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Biomedical subjects

S Gatt

Publications and source records attributed to S Gatt.

At least 127 records · Page 7Linked to original sources
PubMed1980

Utilization of membranous lipid substrates by membranous enzymes. Hydrolysis of sphingomyelin in erythrocyte 'ghosts' and liposomes by the membranous sphingomyelinase of chicken erythrocyte 'ghosts'.

Incubation at 37 degrees C of haemolysed chicken erythrocytes ('chicken erythrocyte ghosts') resulted in hydrolysis of the membrane sphingomyelin, suggesting an activation of a latent sphingomyelinase during the haemolysis procedure. When this incubation was continued for several hours, the entire sphingomyelin of the erythrocyte 'ghosts' was hydrolysed and membranes were obtained that were devoid of sphingomyelin, but had an active sphingomyelinase. Mixing the latter membranes with human erythrocyte 'ghosts' or positively charged liposomes led to hydrolysis of the sphingomyelin in these two membranes. This suggested that, after haemolysis, the activated sphingomyelinase in the membrane of the chicken erythrocyte 'ghosts' could hydrolyse sphingomyelin in its own membrane ('intramembrane utilization') or adjacent membranes ('intermembrane utilization').

Animals↗
PubMed1980

Enzymic hydrolysis of sphingomyelin in the presence of bile salts.

Sphingomyelin in mixed dispersion with bile salts was hydrolysed by the solubilized sphingomyelinase of rat brain lysosomes. In parallel studies, physical properties of these dispersions were determined. The kinetic curves that described the rate of hydrolysis as a function of increasing concentrations of bile salt were multiphasic. A region of very low activity was followed by an ascending portion, a peak, a descending portion, a trough and a second ascending portion. The positions of the initiation points, peaks and troughs were found to be a function of the respective ratios of the bile salt to sphingomyelin for the detergent sodium taurodeoxycholate, but of the absolute concentration of the detergent for sodium taurocholate. Turbidity studies suggested that hydrolysis of sphingomyelin begins at a bile salt concentration that solubilizes the lipid and incorporates it into a mixed micelle with the detergent. Ultracentrifugation studies suggested that the sizes of the mixed aggregates of detergent and lipid were a function of the ratio of taurodeoxycholate to sphingomyelin, but of the absolute concentration of the bile salt, for sodium taurocholate.

Animals↗
PubMed1980

Hydrolysis of di- and trisialo gangliosides in micellar and liposomal dispersion by bacterial neuraminidases.

The hydrolysis of di- and trisialo gangliosides by bacterial neuraminidases was investigated. Slow rates of hydrolysis were obtained with micellar dispersions of the pure gangliosides; the rates increased considerably with mixtures of ganglioside and phospholipids, such as phosphatidylcholine or sphingomyelin. The greatest rates of hydrolysis were obtained with mixtures containing 5-10 mol% ganglioside and 90-95% phospholipid. With the aid of the nonpenetrating reagent trinitrobenzenesulfonic acid, it was ascertained that this mixture consisted of sealed, unilamellar vesicles in which the ganglioside was distributed symmetrically between the two layers of the liposome. When the relative proportion of the ganglioside was increased, the dispersions contained liposomes admixed with micelles of ganglioside and phospholipid. The rates of hydrolysis of the ganglioside could be correlated with the percentage of sealed vesicles in each mixture. Experiments in which another ganglioside (GM1) or cholesterol was incorporated into the mixed dispersions further supported this conclusion. It is suggested that the rate of hydrolysis is affected predominantly by interactions between the carbohydrate chains of ganglioside molecules. The data emphasize that ganglioside metabolism can be best studied when the latter are part of biological or model membranes.

Clostridium perfringens↗
PubMed1980

Hydrolysis of gangliosides in micellar and liposomal dispersion by bacterial neuraminidases.

Aqueous dispersions of pure gangliosides contain micelles of these compounds. In this dispersion state, the rates of hydrolysis of the neuraminyl residues by bacterial neuraminidases are slowest. Incorporation of gangliosides into mixed dispersion with other lipids or into mixed micelles with bile salts considerably increases the reaction rates. The greatest reaction rates are obtained when di- or trisialogangliosides are incorporated into unilamellar vesicles of lecithin or sphingomyelin.

Clostridium perfringens↗
PubMed1980

The synthesis of cholesteryl alkyl ethers.

Seventeen cholesteryl alkyl ethers were synthesized through alcoholysis of cholesterol p-toluenesulfonate. This method was found superior to the etherification of sodium or potassium cholesterylate with alkyl halides or methanesulfonates, especially for the preparation of long-chain unsaturated aklyl ethers of [7(m)-3H]cholesterol of high specific activity.

Cholesterol↗
PubMed1980

A fluorometric determination of sphingomyelinase by use of fluorescent derivatives of sphingomyelin, and its application to diagnosis of Niemann-Pick disease.

