scholarly journals Acid lipase: a histochemical and biochemical study using triton X100-naphtyl palmitate micelles.

1978 ◽  
Vol 26 (9) ◽  
pp. 696-712 ◽  
Author(s):  
T Schaffner ◽  
V M Elner ◽  
M Bauer ◽  
R W Wissler
Keyword(s):  
Enzyme Activity ◽  
Neutral Lipids ◽  
Biochemical Study ◽  
Reticuloendothelial System ◽  
Acid Lipase ◽  
Human Acid ◽  
Specific Localization ◽  
Lipase A ◽  
Sclerotic Plaques ◽  
Triton X

Hydrolsis of a-naphtyl palmitate dispersed with the detergent Triton X-100 at acid pH was studied by a histochemical diazocoupling technique in both fixed sections and cultures of primate tissues as well as by a biochemical assay employing the same chromogenic substrate. Evidence for the exclusive hydrolysis of this artificial fatty acid ester substrate by acid lipases was gathered from (1) comparison of isoelectric focusing zymograms developed with different substrates, (2) kinetic analysis of enzyme activity in the presence or absence of inhibitors, including a natural substrate of acid lipase, trioleylglycerol, (3) specific localization of marked enzyme activity in certain tissues, and (4) absence of detectable enzyme activity in a case of human acid lipase deficiency (Wolman's disease). Histochemically, acid lipase activity was most readily detected in cells active in the uptake and processing of neutral lipids, i.e., the phagocytes of the reticuloendothelial system, the adrenal cortex and the lipid-storing cells in the athero-sclerotic plaques of arteries.

scholarly journals Mucin biosynthesis. Properties of a bovine tracheal mucin β-6-N-acetylglucosaminyltransferase

1985 ◽  
Vol 227 (2) ◽  
pp. 405-412 ◽  
Author(s):  
P W Cheng ◽  
W E Wingert ◽  
M R Little ◽  
R Wei
Keyword(s):  
Enzyme Activity ◽  
Freezing Point ◽  
Cetylpyridinium Chloride ◽  
Sodium Dodecyl ◽  
Sodium Deoxycholate ◽  
Gas Chromatography Mass Spectrometry ◽  
Tween 20 ◽  
Group A ◽  
Michaelis Constants ◽  
Triton X

We have characterized a bovine tracheal mucin beta-6-N-acetylglucosaminyltransferase that catalyses the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine to the C-6 of the N-acetylgalactosamine residue of galactosyl-β 1→3-N-acetylgalactosamine. Optimal enzyme activity was obtained between pH 7.5-8.5, at 5mM-MnCl2, and at 0.06-0.08% (v/v) Triton X-100 (or Nonidet P-40), or 0.5-5.0% (v/v) Tween 20. Ba2+, Mg2+ and Ca2+ could partially replace Mn2+, but Co2+, Fe2+, Cd2+ and Zn2+ could not. Sodium dodecyl sulphate, cetylpyridinium chloride, sodium deoxycholate, octyl beta-D-glucoside, digitonin and alkyl alcohols were less effective in enhancing enzyme activity, and dimethyl sulphoxide was ineffective. The apparent Michaelis constants were 1.25 mM for UDP-N-acetylglucosamine, 0.94-3.34 mM for freezing-point-depressing glycoprotein and 0.19 mM for periodate-treated blood-group-A porcine submaxillary mucin. Asialo ovine submaxillary mucin could not serve as the glycosyl acceptor. The structure of the 14C-labelled oligosaccharide obtained by alkaline-borohydride treatment of the product was identified as Gal beta 1→3(Glc-NAc beta 1→6)N-acetylgalactosaminitol by beta-hexosaminidase treatment, gas chromatography-mass spectrometry and 1H-n.m.r. (270 MHz) analysis. The enzyme is important in the regulation of mucin oligosaccharide biosynthesis.

