Formation of radioactive diacylglycerol from radioisotope-labelled phosphatidylethanolamine by Escherichia coli extracts

1989 ◽  
Vol 67 (6) ◽  
pp. 288-292 ◽  
Author(s):  
H. Aubry ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Phospholipase C ◽  
Substrate Concentration ◽  
Incubation Medium ◽  
Isotope Ratio ◽  
Enzyme Concentration ◽  
Water Soluble ◽  
Ph Optimum ◽  
Direct Incorporation

Radioisotope-labelled phosphatidylethanolamine can be converted to radioactive diacylglycerol in the presence of added unlabelled diacylglycerol. With [14C-glycerol; 3H-acyl]phosphatidylethanolamine as substrate, the conversion to double-labelled diacylglycerol occurred without change in isotope ratio indicating that the whole diacylglycerol moiety of phosphatidylethanolamine was directly involved. With [3H-acyl; 32P]phosphatidylethanolamine, formation of [3H]diacylglycerol occurred without production of labelled water-soluble products and consequently no phospholipase C activity could be detected. Under similar conditions, a conversion of [14C-acyl]- or [3H-acyl]-diacylglycerol to labelled phosphatidylethanolamine could also be shown even in the presence of hydroxylamine. [14C-Glycerol; 3H-acyl] diacylglycerol was converted to double-labelled product without change in isotope ratio which again indicated a direct incorporation of the entire diacylglycerol molecule into phosphatidylethanolamine. Both types of conversions were dependent upon time, enzyme concentration, and substrate concentration, and both displayed a pH optimum of approximately 6 and required no added cofactors. In the presence of increasing concentrations of [14C-acyl]diacylglycerol, added to incubation medium containing [3H-acyl]phosphatidylethanolamine, equal amounts of [14C]phosphatidylethanolamine and [3H]diacylglycerol were formed which matched the decrease in [3H]phosphatidylethanolamine. From these results, we conclude that Escherichia coli has an enzyme that catalyses the exchange between the diacylglycerol moiety of phosphatidylethanolamine and free diacylglycerol, with complete sparing of the phosphoethanolamine moiety.Key words: diacylglycerol, phosphatidylethanolamine, exchange, Escherichia coli.

The lipase activity of a partially purified lipolytic enzyme of Escherichia coli

1978 ◽  
Vol 56 (5) ◽  
pp. 324-328 ◽  
Author(s):  
G. Nantel ◽  
Georgia Baraff ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Lipase Activity ◽  
Slow Rate ◽  
Water Soluble ◽  
Lipolytic Enzyme ◽  
Ph Optimum ◽  
Chain Acyl ◽  
Marked Preference ◽  
Hydrolysis Of ◽  
Long Chain

Escherichia coli lipase was found to have a broad pH optimum between pH 8 and 10. Long-chain acyl triacylglycerols such as trioleoylglycerol were hydrolysed at a relatively slow rate, whereas, the shorter-chain acyl derivative tricapryloylglycerol was not. Triacylglycerols and diacylglycerols were broken down at a rate 10- to 15-fold greater than that for monoacylglycerol. Simple esters such as methyloieate and cetylpalmitate were hydrolysed at rates greater than that of triacylglycerol. Water-soluble esters such as p-nitrophenylacetate were not attacked. Hydrolysis of lipase substrates occurred more readily in the presence of an anionic detergent such as taurocholate. The enzyme had no marked preference for the 1- or 3-position of triacylglycerols but attacked these positions much more readily than position 2. The enzyme also catalyzed transacylation reactions with simple alcohols such as methanol or ethanol.

scholarly journals Phosphatidylinositol phosphodiesterase in higher plants

1980 ◽  
Vol 192 (1) ◽  
pp. 279-283 ◽  
Author(s):  
R F Irvine ◽  
A J Letcher ◽  
R M C Dawson
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Higher Plants ◽  
Water Soluble ◽  
Apium Graveolens ◽  
Soluble Enzyme ◽  
Ph Optimum ◽  
Bivalent Cation ◽  
Detectable Activity ◽  
Ph Range

1. The lower regions of the stem of celery (Apium graveolens L.) contain a soluble enzyme that hydrolyses phosphatidylinositol. 2. The lipoidal product of hydrolysis is diacylglycerol, and the water-soluble products are 1:2-cyclic phosphoinositol and phosphoinositol in the approximate proportions of 60% and 40% respectively: this indicates that a phosphodiesterase (phospholipase C-like) activity is cleaving the phosphatidylinositol. 3. The enzyme requires a bivalent cation, Ca2+ being the most effective activator. 4. The enzyme has a pH optimum, depending on conditions of assay, of pH 5.9-6.6 and in this pH range shows no detectable activity against phosphatidylcholine or phosphatidylethanolamine. 5. The activity is stimulated by phosphatidic acid and slightly inhibited (30% at concentrations equimolar with phosphatidylinositol) by phosphatidylcholine. 6. The phosphodiesterase was also detected (but not quantified) in the tips of the flowers in cauliflowers, in outer leaves of onion and in the elongating stem of daffodils. 7. The enzyme's properties are compared with equivalent mammalian enzymes, and its possible role in the catabolism of phosphatidylinositol in higher plants is discussed.