We synthesized fluorescent derivatives of sphingomyelin (N-acyl-sphingosylphosphocholine) and used them as substrates for several sphingomyelinases. The following five fluorescent probes, each attached to the terminal carbon atom of the fatty acyl residue, were introduced into sphingomyelin: dansyl, pyrene, carbazole, 4-chloro-7-nitrobenz-2-oxa-1,3-diazole, and anthroic acid. We compared the rates at which the fluoro- and radiolabeled sphingomyelins were hydrolyzed. They were the same with the following three spingomyelinases: (a) a purified enzyme from Staphylococcus aureus; (b) at Triton X100-treated extract of human brain (assayed at pH 7.4 in the presence of Mg2+; and (c) aqueous extracts of brain lysosomes, skin fibroblasts, and amniotic cells, assayed at pH 5.0. Homogenates of skin fibroblasts of a patient with Niemann-Pick disease had practically no activity when assayed at pH 5, with fluorosphingomyelin as substrate. When fluorosphingomyelin was mixed in various proportions with natural sphingomyelin, enzymes from each of the three sources hydrolyzed the two substrates at equal rates. The fluorosphingomyelins can be used to estimate with great sensitivity the sphingomyelinase activity in extracts of tissues or cells, in tears, and probably in hair follicles, as well as diagnose Niemann-Pick disease, either pre- or post-natally

Amniotic Fluid↗
PubMed1979

Interaction of membranous enzymes with membranous lipid substrates. Hydrolysis of diacylglycerol by lipase in rat brain microsomes.

The phospholipids in rat brain microsomes were labeled with tritium by intracerebral administration of radioactive fatty acids and converted to diacylglycerol with phospholipase C. The latter lipid was hydrolyzed in situ at pH 4.8, to monoacylglycerol and fatty acid by the endogenous microsomal lipase. This paper provides an experimental approach to determine whether the lipid was degraded by enzyme molecules residing in its own membrane (intramembrane interaction) or an adjacent membrane (intermembrane interaction). Direct interaction between separate membranes containing enzyme or substrate showed the existence of the inter-membrane route while dilution experiments provided evidence for the presence of the intramembrane interaction as well. A probable difference in the mechanisms of these two interactions is suggested by different shapes of the curves that describe the reaction rate as a function of the endogenous substrate. The curve resulting from the intermembrane interaction was hyperbolic while that representing the intramembrane route was of a parabola-like shape. Competition experiments suggested that when given a choice between the two, the enzyme utilized preferentially the substrate molecules in its own membrane.

Animals↗
PubMed1978

Magnesium-dependent sphingomyelinase of infantile brain. Effect of detergents and a heat-stable factor.

The properties of the Mg2+-dependent sphingomyelinase, whose pH optimum is between 7 and 8, were investigated using post-mortem infantile brain. The enzyme could be extracted with 0.2% Triton X-100 and remained soluble when centrifuged at 170,000 X g. Subsequent removal of the detergent with SM2-Biobeads resulted in resedimentation of the enzyme at 80,000 X g. A detergent was needed for assaying enzymatic activity; either Triton X-100 or bile salts could be used. With increasing concentrations of detergent, the rates of hydrolysis of sphinomyelin increased, reached an optimum and then decreased, suggesting inhibition of the enzyme. The concentrations of detergent which resulted in optimal reaction rates were directly related to the protein concentration of the enzymatic preparation. A heat-stable factor which counteracts inhibition by the above detergents is present in brain as well as several other tissues. A lipid extract of the enzymatic preparation, or several purified lipids could not mimic the effect of the heat-stable factor. The interrelationship between enzyme, detergent and the heat-stable factor was investigated.

Brain↗
PubMed1978

Synthesis of trinitrophenylaminolauric acid and the use of its glyceryl esters for assaying lipase by a spectrophotometric procedure.

Trinitrophenylaminolauric acid was synthesized from omega-aminolauric acid and trinitrobenzenesulfonic acid and then condensed with glycerol to yield mono-, di- and triacylglyceryl esters of this acid. Hydrolysis of these glycerides was followed by isolation of the yellow fatty acid with the aid of one solvent extraction step and estimating its content by spectrophotometry. This procedure was used to assay the activities of lipases from hog pancreas, rat bile, microsomes of rat brain and Rhizopus arrhizus delamar.

Animals↗
PubMed1978

A spectrophotometric method for determination of sphingomyelinase.

A colored derivative of sphingomyelin was synthesized and used as substrate for several sphingomyelinases. The compound is N-omega-trinitrophenyl-aminolaurylsphingosylphosphorylcholine. The rate of hydrolysis of this substrate was compared to that of bovine brain sphingomyelin, labelled with tritium in the choline moiety. The following enzyme preparations were used: homogenate-less debris of brain, assayed at pH 5.0 or 7.4; a solubilized preparation derived from rat brain lysosomes, assayed at pH 5.0 and a purified enzyme of Staphylococcus aureus. With all preparations, the rates of hydrolysis of the yellow derivative were very similar to those of the brain sphingomyelin. Extracts of skin fibroblasts of normal and Niemann-Pick patients as well as amniotic cells were also used. Again, the rates of hydrolysis of the yellow derivative practically equalled those using brain sphingomyelin.

Animals↗
PubMed1978

The hydrolysis of triacylglycerol and diacylglycerol by a rat brain microsomal lipase with an acidic pH optimum.

Lipase activity towards triacylglycerol and diacylglycerol was measured at pH 4.8 using a microsomal preparation from rat brain as the enzyme source. The optimal pH for the hydrolysis of triacylglycerol was 4.8, with only minor lipolytic activity in the alkaline pH range. Diacylglycerol was the major product of triacylglycerol hydrolysis, with only little monoacylglycerol being formed. When diacylglycerol was the starting substrate it was hydrolyzed at a rate 10-fold greater than triacylglycerol, and the product was monoacylglycerol. The enzyme showed positional specificity for the fatty acid moieties located at the primary positions of sn-glycerol. 1,3-Diacylglycerol was hydrolyzed at greater than twice the rate of the corresponding 1,2(2,3)-isomer.

Animals↗