scholarly journals A shift in the optimum pH of phospholipase D produced by activating long-chain anions

1969 ◽  
Vol 112 (5) ◽  
pp. 795-799 ◽  
Author(s):  
R. H. Quarles ◽  
R. M. C. Dawson
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Activity Profile ◽  
Ph Optimum ◽  
Surface Ph ◽  
Electrophoretic Mobilities ◽  
Low Concentrations ◽  
Optimum Ph ◽  
Long Chain

1. The activity of phospholipase D (phosphatidylcholine phosphatidohydrolase, EC 3.1.4.4) towards ultrasonically treated phosphatidylcholine or large phosphatidylcholine particles activated with ether was maximal near pH5, and there was little activity above pH6. 2. When the enzyme was activated by the addition of phosphatidic acid to large phosphatidylcholine particles the pH optimum was shifted to pH6·5 irrespective of the amount of activator added. 3. When the enzyme was activated with low concentrations of dodecyl sulphate the pH optimum was 5·5 with little activity above pH6. With higher concentrations of dodecyl sulphate the pH–activity profile was shifted upwards towards a pH optimum of 6·5–6·6, the magnitude of the shift depending on the extent of the hydrolysis. 4. The shifts in the pH–activity profiles cannot be correlated with changes in the ‘surface pH’ of the substrate particles calculated from the measurement of their ζ-potentials (electrophoretic mobilities).

scholarly journals The hydrolysis of monolayers of phosphatidyl-[Me−14C]choline by phospholipase D

1969 ◽  
Vol 113 (4) ◽  
pp. 697-705 ◽  
Author(s):  
R. H. Quarles ◽  
R. M. C. Dawson
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
High Pressures ◽  
Specific Radioactivity ◽  
Maximal Activity ◽  
Enzymic Hydrolysis ◽  
Ph Optimum ◽  
High Specific Radioactivity ◽  
Rapid Hydrolysis ◽  
Hydrolysis Of

1. The hydrolysis of monolayers of phosphatidyl[Me−14C]choline at the air/water interface by phospholipase D (phosphatidylcholine phosphatidohydrolase) was investigated by a surface-radioactivity technique by using a flow counter. 2. Phosphatidylcholine of high specific radioactivity was prepared biosynthetically in good yield from [Me−14C]choline by using Saccharomyces cerevisiae. 3. At initial monolayer pressures between 12 and 25 dynes/cm. the hydrolysis occurred in two stages, an initial slow hydrolysis followed by a rapid hydrolysis. Below 3dynes/cm. and above 28dynes/cm. no enzymic hydrolysis of pure phosphatidylcholine monolayers could be detected. 4. The rapid hydrolysis was proportional to the enzyme concentration in the subphase, its pH optimum was 6·6, and 0·2mm-Ca2+ was required for maximal activity. 5. Hydrolysis of the film was accompanied by a pronounced fall in the surface pressure even though the phosphatidic acid formed did not leave the film. When the pressure fell to low values the hydrolysis ceased even if the film was only partially hydrolysed. 6. Above monolayer pressures of 28dynes/cm. enzymic hydrolysis could be initiated by inclusion of phosphatidic acid (and less effectively stearyl hydrogen sulphate) in the film, although the rates were not appreciably higher than those observed at 25dynes/cm. with a pure phosphatidylcholine film. 7. The initiation of the hydrolysis by phosphatidic acid was facilitated by the inclusion of high Ca2+ concentrations and certain carboxylic acid buffer anions in the subphase, although these did not activate by themselves. 8. The initiation of the hydrolysis at high pressures could not be related to any change in the surface potential brought about by the addition of the long-chain anions to the film, nor could it be ascribed to a surface dilution effect. 9. The results are discussed in relation to previous studies on the hydrolysis of phosphatidylcholine particles by the enzyme and also similar investigations on phosphatidylcholine monolayers with other phospholipases.

