Improvement of transphosphatidylation reaction model of phospholipase D from Streptoverticillium cinnamoneum

2002 ◽  
Vol 10 (2) ◽  
pp. 115-121 ◽  
Author(s):  
Chiaki Ogino ◽  
Yuuichi Yasuda ◽  
Akihiko Kondo ◽  
Nobuaki Shimizu ◽  
Hideki Fukuda
Keyword(s):  
Phospholipase D ◽  
Reaction Model ◽  
Transphosphatidylation Reaction

scholarly journals A real-time, click chemistry imaging approach reveals stimulus-specific subcellular locations of phospholipase D activity

2019 ◽  
Vol 116 (31) ◽  
pp. 15453-15462 ◽  
Author(s):  
Dongjun Liang ◽  
Kane Wu ◽  
Reika Tei ◽  
Timothy W. Bumpus ◽  
Johnny Ye ◽  
...  
Keyword(s):  
Real Time ◽  
Phospholipase D ◽  
Second Messengers ◽  
Primary Alcohols ◽  
Fluorescent Reporter ◽  
Lipid Trafficking ◽  
Tyrosine Kinase Signaling ◽  
Inverse Electron Demand ◽  
Transphosphatidylation Reaction ◽  
Phospholipid Transport

The fidelity of signal transduction requires spatiotemporal control of the production of signaling agents. Phosphatidic acid (PA) is a pleiotropic lipid second messenger whose modes of action differ based on upstream stimulus, biosynthetic source, and site of production. How cells regulate the local production of PA to effect diverse signaling outcomes remains elusive. Unlike other second messengers, sites of PA biosynthesis cannot be accurately visualized with subcellular precision. Here, we describe a rapid, chemoenzymatic approach for imaging physiological PA production by phospholipase D (PLD) enzymes. Our method capitalizes on the remarkable discovery that bulky, hydrophilic trans-cyclooctene–containing primary alcohols can supplant water as the nucleophile in the PLD active site in a transphosphatidylation reaction of PLD’s lipid substrate, phosphatidylcholine. The resultant trans-cyclooctene–containing lipids are tagged with a fluorogenic tetrazine reagent via a no-rinse, inverse electron-demand Diels–Alder (IEDDA) reaction, enabling their immediate visualization by confocal microscopy in real time. Strikingly, the fluorescent reporter lipids initially produced at the plasma membrane (PM) induced by phorbol ester stimulation of PLD were rapidly internalized via apparent nonvesicular pathways rather than endocytosis, suggesting applications of this activity-based imaging toolset for probing mechanisms of intracellular phospholipid transport. By instead focusing on the initial 10 s of the IEDDA reaction, we precisely pinpointed the subcellular locations of endogenous PLD activity as elicited by physiological agonists of G protein-coupled receptor and receptor tyrosine kinase signaling. These tools hold promise to shed light on both lipid trafficking pathways and physiological and pathological effects of localized PLD signaling.

scholarly journals Role of Phospholipase D in Pasteurella haemolytica Leukotoxin-Induced Increase in Phospholipase A2 Activity in Bovine Neutrophils

1999 ◽  
Vol 67 (8) ◽  
pp. 3768-3772 ◽  
Author(s):  
Zuncai Wang ◽  
Cyril R. Clarke ◽  
Kenneth D. Clinkenbeard
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Pasteurella Haemolytica ◽  
Time Dependent ◽  
Regulatory Interaction ◽  
Pneumonic Pasteurellosis ◽  
Transphosphatidylation Reaction ◽  
Bovine Neutrophils ◽  
Phospholipase A2 Activity

ABSTRACT The effects of Pasteurella haemolytica leukotoxin (LKT) on the activity of phospholipase D (PLD) and the regulatory interaction between PLD and phospholipase A2 (PLA2) were investigated in assays using isolated bovine neutrophils labeled with tritiated phospholipid substrates of the two enzymes. Exposure of [3H]lysophosphatidylcholine-labeled neutrophils to LKT caused concentration- and time-dependent production of phosphatidic acid (PA), the product of PLD. LKT-induced generation of PA was dependent on extracellular calcium. Both production of PA and metabolism of [3H]-arachidonate ([3H]AA)-labeled phospholipids by PLA2 were inhibited when ethanol was used to promote the alternative PLD-mediated transphosphatidylation reaction, resulting in the production of phosphatidylethanol rather than PA. The role of PA in regulation of PLA2 activity was then confirmed by means of an add-back experiment, whereby addition of PA in the presence of ethanol restored PLA2-mediated release of radioactivity from neutrophil membranes. Considering the involvement of chemotactic phospholipase products in the pathogenesis of pneumonic pasteurellosis, development and use of anti-inflammatory agents that inhibit LKT-induced activation of PLD and PLA2 may improve therapeutic management of the disease.