Lysosomal Acid Lipase, a Novel Biomarker for Hepatic Fibrosis

2021 ◽  
Author(s):  
Lin An ◽  
Mi Zhang ◽  
Yuefang Lin ◽  
Ting Jiang ◽  
Liming Cai ◽  
...  
Keyword(s):  
Hepatic Fibrosis ◽  
Acid Lipase ◽  
Lysosomal Acid ◽  
Lysosomal Acid Lipase ◽  
Lipase A

scholarly journals Differential effects of non-ionic detergents on microsomal and sarcolemmal adenylate cyclase in cardiac muscle

1978 ◽  
Vol 175 (1) ◽  
pp. 171-180 ◽  
Author(s):  
Prakash V. Sulakhe ◽  
Njanoor Narayanan
Keyword(s):  
Enzyme Activity ◽  
Sarcoplasmic Reticulum ◽  
Adenylate Cyclase ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Protein Ratio ◽  
Guinea Pig Heart ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Ionic Detergent ◽  
Triton X

1. About 4 and 23% of the homogenate adenylate cyclase activity was recovered in the microsomal and sarcolemmal fractions isolated from guinea-pig heart ventricles. 2. Cardiac microsomal adenylate cyclase activity [basal as well as p[NH]ppG (guanyl-5′-yl imidodiphosphate)- and NaF-stimulated] was increased over 2-fold in the presence of Lubrol-PX (0.01–0.1%). 3. The sarcolemmal enzyme, however, showed concentration-dependent inhibition caused by the detergent under all assay conditions, except when p[NH]ppG was included in the assay. In the latter case, the detergent (0.01–0.02%) caused a modest increase (30–45%) in enzyme activity. 4. Another non-ionic detergent, Triton X-100, also stimulated the microsomal cyclase and inhibited the sarcolemmal enzyme. 5. With either membrane fraction, Lubrol-PX solubilized the enzyme when the detergent/membrane protein ratio was 2.5 (μmol of detergent/mg of protein). 6. The findings with homogenate and a washed particulate fraction resembled those obtained with sarcolemma, and those with isolated sarcoplasmic reticulum resembled those with microsomal preparations. 7. p[NH]ppG, and to some extent NaF, protected the detergent-induced inactivation of the enzyme observed at higher detergent concentrations (0.5% Lubrol-PX and 0.05–0.5% Triton X-100). 8. In the absence of detergents, p[NH]ppG increased the basal enzyme activity about 2-fold in microsomal fractions, but did not appreciably stimulate the sarcolemmal enzyme. Isoproterenol, on the other hand, increased the sarcolemmal enzyme activity (>2-fold) in the presence of p[NH]ppG and caused only moderate stimulation (31%) of the microsomal enzyme under these conditions. 9. These findings support the view that, although the bulk of adenylate cyclase resides in heart sarcolemma (plasma membrane), the microsomal activity cannot be accounted for solely by contamination of the microsomal fraction with sarcolemma, as has been suggested by others [Besch, Jones & Watanabe (1976) Circ. Res.39, 586–595; Engelhard, Plut & Storm (1976) Biochim. Biophys. Acta451, 48–61]. Further, the results of this study show that cardiac sarcoplasmic-reticulum membranes possess this enzyme.

Proposed Treatment for Infants With Wolman Disease

PEDIATRICS ◽  
1989 ◽  
Vol 83 (6) ◽  
pp. 1074-1075
Author(s):  
MOSHE WOLMAN
Keyword(s):  
Lysosomal Enzyme ◽  
Cholesterol Ester ◽  
Reticuloendothelial System ◽  
Cholesterol Esters ◽  
Enzyme Molecule ◽  
Acid Lipase ◽  
Wolman Disease ◽  
Cholesterol Ester Hydrolase ◽  
Ester Hydrolase ◽  
Available Information

Presently available information indicates that Wolman disease is due to a defect in a single lysosomal enzyme molecule, called acid lipase, esterase, or cholesterol ester hydrolase (EC3.1.1.13). Cells of the reticuloendothelial system, hepatocytes, adrenocortical, and presumably many other cells normally, contain a neutral esterase associated with their microsomes, which is not deficient in Wolman disease. Theoretically, catabolism of intracellular hydrophobic cholesterol (esters and triglycerides) should proceed normally in Wolman disease cells whenever the metabolic chain does not depend on lysosomal hydrolysis.

