2 homologue I (LcPLH-I),...
Isolation, properties and amino acid sequences of a phospholipase A2 and its homologue without activity from the venom of a sea snake, Laticauda colubrina, from the Solomon Islands.
A phospholipase A2, Laticauda colubrina phospholipase A2 II (LcPLA-II), and a phospholipase A2 homologue, Laticauda colubrina phospholipase A2 homologue I (LcPLH-I), were isolated from the venom of the yellow-lipped sea snake, Laticauda colubrina, from the Solomon Islands. LcPLA-II showed phospholipase A2 activity towards egg-yolk phosphatidylcholine (24 mumol/min per mg at optimal conditions at 37 degrees C) and lethal potency (LD50 45 micrograms/kg body wt. intravenously in mice). Both of the activities were lost by treatment with p-bromophenacyl bromide. LcPLH-I showed neither phospholipase A2 activity nor lethal potency at a dose of 4.5 mg/kg body wt. in mice. It was not modified by the treatment with p-bromophenacyl bromide. LcPLA-II and LcPLH-I bound Ca2+ at a 1:1 molar ratio with KCa values of 105 microM and 44 microM at pH 8.0 respectively. Elucidation of the amino acid sequences of these two proteins showed that each protein consisted of a single chain of 118 amino acid residues, including 14 half-cystine residues. The two sequences are different from each other at 22 residues and highly homologous to those from other sources. The essential histidine residue for the phospholipase A2 activity at position 48 is replaced by an asparagine residue in the homologue LcPLH-I. Details of the separation of the peptides obtained by proteinase digestions of LcPLA-II and LcPLA-I and the determination of their amino acid sequences are given in Supplementary Publication SUP 50145 (14 pages), which has been deposited at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1988) 249, 5.
Topics: Acetophenones; Amino Acid Sequence; Animals; Chromatography, Gel; Elapid Venoms; Isoenzymes; Lethal Dose 50; Molecular Sequence Data; Peptide Fragments; Phospholipases; Phospholipases A; Phospholipases A2; Sequence Homology, Nucleic Acid
PubMed: 3178739
DOI: 10.1042/bj2530869 -
Journal of Clinical Microbiology Feb 1986The results of recent studies support the concept that Ureaplasma urealyticum may be a major cause of perinatal infection in both term and preterm infants. It has been...
The results of recent studies support the concept that Ureaplasma urealyticum may be a major cause of perinatal infection in both term and preterm infants. It has been postulated that phospholipase degradation of placental phospholipids by microorganisms triggers the onset of premature labor. Since the presence of ureaplasmas in placentas is associated with pregnancy loss, prematurity, and neonatal morbidity, we assayed U. urealyticum for the presence of phospholipase A1, A2, and C activities. Phospholipase A1 activity was low in lysates of exponential-phase cells of U. urealyticum. Phospholipase A2 activity was present and was 100-fold higher than the activity of phospholipase A1 in serotypes 3,4, and 8. The total activity and specific activity of phospholipase A2 in serotype 8 were nearly threefold higher than the activities in serotypes 3 and 4. Cell lysates of all three serotypes showed the presence of phospholipase C activity during the exponential phase of growth, and no significant difference in activity was observed among the three serotypes. In stationary-phase cells the phospholipase C activity was 10-fold lower than the activity in exponential-phase cells. Our results demonstrate that phospholipase activities are present in U. urealyticum cells and that the specific activities of phospholipase A2 differed among the three serotypes tested, while the activities of phospholipases A1 and C were similar.
Topics: Humans; Phospholipases; Phospholipases A; Phospholipases A1; Phospholipases A2; Phospholipids; Substrate Specificity; Type C Phospholipases; Ureaplasma
PubMed: 3700618
DOI: 10.1128/jcm.23.2.354-359.1986 -
Free Radical Biology & Medicine Jun 2013This article presents a radiometric assay to determine the enzymatic activity of platelet-activating factor (PAF) acetylhydrolase (PAF-AH), also known as...
This article presents a radiometric assay to determine the enzymatic activity of platelet-activating factor (PAF) acetylhydrolase (PAF-AH), also known as lipoprotein-associated phospholipase A2 and phospholipase A2 group 7A. The method is based on the release of radioactively labeled acetate from sn-2-labeled PAF and separation of substrate and product using reversed-phase column chromatography on octadecyl silica gel cartridges. The assay is fast, convenient, reproducible, sensitive, and inexpensive. The instrumentation required includes standard laboratory equipment and a liquid scintillation counter. The assay is also useful to determine the activity of intracellular PAF-AH (PAF-AH II), provided that a few modifications are included. The enzymatic activity determined using PAF as the substrate is a direct indication of the ability of plasma samples, purified preparations, and cellular and tissue lysates to hydrolyze short- and medium-chain phospholipids that may or may not harbor oxidized functionalities. In addition, the assay can be used to test the suitability of other phospholipids, including species containing oxidized, long-chain sn-2 fatty acyl groups, as PAF-AH substrates. This versatile assay can be used to accurately determine PAF-AH activity in biological samples and preliminarily assess affinity and efficiency of the hydrolysis of potential substrates present in complex mixtures.
Topics: 1-Alkyl-2-acetylglycerophosphocholine Esterase; Enzyme Activation; Humans; Phospholipases
PubMed: 22659315
DOI: 10.1016/j.freeradbiomed.2012.05.031