Diacylglycerol generation in fluoride-treated neutrophils: involvement of phospholipase D

Blood ◽  
1991 ◽  
Vol 77 (12) ◽  
pp. 2746-2756
Author(s):  
D English ◽  
G Taylor ◽  
JG Garcia
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Superoxide Anion ◽  
Second Messenger ◽  
Complete Inhibition ◽  
Fluoride Ion ◽  
Extracellular Medium ◽  
Time Points ◽  
Acid Generation ◽  
Transphosphatidylation Reaction

Neutrophils exposed to fluoride ion (F-) respond with a delayed and sustained burst of superoxide anion release that is both preceded by and dependent on the influx of Ca2+ from the extracellular medium. The results of this study demonstrate a similarly delayed and sustained generation of 1,2-diglyceride in F(-)-treated neutrophils, over 90% of which was 1,2-diacylglycerol. Diacylglycerol generation was not dependent on the presence of extracellular Ca2+. Conversely, in contrast to results obtained with other agonists, removal of extracellular Ca2+ markedly potentiated synthesis of diacylglycerol in F(-)-treated neutrophils. This effect was accompanied by a corresponding decrease in the recovery of phosphatidic acid. In either the presence or absence of extracellular Ca2+, phosphatidic acid accumulated before diacylglycerol in F(-)-treated cells, suggesting the latter was derived from the former. Consistent with this hypothesis, the phosphatidic acid phosphohydrolase inhibitor, propranolol, suppressed generation of diacylglycerol as it potentiated the accumulation of phosphatidic acid in F(-)-treated neutrophils. This effect was observed both in the presence and absence of extracellular Ca2+. Moreover, high levels of propranolol (160 mumol/L) effected complete inhibition of diacylglycerol generation in F(-)-treated neutrophils with a corresponding increase in phosphatidic acid generation. Phosphatidylethanol accumulated in neutrophils stimulated with F- in the presence of ethanol. The extent of phosphatidylethanol accumulation at all time points after addition of F- corresponded to decreased levels of both phosphatidic acid and diacylglycerol, indicating that phosphatidylethanol was derived from the phospholipase D-catalysed transphosphatidylation reaction. The results indicate that F- activates a Ca(2+)-independent phospholipase D, which appears to be the major, if not sole, catalyst for both phosphatidic acid and diacylglycerol generation in F(-)-treated neutrophils. Ca2+, mobilized as a result of F- stimulation and possibly as a consequence of phospholipase D activation, exerts a profound effect on cellular second messenger levels by modulating the conversion of phosphatidic acid to diacylglycerol.

scholarly journals Polar Head Modified Phospholipids by Phospholipase D-Catalyzed Transformations of Natural Phosphatidylcholine for Targeted Applications: An Overview

Catalysts ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 997
Author(s):  
Chiara Allegretti ◽  
Francesca Denuccio ◽  
Letizia Rossato ◽  
Paola D’Arrigo
Keyword(s):  
Phospholipase D ◽  
Chemical Properties ◽  
Physical And Chemical Properties ◽  
Reaction Conditions ◽  
Chemical Methods ◽  
Polar Head ◽  
Transphosphatidylation Reaction ◽  
Physical And Chemical ◽  
Hydrolysis Of ◽  
And Chemical Properties