The improvement of lipase secretion and stability by addition of inert compounds into Acinetobacter calcoaceticus cultures

2002 ◽  
Vol 48 (12) ◽  
pp. 1056-1061 ◽  
Author(s):  
Daniela A Martinez ◽  
B Clara Nudel
Keyword(s):  
Enzyme Activity ◽  
Gum Arabic ◽  
Acinetobacter Calcoaceticus ◽  
Glass Beads ◽  
High Yield ◽  
Poly Ethylene Glycol ◽  
Poly Ethylene ◽  
Cell Fractions ◽  
Triton X ◽  
Effective Additive

Acinetobacter calcoaceticus BD413 produces variable amounts of an exocellular lipase that becomes rapidly inactivated upon secretion. To achieve high yield and protect the enzyme, we assayed the addition of several inert compounds to cell-free supernatants, cell fractions, and whole cultures. Glass beads, poly(ethylene glycol) 600, Triton X-100, saccharose, gum arabic, and β-cyclodextrin were among the compounds tested. β-Cyclodextrin and gum arabic (and saccharose to a lesser extent) were effective enzyme stabilizers in cell-free supernatants, while gum arabic, glass beads, and Triton X-100 improved lipase secretion from cells, and, therefore, total lipase yield (30–50%, according to the additive). In whole cultures, β-cyclodextrin was the most effective additive, particularly in combination with glass beads or gum arabic. Indeed, cultures containing β-cyclodextrin plus gum arabic were able to maintain 95% (±1.5%) of the initial lipase activity for more than 16 h, while control cultures with no additives maintained only 10% (±4%) of the enzyme activity after the same period. In conclusion, the addition of inert compounds in cultures may be considered a useful approach for achieving increased yield and lipase stabilization, amenable for downstream processing.Key words: Acinetobacter calcoaceticus, lipase, secretion, stabilization, inert additives.

Lysosomal acid lipase A and the hypercholesterolaemic phenotype

2013 ◽  
Vol 24 (4) ◽  
pp. 332-338 ◽  
Author(s):  
Sigrid W. Fouchier ◽  
Joep C. Defesche
Keyword(s):  
Acid Lipase ◽  
Lysosomal Acid ◽  
Lysosomal Acid Lipase ◽  
Lipase A

scholarly journals Multiple forms of acetylcholinesterase from human erythrocytes

1973 ◽  
Vol 133 (3) ◽  
pp. 521-527 ◽  
Author(s):  
David L. Wright ◽  
David T. Plummer
Keyword(s):  
Enzyme Activity ◽  
Electrophoretic Pattern ◽  
Human Erythrocytes ◽  
Serum Cholinesterase ◽  
Main Peak ◽  
Multiple Forms ◽  
Molecular Weights ◽  
Enzyme Substrate ◽  
Triton X ◽  
Naphthyl Acetate

1. Acetylcholinesterase from human erythrocytes was solubilized with Triton X-100 in strong salt solution and partially purified by (NH4)2SO4 fractionation. This preparation showed three main bands of enzyme activity after electrophoresis on polyacrylamide gel and incubation with either α-naphthyl acetate or acetylthiocholine as enzyme substrate. Two of the multiple forms were completely inhibited by 10μm-eserine and one only partially. Treatment with neuraminidase had no effect on the electrophoretic pattern; therefore sialic acid does not appear to determine or affect the ratios of the acetylcholinesterase multiple forms, unlike those of the serum cholinesterase. 2. Chromatography of the preparation on Sephadex G-200 revealed one major peak of enzyme activity and a suggestion of two minor zones of mol.wt. 546000, 184000 and 93000 (i.e. in the proportion 6:2:1). The main peak was almost completely separated from the Triton X-100 and the overall purification was about 600-fold. Further attempts to purify the enzyme by absorption on calcium phosphate gels were unsuccessful. 3. Electrophoresis of the enzyme preparation on a polyacrylamide gradient for 24h revealed three main bands that corresponded to the three values for molecular weights obtained by column chromatography. After 70h of electrophoresis a further three zones of activity developed making six molecular entities, the molecular weights of which were simple multiples of a monomer, thus resembling the cholinesterase found in serum.