Interleukin-1 stimulates phosphatidic acid-mediated phospholipase D activity in human mesangial cells

1994 ◽  
Vol 266 (4) ◽  
pp. C1093-C1104 ◽  
Author(s):  
S. L. Bursten ◽  
W. E. Harris
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Mesangial Cells ◽  
Interleukin 1 ◽  
Type I ◽  
Human Mesangial Cells ◽  
Low Concentrations ◽  
Phosphatidate Phosphohydrolase ◽  
High Concentrations ◽  
Stimulation Of

Previous studies suggest that signal transduction mediated by interleukin-1 (IL-1), acting through an IL-1 receptor type found on T-cells and mesangial cells, may use phosphatidylethanolamine (PE) as a signaling molecule. Evidence presented here indicates that stimulation of human mesangial cells by IL-1 results in activation of a phospholipase D (PLD) that hydrolyzes PE to phosphatidic acid (PA). PLD acts on a subfraction of PE enriched in 1-o-alkyl and 1-o-alkenyl, sn-2-unsaturated species, generating a unique PA subspecies 30-120 s after stimulation. This PA species is subsequently converted to diradylglycerols by phosphatidate phosphohydrolase. The PE-directed PLD activity is abolished by antibodies against the IL-1 type I receptor and against IL-1. This specific PLD activity is also stimulated by low concentrations of 1,2-sn-dilinoleoyl PA, but not by high concentrations of 1-palmitoyl or 1-oleoyl lyso-PA. Blockade of PLD activation by IL-1 antibodies or antibody against the IL-1 receptor is bypassed by stimulation of human mesangial cells with 1,2-sn-dilinoleoyl PA. A novel system of signal cytokine mediation through PA self-amplification is indicated.

Phorbol Esters Stimulate Phospholipase D Activity in C-6 Glioma Cells

1989 ◽  
Vol 16 (3) ◽  
pp. 257-262
Author(s):  
Lena Gustavsson ◽  
Christofer Lundqvist ◽  
Christer Ailing
Keyword(s):  
Arachidonic Acid ◽  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Culture Medium ◽  
Phorbol Esters ◽  
Glioma Cells

The effects of phorbol esters on phospholipase D activity were studied in C-6 glioma cells. The cell lipids were prelabelled with [3H]-glycerol or [14C]-arachidonic acid. Phosphatidylethanol was formed during stimulation with 100nM 12-0-tetradecanoylphorbol-13-acetate (TPA), when ethanol was present in the culture medium. After 30 minutes of stimulation, phosphatidylethanol constituted 2.6% of the [3H]-glycerol-labelled lipids. Stimulating the cells with TPA in the absence of ethanol caused a significant increase in labelled phosphatidic acid. This increase was inhibited by ethanol. The present findings demonstrate that TPA stimulates phospholipase D activity in cultured C-6 glioma cells.

Phospholipase D1b and D2a generate structurally identical phosphatidic acid species in mammalian cells

2001 ◽  
Vol 360 (3) ◽  
pp. 707-715 ◽  
Author(s):  
Trevor R. PETTITT ◽  
Mark McDERMOTT ◽  
Khalid M. SAQIB ◽  
Neil SHIMWELL ◽  
Michael J. O. WAKELAM
Keyword(s):  
Liquid Chromatography ◽  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Mammalian Cells ◽  
Inducible Promoter ◽  
In Vitro Assays ◽  
Protein Levels ◽  
Intracellular Messenger

Mammalian cells contain different phospholipase D enzymes (PLDs) whose distinct physiological roles are poorly understood and whose products have not been characterized. The development of porcine aortic endothelial (PAE) cell lines able to overexpress PLD-1b or −2a under the control of an inducible promoter has enabled us to characterize both the substrate specificity and the phosphatidic acid (PtdOH) product of these enzymes under controlled conditions. Liquid chromatography–MS analysis showed that PLD1b- and PLD2a-transfected PAE cells, as well as COS7 and Rat1 cells, generate similar PtdOH and, in the presence of butan-1-ol, phosphatidylbutanol (PtdBut) profiles, enriched in mono- and di-unsaturated species, in particular 16:0/18:1. Although PtdBut mass increased, the species profile did not change in cells stimulated with ATP or PMA. Overexpression of PLD made little difference to basal or stimulated PtdBut formation, indicating that activity is tightly regulated in vivo and that factors other than just PLD protein levels limit hydrolytic function. In vitro assays using PLD-enriched lysates showed that the enzyme could utilize both phosphatidylcholine and, much less efficiently, phosphatidylethanolamine, with slight selectivity towards mono- and di-unsaturated species. Phosphatidylinositol was not a substrate. Thus PLD1b and PLD2a hydrolyse a structurally similar substrate pool to generate an identical PtdOH product enriched in mono- and di-unsaturated species that we propose to function as the intracellular messenger forms of this lipid.