This review describes the use of phospholipase D (PLD) to perform the transphosphatidylation of the most common natural phospholipid (PL), phosphatidylcholine (PC) to obtain polar head modified phospholipids with real targeted applications. The introduction of different polar heads with distinctive physical and chemical properties such as charge, polarity and dimensions allows the obtainment of very different PLs, which can be exploited in very diverse fields of application. Moreover, the inclusions of a bioactive moiety in the PL polar head constitutes a powerful tool for the stabilization and administration of active ingredients. The use of this biocatalytic approach allows the preparation of compounds which cannot be easily obtained by classical chemical methods, by using mild and green reaction conditions. PLD is a very versatile enzyme, able to catalyze both the hydrolysis of PC to choline and phosphatidic acid (PA), and the transphosphatidylation reaction in the presence of an appropriate alcohol. The yield of production of the desired product and the ratio with the collateral PA formation is highly dependent on parameters such as the nature and concentration of the alcohol and the enzymatic source. The application of PLD catalyzed transformations for the production of a great number of PLs with important uses in medical, nutraceutical and cosmetic sectors will be discussed in this work.

Cholinergic activation of phospholipase D in lacrimal gland acini is independent of protein kinase C and calcium

1995 ◽  
Vol 268 (3) ◽  
pp. C713-C720 ◽  
Author(s):  
D. Zoukhri ◽  
D. A. Dartt
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase D ◽  
Lacrimal Gland ◽  
Phorbol Esters ◽  
Adrenergic Agonist ◽  
Kinase C ◽  
Cellular Lipids ◽  
Transphosphatidylation Reaction ◽  
Cholinergic Activation

To determine if rat lacrimal gland acini contain phospholipase D (PLD) activity, we took advantage of PLD's unique ability, in the presence of ethanol, to catalyze a transphosphatidylation reaction to produce phosphatidylethanol (PEth). Lacrimal gland acini were labeled for 3 h with [14C]stearic acid, preincubated for 20 min in the presence of 2% ethanol, and incubated for 20 min with or without agonists. Total cellular lipids were then extracted and analyzed by thin-layer chromatography, and the radioactivity was determined by liquid scintillation counting. Carbachol (1 mM), a cholinergic agonist, stimulated the production of both [14C]PEth and [14C]phosphatidic acid ([14C]PA) twofold. This effect was completely blocked by the muscarinic antagonist atropine (10 microM). [14C]PEth accumulation was also stimulated twofold by the active phorbol esters, 4 beta-phorbol 12-myristate 13-acetate and 4 beta-phorbol 12,13-dibutyrate at 1 microM. Ionomycin (1 microM), a Ca2+ ionophore, also stimulated the production of [14C]PEth twofold. In contrast to carbachol, neither phorbol esters nor ionomycin stimulated [14C]PA production. Neither [14C]PEth nor [14C]PA production was altered by epinephrine (1 mM), a nonselective adrenergic agonist, or phenylephrine (0.1 and 1 mM), a specific alpha 1-adrenergic agonist. We concluded that PLD activity, modulated by muscarinic receptors, protein kinase C, and Ca2+, but not by adrenergic receptors, is present in rat lacrimal gland acini. We also concluded that cholinergic activation of PLD appears to be independent of PKC and Ca2+.

Stimulation of phospholipase D activity and phosphatidic acid production by norepinephrine in rat aorta

1993 ◽  
Vol 264 (3) ◽  
pp. C609-C616 ◽  
Author(s):  
A. W. Jones ◽  
S. D. Shukla ◽  
B. B. Geisbuhler
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Time Course ◽  
Rat Aorta ◽  
Functional Responses ◽  
Phosphatidylinositol Bisphosphate ◽  
Phosphatidylinositol Phosphate ◽  
Transphosphatidylation Reaction ◽  
Hydrolysis Of ◽  
Stimulation Of

