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Biomedical subjects

A G Lacko

Publications and source records attributed to A G Lacko.

At least 55 records · Page 3Linked to original sources
PubMed1983

Lecithin: cholesterol acyltransferase in Down's syndrome.

Based on earlier reports indicating that Down's syndrome may represent an atheroma-free human model, two groups of institutionalized subjects were compared with respect to various parameters of their plasma lipid transport system. One group of subjects was comprised of Down's syndrome subjects and the second, a group of mentally retarded individuals. Parameters measured included plasma cholesterol, triglyceride, HDL-cholesterol, apolipoprotein levels (A-I, B, C-III, and E), lecithin:cholesterol acyltransferase (LCAT) activity, body mass and blood pressure. Statistical analyses indicated no significant differences between the two groups except for the lower fractional rate of cholesterol esterification (% cholesterol esterified per hour, p = 0.0049) in the Down's syndrome subjects. Adjustment for the effects of body mass and age revealed no other significant differences between the two groups except for a lower molar rate of esterification (nmol cholesterol esterified X h-1 X ml-1, p less than 0.0063) in the Down's syndrome subjects. Additional differences between the two groups were revealed by partial correlational analyses of LCAT activity with the measured parameters or ratios of these parameters which suggests that the composition and/or metabolism of lipoproteins may differ between these two groups. Whether the lower LCAT activity and the other differences reflected by the correlational analyses contribute to the decreased incidence of atherosclerotic lesions in Down's syndrome remains to be elucidated.

Adolescent↗
PubMed1983

Characterization of lecithin:cholesterol acyltransferase from human plasma: II. Physical properties of the enzyme.

The physical properties of purified human plasma lecithin:cholesterol acyltransferase (LCAT) were investigated by techniques including analytical ultracentrifugation, ultraviolet spectroscopy, electrofocusing, and circular dichroism. The partial specific volume of LCAT was determined by sedimentation equilibrium ultracentrifugation experiments in H2O and D2O solutions (0.702 ml/g). The Mr was 67,000 by sodium dodecyl sulfate (SDS)-gel electrophoresis and 60,000 by sedimentation equilibrium ultracentrifugation. The discrepancy between the two sets of data presumably arose from the glycoprotein nature of the enzyme. Studies of the ultraviolet spectrum indicated that LCAT contained 6.5% (w/w) tyrosine which corresponds to approximately 18 tyrosine residues/mol of LCAT (polypeptide Mr 45,000). Spectrophotometric titration of the ionizable phenolic side chains indicated that nearly all the tyrosine residues were buried at neutral pH while they became gradually exposed at higher pH. The apparent pK of this transition was about 12.0 contrasted with 9.8, the apparent pK of ionization of the free tyrosyl groups.

Centrifugation, Density Gradient↗
PubMed1983

Characterization of lecithin-cholesterol acyltransferase from human plasma. 3. Chemical properties of the enzyme.

The polypeptide molecular weight of lecithin-cholesterol acyltransferase (LCAT) (45000) was obtained by deducting the weight of carbohydrate moiety (25%, w/w) from the total molecular weight of 60000. LCAT was found to have a relatively high content of glutamic acid, aspartic acid, glycine, and leucine residues and four half-cystines. The carbohydrate content was found to be about 25% (w/w): hexoses, 13%; hexosamines, 6.2%; and sialic acid, 5.4%. The total number of 408 amino acid residues per mole and the mean residue weight of 110.3 were found. From fluorescence spectroscopy analysis, 6-7 mol of tryptophan were found per mole of LCAT in 10 mM phosphate (pH 7.4). However, when LCAT was digested by the mixture of chymotrypsin and pronase the tryptophan residues increased to 10-11 mol/mol of LCAT, which agrees well with data obtained previously by ultraviolet absorption spectroscopy. A partial specific volume of 0.707 mL/g was determined by compositional analysis. Human LCAT was found to have a relatively high extinction coefficient (E1%1cm) of 21 at 280 nm and neutral pH. Two residues of cysteine per mole of LCAT were estimated both in the presence or absence of sodium dodecyl sulfate by titration with 5,5'-dithiobis-2-nitrobenzoic acid. The enzyme showed a lower tendency to staining with Coomassie blue R-250 than bovine serum albumin. The enzyme was rapidly inactivated by diisopropyl fluorophosphate (DFP), regardless of whether the free sulfhydryl were blocked or not. The enzyme was also irreversibly inhibited by cysteine above concentrations of 1 mM.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1983