scholarly journals Properties of pyruvate kinase and phosphoenolpyruvate carboxykinase in relation to the direction and regulation of phosphoenolpyruvate metabolism in muscles of the frog and marine invertebrates

1978 ◽  
Vol 174 (3) ◽  
pp. 979-987 ◽  
Author(s):  
Victor A. Zammit ◽  
Eric A. Newsholme
Keyword(s):  
Pyruvate Kinase ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Marine Invertebrates ◽  
Adductor Muscle ◽  
Sea Anemone ◽  
Anaerobic Conditions ◽  
Fructose Bisphosphate ◽  
Low Concentrations ◽  
Optimum Ph ◽  
Hill Coefficients

1. The properties of pyruvate kinase and, if present, phosphoenolpyruvate carboxykinase from the muscles of the sea anemone, scallop, oyster, crab, lobster and frog were investigated. 2. In general, the properties of pyruvate kinase from all muscles were similar, except for those of the enzyme from the oyster (adductor muscle); the pH optima were between 7.1 and 7.4, whereas that for oyster was 8.2; fructose bisphosphate lowered the optimum pH of the oyster enzyme from 8.2 to 7.1, but it had no effect on the enzymes from other muscles. Hill coefficients for the effect of the concentration of phosphoenolpyruvate were close to unity in the absence of added alanine for the enzymes from all muscles except oyster adductor muscle; it was 1.5 for this enzyme. Alanine inhibited the enzyme from all muscles except the frog; this inhibition was relieved by fructose bisphosphate. Low concentrations of alanine were very effective with the enzyme from the oyster (50% inhibition was observed at 0.4mm). Fructose bisphosphate activated the enzyme from all muscles, but extremely low concentrations were effective with the oyster enzyme (0.13μm produced 50% activation). 3. In general, the properties of phosphoenolpyruvate carboxykinase from the sea anemone and oyster muscles are similar: the Km values for phosphoenolpyruvate are low (0.10 and 0.13mm); the enzymes require Mn2+ in addition to Mg2+ for activity; and ITP inhibits the enzymes and the inhibition is relieved by alanine. These latter compounds had no effect on enzymes from other muscles. 4. It is suggested that changes in concentrations of fructose bisphosphate, alanine and ITP produce a coordinated mechanism of control of the activities of pyruvate kinase and phosphoenolpyruvate carboxykinase in the sea anemone and oyster muscles, which ensures that phosphoenolpyruvate is converted into oxaloacetate and then into succinate in these muscles under anaerobic conditions. 5. It is suggested that in the muscles of the crab, lobster and frog, phosphoenolpyruvate carboxykinase catalyses the conversion of oxaloacetate into phosphoenolpyruvate. This may be part of a pathway for the oxidation of some amino acids in these muscles.

scholarly journals Transformation of Construction Cement to a Self-Healing Hybrid Binder

2019 ◽  
Vol 20 (12) ◽  
pp. 2948 ◽  
Author(s):  
Werner E.G. Müller ◽  
Emad Tolba ◽  
Shunfeng Wang ◽  
Qiang Li ◽  
Meik Neufurth ◽  
...  
Keyword(s):  
Soil Bacteria ◽  
Construction Material ◽  
Water Soluble ◽  
The Self ◽  
Average Chain Length ◽  
Self Healing ◽  
Cement Concrete ◽  
Low Concentrations ◽  
Natural Mechanism ◽  
Long Chain

A new biomimetic strategy to im prove the self-healing properties of Portland cement is presented that is based on the application of the biogenic inorganic polymer polyphosphate (polyP), which is used as a cement admixture. The data show that synthetic linear polyp, with an average chain length of 40, as well as natural long-chain polyP isolated from soil bacteria, has the ability to support self-healing of this construction material. Furthermore, polyP, used as a water-soluble Na-salt, is subject to Na+/Ca2+ exchange by the Ca2+ from the cement, resulting in the formation of a water-rich coacervate when added to the cement surface, especially to the surface of bacteria-containing cement/concrete samples. The addition of polyP in low concentrations (<1% on weight basis for the solids) not only accelerated the hardening of cement/concrete but also the healing of microcracks present in the material. The results suggest that long-chain polyP is a promising additive that increases the self-healing capacity of cement by mimicking a bacteria-mediated natural mechanism.

Sign in / Sign up

Export Citation Format

Share Document