scholarly journals Localization of oxidized nocotinamide–adenine dinucleotide glycohydrolase in the mouse liver nuclear envelope

1979 ◽  
Vol 178 (2) ◽  
pp. 467-473 ◽  
Author(s):  
P Tamulevicius ◽  
C Streffer ◽  
O Roscic ◽  
E Hubert
Keyword(s):  
Electron Microscopy ◽  
Enzyme Activity ◽  
Nuclear Envelope ◽  
Mouse Liver ◽  
Residual Activity ◽  
Latent Form ◽  
Nad Glycohydrolase ◽  
Nuclear Membranes ◽  
Deoxyribonuclease I ◽  
Triton X

NAD+ glycohydrolase activity located in the nuclear envelope was maximally solubilized by treatment with 0.1–0.2% Triton X-100. The residual activity largely represents the chromatin-associated NAD+ glycohydrolase. Under these conditions the phospholipids were extensively solubilized (over 90%) while leaving the nuclei physically stable, although the nuclear membranes were removed, as shown by electron microscopy. After Triton X-100 treatment, deoxyribonuclease I did not significantly affect the residual NAD+ glycohydrolase activity, although the DNA was completely broken down. This enzyme activity can be released from the nuclear pellet by incubation with phospholipase C. For comparative studies, the glucose 6-phosphatase activity, known to be present in the nuclear envelope, was investigated. Treatment with 0.01% Triton X-100 released 10–20% of the phospholipids, but without solubilizing either glucose 6-phosphatase or NAD+ glycohydrolase. Higher Triton X-100 concentrations (0.1–1.0%) inhibited glucose 6-phosphatase, but not NAD+ glycohydrolase activity. NAD+ glycohydrolase is apparently present in a latent form in the nuclear envelope. Glucose 6-phosphatase, However, shows no such latency.

scholarly journals The effect of phenobarbital on the submicrosomal distribution of uridine diphosphate glucuronyltransferase from rat liver

1970 ◽  
Vol 117 (2) ◽  
pp. 319-324 ◽  
Author(s):  
G. J. Mulder
Keyword(s):  
Enzyme Activity ◽  
Density Gradient ◽  
Rat Liver ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Sucrose Density Gradient ◽  
Male Rats ◽  
Uridine Diphosphate ◽  
Triton X

1. The detergent Triton X-100 activates UDP glucuronyltransferase from rat liver in vitro six- to seven-fold with p-nitrophenol as substrate. The enzyme activity when measured in the presence of Triton X-100 is increased significantly by pretreatment of male rats with phenobarbital for 4 days (90mg/kg each day intraperitoneally). If no Triton X-100 is applied in vitro such an increase could not be shown. In all further experiments the enzyme activity was measured after activation by Triton X-100. 2. The Km of the enzyme for the substrate p-nitrophenol does not change on phenobarbital pretreatment. 3. When the microsomal fraction from the liver of untreated rats is subfractionated on a sucrose density gradient, 47% of the enzyme activity is recovered in the rough-surfaced microsomal fraction, which also has a higher specific activity than the smooth-surfaced fraction. 4. Of the increase in activity after the phenobarbital pretreatment 50% occurs in the smooth-surfaced fraction, 19% in the rough-surfaced fraction and 31% in the fraction located between the smooth- and rough-surfaced microsomal fractions on the sucrose density gradient. 5. The latency of the enzyme in vitro, as shown by the effect of the detergent Triton X-100, is discussed in relation to the proposed heterogeneity of UDP glucuronyltransferase.

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