We sought to relate norepinephrine (NE) stimulation of phosphatidic acid (PA) production to functional responses of rat aorta and pathways for PA production. The time course for changes in PA was closely related to Ca-dependent tonic responses in 42K efflux and contraction. NE (30 microM for 1 min) increased PA and reduced phosphatidylcholine (PC) and phosphatidylinositol (PI) based on Pi analyses and 32P labeling of phospholipids. The 32P-to-Pi ratio in PA (0.8 +/- 0.2, n = 13) was similar to PC (0.8 +/- 0.1, n = 14) but was significantly lower (P < 0.001) than PI (4.6 +/- 0.5, n = 14). The 32P-to-Pi ratio in PA was also lower (P < 0.02) than phosphatidylinositol phosphate and phosphatidylinositol bisphosphate. NE also increased [3H]PA twofold (P < 0.05) when PC was selectively labeled with [3H]myristic acid. These observations are more consistent with PA being formed from the hydrolysis of PC by phospholipase D (PLD) than by the phosphorylation of diacylglycerol produced by the action of phospholipase C. PLD was assayed by the formation of phosphatidylethanol (PEt) via a transphosphatidylation reaction with ethanol (half-maximal stimulation at 0.4-0.5% vol/vol). The time course for PLD stimulation by NE was similar to PA, with significant increases (P < 0.002) during 10 s to 30 min exposure. Once formed, PEt was degraded slowly, with a half time > 3 h. It is concluded that NE stimulates PLD in rat aorta, which forms a significant amount of PA from the hydrolysis of PC.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Diacylglycerol generation in fluoride-treated neutrophils: involvement of phospholipase D

Blood ◽  
1991 ◽  
Vol 77 (12) ◽  
pp. 2746-2756 ◽  
Author(s):  
D English ◽  
G Taylor ◽  
JG Garcia
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Superoxide Anion ◽  
Second Messenger ◽  
Complete Inhibition ◽  
Fluoride Ion ◽  
Extracellular Medium ◽  
Time Points ◽  
Acid Generation ◽  
Transphosphatidylation Reaction

Abstract Neutrophils exposed to fluoride ion (F-) respond with a delayed and sustained burst of superoxide anion release that is both preceded by and dependent on the influx of Ca2+ from the extracellular medium. The results of this study demonstrate a similarly delayed and sustained generation of 1,2-diglyceride in F(-)-treated neutrophils, over 90% of which was 1,2-diacylglycerol. Diacylglycerol generation was not dependent on the presence of extracellular Ca2+. Conversely, in contrast to results obtained with other agonists, removal of extracellular Ca2+ markedly potentiated synthesis of diacylglycerol in F(-)-treated neutrophils. This effect was accompanied by a corresponding decrease in the recovery of phosphatidic acid. In either the presence or absence of extracellular Ca2+, phosphatidic acid accumulated before diacylglycerol in F(-)-treated cells, suggesting the latter was derived from the former. Consistent with this hypothesis, the phosphatidic acid phosphohydrolase inhibitor, propranolol, suppressed generation of diacylglycerol as it potentiated the accumulation of phosphatidic acid in F(-)-treated neutrophils. This effect was observed both in the presence and absence of extracellular Ca2+. Moreover, high levels of propranolol (160 mumol/L) effected complete inhibition of diacylglycerol generation in F(-)-treated neutrophils with a corresponding increase in phosphatidic acid generation. Phosphatidylethanol accumulated in neutrophils stimulated with F- in the presence of ethanol. The extent of phosphatidylethanol accumulation at all time points after addition of F- corresponded to decreased levels of both phosphatidic acid and diacylglycerol, indicating that phosphatidylethanol was derived from the phospholipase D-catalysed transphosphatidylation reaction. The results indicate that F- activates a Ca(2+)-independent phospholipase D, which appears to be the major, if not sole, catalyst for both phosphatidic acid and diacylglycerol generation in F(-)-treated neutrophils. Ca2+, mobilized as a result of F- stimulation and possibly as a consequence of phospholipase D activation, exerts a profound effect on cellular second messenger levels by modulating the conversion of phosphatidic acid to diacylglycerol.

scholarly journals Formation of bis(phosphatidyl)inositol and phosphatidic acid by phospholipase D action on phosphatidylinositol

1981 ◽  
Vol 195 (2) ◽  
pp. 521-523 ◽  
Author(s):  
N G Clarke ◽  
R F Irvine ◽  
R M C Dawson
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Phosphatidyl Inositol ◽  
Transphosphatidylation Reaction

Phosphatidylinositol is readily attacked by phospholipase D. Although some phosphatidic acid is produced by hydrolysis, the main products are bis(phosphatidyl)inositols most likely formed by a transphosphatidylation reaction between two substrate molecules.

Sign in / Sign up

Export Citation Format

Share Document