Distribution of apolipoprotein A-IV between the lipoprotein and the lipoprotein-free fractions of rat plasma: possible role of lecithin:cholesterol acyltransferase.

Plasma samples were incubated under various conditions to study the effect of in vitro incubation on apolipoprotein A-IV distribution between the lipoprotein and lipoprotein-free fractions. When plasma was fractionated immediately after bleeding, apolipoprotein A-IV was present in equal concentrations in the lipoprotein and lipoprotein-free fractions. After a 4-hr, 37 degrees C incubation, greater than 90% of total plasma apolipoprotein A-IV was present in the lipoprotein fraction and the percentage of plasma cholesterol present as cholesteryl ester increased from 58% to 74%. When plasma was incubated for 4 hr at 37 degrees C in the presence of 1.5 mM 5,5-dithiobis(2-nitrobenzoic acid) (DTNB), greater than 90% of total plasma apoA-IV was present in the lipoprotein-free fraction, whereas plasma cholesteryl ester concentration did not change. Incubating heat-inactivated plasma for 4 hr also resulted in the redistribution of apolipoprotein A-IV from the lipoprotein fraction to the lipoprotein-free fraction, concurrent with no change in cholesterol esterification. When heat-inactivated plasma was incubated in the presence of a purified lecithin:cholesterol acyltransferase preparation, cholesterol esterification was restored and apolipoprotein A-IV was redistributed from the lipoprotein-free fraction to the lipoprotein fraction in such a manner that greater than 90% was present in the lipoprotein fraction. No changes in apolipoprotein A-I and apolipoprotein E distributions were found under any of the above conditions. Thus, the in vitro plasma incubations show that apolipoprotein A-IV can move bidirectionally between lipoprotein and lipoprotein-free fractions; the direction of this movement depends on the condition of the incubation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1982

Study of the lecithin: cholesterol acyltransferase reaction with liposome and high density lipoprotein substrates.

The activity of highly purified preparations of human plasma lecithin: cholesterol acyltransferase were stabilized by precipitating the enzyme with ammonium sulfate and using the dilutions of the particulate lecithin: cholesterol acyltransferase suspension for enzyme assays. Ammonium sulfate concentrations in the assay mix up to 0.1 M had no significant effect on lecithin: cholesterol acyltransferase activity. The basic enzymatic properties of lecithin: cholesterol acyltransferase were investigated using liposomes and high density lipoprotein (HDL) substrates. pH optima for both substrates was approximately 8.0. The temperature dependence of lecithin: cholesterol acyltransferase activity resulted in non-linear Arrhenius plots with both substrates. The activity vs. temperature (degrees C) curves showed slight inflections at 30 degrees C, which may have been due to the relatively rapid inactivation of the enzyme above this temperature. HDL3 was found to be a better substrate than HDL or HDL2. HDL3 was also considerably better than egg phosphatidylcholine/cholesterol liposomes as an lecithin: cholesterol acyltransferase substrate. Addition of HDL2 to a reaction mix of enzyme and HDL3 indicated that HDL2 acts as an inhibitor of cholesterol esterification in this system.

Apolipoprotein A-I↗
PubMed1981

Separation and detection of lipoproteins in human serum by use of size-exclusion liquid chromatography: a preliminary report.