The uptake and acylation of exogenous lysophosphatidylethanolamine by Escherichia coli cells

1980 ◽  
Vol 58 (12) ◽  
pp. 1381-1386 ◽  
Author(s):  
P. Hellion ◽  
F. Landry ◽  
P. V. Subbaiah ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Isotope Ratio ◽  
Inorganic Phosphate ◽  
Cell Envelope ◽  
Membrane Lipid ◽  
Water Soluble ◽  
Absorption Process ◽  
Outer Membranes ◽  
Escherichia Coli Cells

Escherichia coli envelopes were fractionated to yield inner and outer membrane fractions. Both these fractions were found to convert [14C]lysophosphatidylethanolamine to its diacyl analogue. Intact Escherichia coli cells were capable of absorbing exogenous labelled lysophosphatidylethanolamine and converting it to phosphatidylethanolamine. When the 14C- and 32P-labelled lyso analogue was used, both the absorption process and the conversion to diacyl analogue proceeded without a significant change in isotope ratio either in the presence or in the absence of added inorganic phosphate. The absorption process was not markedly stimulated by Ca2+ in the medium; it proceeded to an amount representing 25–30% of the endogenous membrane lipid and was accompanied by some degradation to water-soluble products which accumulated in the cell mainly, but also in the incubation medium. The absorbed lipid was recovered in both the inner and outer membrane fractions of the cell envelope. The results indicate that Escherichia coli inner and outer membranes are capable of absorbing exogenous lysophosphoglyceride and converting it into structurally useful diacyl analogue.

scholarly journals Lysolecithin Metabolism by Human Platelets

Blood ◽  
1971 ◽  
Vol 37 (6) ◽  
pp. 675-683 ◽  
Author(s):  
PETER ELSBACH ◽  
PENELOPE PETTIS ◽  
AARON MARCUS
Keyword(s):  
Fatty Acid ◽  
Free Fatty Acid ◽  
Incubation Medium ◽  
Human Platelets ◽  
Water Soluble ◽  
Soluble Product ◽  
Ph Optimum ◽  
Acid Addition ◽  
Reaction Conversion ◽  
Direct Acylation

Abstract Human platelets incubated with 32P lysolecithin convert this compound to platelet lecithin and the water-soluble product glycerylphosphorylcholine. Lecithin synthesis from lysolecithin by intact platelets has a pH optimum of 9, whereas platelet homogenates show a pH optimum of 7.5 for the same reaction. Conversion of lysolecithin to platelet lecithin takes place by direct acylation of lysolecithin with free fatty acid. Addition of thrombin or ADP to the incubation medium has no effect on lysolecithin acylation or breakdown and does not affect the degradation of 14C choline-labeled platelet lecithin. Polystyrene particles stimulate lecithin synthesis and lysolecithin breakdown to glycerylphosphorylcholine by intact platelets but have no effect when added to platelet homogenates. By converting lysolecithin to lecithin, platelets add 2% to their lecithin content in 1 hr.

scholarly journals Optimization of Chitosan hydrolysis by Cellulase Enzyme to produce Chitooligosaccharide

2017 ◽  
Vol 20 (K3) ◽  
pp. 74-82
Author(s):  
Hoai Van Bui ◽  
Quang An Dao ◽  
Nghiep Dai Ngo
Keyword(s):  
Substrate Concentration ◽  
Water Solubility ◽  
Enzyme Concentration ◽  
Water Soluble ◽  
Hydrolysis Time ◽  
Surface Response Method ◽  
Optimal Values ◽  
Response Method ◽  
Central Composite ◽  
Chitosan Hydrolysis

In order to increase the water solubility of chitosan and potential application to products that is good for human health. Chitosan was caried out the optimization of hydrolysis by cellulase to produce chitooligosaccharide. Chitosan with degree of deacetyl more than 80% and cellulase used in this study. Surface Response Method (RSM) - Central Composite Design (CCD) option is used to optimize the hydrolysis. Results of the study showed optimal values for hydrolysis such as 49 oC temperature, 5.9 pH, 0.76% substrate concentration, and 8.97 U/g enzyme concentration, 180 minutes hydrolysis time. More than 90% of the oligosaccharides produced were in the range less than 10 kDa. The research results are the premise for production of COS powder to water-soluble.

Standardization and bacteria inhibitory test of purified extract of mahogany (Swietenia mahagoni (L.) Jacq) seeds and leaves

2019 ◽  
Vol 10 (3) ◽  
pp. 2132-2138
Author(s):  
Virsa Handayani ◽  
Rezki Amriati Syarif ◽  
Ahmad Najib ◽  
Aktsar Roskiana Ahmad ◽  
Abdullah Mahmud ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Heavy Metal ◽  
Bitter Taste ◽  
Drying Shrinkage ◽  
Ash Content ◽  
Water Soluble ◽  
Lead Contamination ◽  
Specific Parameter ◽  
Chemical Content ◽  
Swietenia Mahagoni

Mahogany (Swietenia mahagoni (L.) Jacq) is one of the plants that is often used by the community as traditional medicine. One of them is antifungal, antibacterial, antidiabetic, and eczema. This study aims to obtain standardized extracts from mahogany seeds and leaves. Standardization of purified extract of mahogany has been carried out according to the monographs of extract standardization guidelines, which include testing of specific and non-specific parameters. The results of the specific parameter testing showed that the purified extract of mahogany seeds is a thick extract, brown to reddish, smells distinctive and has a bitter taste. While the purified extract of mahogany leaves is a thick extract, greenish-brown in color, distinctive smell and has a bitter taste. The chemical content of purified extract of mahogany seeds and leaves showed the presence of flavonoids, alkaloids, terpenoids and saponins. Water-soluble essence levels in mahogany seeds and leaves was 14.84% and 10.28%. While the ethanol-soluble essence levels in mahogany seeds and leaves were 15.38% and 12.43%. Testing of non-specific parameters on mahogany seeds and leaves showed the results of drying shrinkage levels of 0.22% and 8.84%, moisture content of 2.60% and 4.04%, total ash content of 1.71% and 1.93%, levels acidic insoluble ash 0.38% and 0.32%, Total Plate Number (ALT) of mahogany seed bacteria 1x102 colonies/g, Number of mahogany mold seeds 4x10 colonies/g, heavy metal lead contamination and cadmium in mahogany seeds 0.0607µg/g and<0.003µg/g. The inhibitory diameter of each concentration of seeds against Escherichia coli, 3%, 5%, 7%, and 9%, is 12,67; 13,67; 17,67; and 19,67 mm, respectively. The inhibitory diameter of each concentration of leaves against Escherichia coli, 3%, 5%, 7%, and 9%, is 10,27; 10,90; 13,46; and 15,68 mm, respectively.

scholarly journals The determination of binding parameters when the total and free substrate concentrations are not approximately equal

1979 ◽  
Vol 179 (3) ◽  
pp. 697-700 ◽  
Author(s):  
N Gains
Keyword(s):  
Substrate Concentration ◽  
Graphical Method ◽  
Enzyme Concentration ◽  
Binding Parameters ◽  
Free Concentration ◽  
Equilibrium Binding ◽  
Substrate Concentrations

By using a standard graphical method values of Km and V may be found that are independent of the conditions and assumptions that the total substrate concentration approximates to its free concentration and that Km is much larger than the enzyme concentration. The procedure is also applicable to the determination of equilibrium binding parameters of a ligand to a macromolecule.

scholarly journals Identification and characterization of a phospholipase C activity in resident mouse peritoneal macrophages. Inhibition of the enzyme by phenothiazines

1981 ◽  
Vol 197 (2) ◽  
pp. 523-526 ◽  
Author(s):  
Paul D. Wightman ◽  
Mary Ellen Dahlgren ◽  
James C. Hall ◽  
Philip Davies ◽  
Robert J. Bonney
Keyword(s):  
Phospholipase C ◽  
High Activity ◽  
Peritoneal Macrophages ◽  
Ph Optimum ◽  
Reaction Velocity ◽  
Neutral Ph ◽  
Maximum Reaction ◽  
Mouse Peritoneal Macrophages ◽  
Identification And Characterization

Resident mouse peritoneal macrophages contain a phospholipase C of high activity that is specific for phosphatidylinositol. The activity has a neutral pH optimum, is Ca2+-dependent and has a maximum reaction velocity of 525nmol/h per mg of protein. Certain phenothiazines are potent inhibitors of this activity.

scholarly journals Thermostable glucose isomerase from psychrotolerant Streptomyces species

1970 ◽  
Vol 1 ◽  
pp. 6-10 ◽  
Author(s):  
Bidur Dhungel ◽  
Manoj Subedi ◽  
Kiran Babu Tiwari ◽  
Upendra Thapa Shrestha ◽  
Subarna Pokhrel ◽  
...  
Keyword(s):  
Specific Activity ◽  
Fold Increase ◽  
Glucose Isomerase ◽  
Enzyme Concentration ◽  
Maximal Velocity ◽  
Ph Optimum ◽  
Streptomyces Spp ◽  
Optimum Ph ◽  
Optimum Reaction ◽  
Sepharose 4B

Glucose isomerase (EC 5.3.1.5) was extracted from Streptomyces spp., isolated from Mt. Everest soil sample, and purified by ammonium sulfate fractionation and Sepharose-4B chromatography. A 7.1 fold increase in specific activity of the purified enzyme over crude was observed. Using glucose as substrate, the Michaelis constant (KM<) and maximal velocity (Vmax) were found to be 0.45M and 0.18U/mg. respectively. The optimum substrate (glucose) concentration, optimum enzyme concentration, optimum pH, optimum temperature, and optimum reaction time were 0.6M, 62.14μg/100μl, 6.9, 70ºC, and 30 minutes, respectively. Optimum concentrations of Mg2+ and Co2+ were 5mM and 0.5mM, respectively. The enzyme was thermostable with half-life 30 minutes at 100ºC.DOI: 10.3126/ijls.v1i0.2300 Int J Life Sci 1 : 6-10

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