Components of human serum can be separated on the basis of differences in relative molecular mass by using size-exclusion "high-performance" liquid chromatography. Lipoproteins in fractions of the eluate can be quantitated by conventional chemical and enzymatic methods. Alternatively, if lipoproteins in the serum are selectively prestained with diformazan dye, the column effluent can be monitored spectrophotometrically at 580 nm, so that only the lipoprotein components of serum are detected. Samples of purified low-density lipoproteins, so stained and analyzed, provide peak-area values that are proportional to their concentration as evaluated by chemical methods. With this technique, the various lipoprotein classes can be quickly separated and their concentration estimated. These techniques should prove useful in clinical and research laboratories.

Azo Compounds↗
PubMed1981

A study of the interaction of lecithin: cholesterol acyltransferase with subfractions of high density lipoproteins.

High density lipoproteins (HDL) were isolated by a chromatographic procedure and subsequently fractionated on a DEAE cellulose (DE-52) column. Four fractions were separated and analyzed for lipid and protein composition and molecular weight. During ion exchange chromatography, one of the four fractions consistently coincided with lecithin:cholesterol acyltransferase (LCAT) activity. When the HDL fractions were incubated with highly purified LCAT preparations, the LCAT activity showed a dependence on unesterified cholesterol concentrations. The HDL subfraction eluting at the highest ionic strength was found to be the best substrate for LCAT. This subfraction exhibited apoprotein and lipid composition similar to HDL3 and contained 31% of the total apoprotein D present in all the subfractions. A positive correlation was found between LCAT activity and the cholesteryl ester/unesterified cholesterol ratio, and a negative correlation was found between LCAT substrate potential and apparent molecular weight of the HDL subfractions when these subfractions were incubated with LCAT. No correlation was apparent between LCAT activity, and the phospholipid/unesterified cholesterol ratio or with the apoA-I/apoA-II ratio.U

Cholesterol↗
PubMed1980

A novel chromatographic method for the preparation of high density lipoproteins.

High density lipoproteins (HDL) were isolated by a procedure employing polyanion precipitation and column chromatography. The product polyanion precipitation and column chromatography. The product was free of low denisty lipoproteins (LDL) but serum albumin (HSA) was still present. The remaining HSA was removed by an immunoadsorbent column. The HDL isolated by our method was compared to another HDL preparation isolated from the same plasma sample by the combination of ultracentrifugation and gel chromatgraphy. It was found to have approximately the same lipid and protein composition as the HDL isolated by conventional techniques. Minor differences included a higher phospholipid and apoprotein E content and lower triglyceride and ApoC II content of the HDL isolated by column chromatography. The method described here is considerably less tedious than earlier techniques, can be scaled up without substantial increase in labor and results in an approximately 30% higher yield than the method described by Rudel et al.

Adult↗
PubMed1978

Serum cholesterol esterification in hyperthyroidism and hypothyroidism.

The rate of serum cholesterol esterification was measured in twenty healthy subjects and compared to similar data obtained with seventeen hyperthyroid and ten hypothyroid subjects. No significant differences were noted in the rate of cholesterol esterification while differences in the fractional rates were highly significant (p less than 0.001); the hyperthyroid group being higher and the hypothyroid group lower than normal. There were no clear trends observed in the changes of the rate of cholesterol esterification upon therapy. However, the fractional rates always increased when hypothyroid patients became euthyroid and always decreased in hyperthyroid patients as the result of therapy.

Cholesterol↗
PubMed1976

On the interpretation and potential diagnostic value of the measurements related to lecithin:cholesterol acyltransferase activity.

At least one reliable method is available for measuring LCAT activity in human subjects(25), and this assay may help to probe even subtle changes in plasma lipids and lipoproteins. Since plasma lipids and lipoproteins are known to be present in abnormal amounts in a large number of metabolic diseases, the method of measuring the rate of cholesterol esterification in plasma could assume a substantial role in the diagnosis and (or) monitoring of treatment of pathologic states.

Acyltransferases↗