scholarly journals Palmitate activation and esterification in microsomal fractions of rat liver

1975 ◽  
Vol 152 (1) ◽  
pp. 39-49 ◽  
Author(s):  
K A Lloyd-Davies ◽  
D N Brindley
Keyword(s):  
Rat Liver ◽  
Maximum Rate ◽  
Microsomal Fraction ◽  
Synthetase Activity ◽  
Optimum Conditions ◽  
Glycerol Phosphate ◽  
Glycerolipid Synthesis ◽  
The Relationship ◽  
Rate Limiting ◽  
Substrate Concentrations

1. Palmitoyl-CoA synthetase activity in the microsomal fraction of rat liver was measured directly by palmitoyl-CoA production, and indirectly by converting the palmitoyl-CoA into palmitoylcarnitine under optimum conditions. Even in the latter system, palmitoyl-CoA accumulated. The rate of palmitoyl-CoA hydrolysis and the inhibition of palmitoyl-CoA synthetase by palmitoyl-CoA were each estimated to be less than 10% of the maximum rate of palmitoyl-CoA production. The concentration of palmitoyl-CoA present in the assay systems used for measuring palmitate esterification to glycerol phosphate and the activity of palmitoyl-CoA synthetase by using the carnitine-linked determination were measured. These concentrations were not altered by the addition of glycerol phosphate, or of carnitine plus carnitine palmitoyltransferase. The relationship between the activity of palmitoyl-CoA synthetase and the rate of glycerolipid synthesis was investigated. The latter activity was measured by using palmitoyl-CoA generated from palmitate, palmitoyl-AMP or palmitoylcarnitine. It is concluded that, at optimum substrate concentrations, the activity of glycerol phosphate acyltransferase is rate-limiting in the synthesis of phosphatidate by rat liver microsomal fractions. The implications of these results in the measurement of palmitoyl-CoA synthetase and in the control of glycerolipid synthesis are discussed.

scholarly journals The relationship between palmitoyl-coenzyme A synthetase activity and esterification of sn-glycerol 3-phosphate in rat liver mitochondria

1973 ◽  
Vol 132 (4) ◽  
pp. 697-706 ◽  
Author(s):  
Mariana Sánchez ◽  
David G. Nicholls ◽  
David N. Brindley
Keyword(s):  
Rat Liver ◽  
Specific Activity ◽  
Liver Mitochondria ◽  
Carnitine Palmitoyltransferase ◽  
Rat Liver Mitochondria ◽  
Synthetase Activity ◽  
Palmitoyl Carnitine ◽  
The Mean ◽  
Glycerolipid Synthesis ◽  
Rate Limiting

1. The specific activities for palmitoyl-CoA synthetase and for sn-glycerol 3-phosphate esterification, with palmitoyl-CoA generated either by the endogenous synthetase or from palmitoyl-(-)-carnitine, CoA and excess of carnitine palmitoyltransferase, were measured with rat liver mitochondria. 2. The mean specific activity of palmitoyl-CoA synthetase was approximately five- and seven-fold the rates of sn-glycerol 3-phosphate esterification from palmitate and palmitoyl-(-)-carnitine respectively. No significant correlation was found in different rats between the activities of palmitoyl-CoA synthetase and sn-glycerol 3-phosphate esterification from either acyl precursor. However, there was a significant correlation (r=0.83, P<0.001) between the rates of glycerolipid synthesis from palmitate and palmitoyl-(-)-carnitine. 3. The mean molar composition of the glycerolipid synthesized from palmitate was 58% lysophosphatidate, 31% phosphatidate and 11% neutral lipid. With palmitoyl-(-)-carnitine the equivalent values were 70, 23 and 7%, which were significantly different. 4. When palmitoyl-CoA synthetase had been inactivated by 60–70% after preincubation of mitochondria at 37°C, it became rate-limiting in glycerolipid biosynthesis. Additions of 1–5mm-ATP prevented inactivation of palmitoyl-CoA synthetase. 5. Preincubation also inhibited the oxidation of palmitate, palmitoyl-CoA, palmitoyl-(-)-carnitine and malate plus glutamate. These inhibitions could not be prevented by addition of ATP. 6. Diversion of palmitoyl-CoA to form palmitoyl-(-)-carnitine did not inhibit sn-glycerol 3-phosphate esterification. 7. The palmitoyl-CoA pool synthesized by the palmitoyl-CoA synthetase was augmented by adding partially purified synthetase or carnitine palmitoyltransferase and palmitoyl-(-)-carnitine. No stimulation of palmitate incorporation into glycerolipids occurred. 8. At low concentrations of Mg2+, palmitate, ATP and CoA the velocity with palmitoyl-CoA synthetase decreased more than that of glycerolipid synthesis from palmitate. 9. It is concluded that in the presence of optimum substrate concentrations the activity of sn-glycerol 3-phosphate acyltransferase and not of palmitoyl-CoA synthetase is rate-limiting in the synthesis of phosphatidate and lysophosphatidate in isolated rat liver mitochondria.

scholarly journals The effect of chronic diabetes, induced by streptozotocin, on the activities of some enzymes of glycerolipid synthesis in rat liver

1977 ◽  
Vol 168 (2) ◽  
pp. 147-153 ◽  
Author(s):  
P H Whiting ◽  
M Bowley ◽  
R G Sturton ◽  
P H Pritchard ◽  
D N Brindley ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Single Dose ◽  
Rat Liver ◽  
Diacylglycerol Acyltransferase ◽  
Diabetic Rats ◽  
Glycerol Phosphate ◽  
Choline Phosphotransferase ◽  
Phosphatidate Phosphohydrolase ◽  
Glycerolipid Synthesis ◽  
Increased Activity

1. Rats were injected with a single dose of 35mg of streptozotocin/kg body wt. They exhibited a diabetes that was characterized by glycosuria, polyuria, polydipsia, hyperphagia, hyperglycaemia, increased concentrations of unesterified fatty acids, glycerol and triacylglycerols in the serum and an increased activity of glucose 6-phosphatase in the liver. 2. After 10 weeks the hepatic activities of the microsomal glycerol phosphate acyltransferase, phosphatidate phosphohydrolase, phosphatidate cytidylyltransferase, diacylglycerol acyltransferase, choline phosphotransferase, CDP-diacylglycerolx—inositol phosphatidyltransferase and the soluble phosphatidate phosphohydrolase were measured. 3. The only significant changes were an increase in the activity of the soluble phosphatidate phosphohydrolase and a decrease in that of the CDP-diacylglycerol—inositol phosphatidyltransferase in the diabetic rats. 4. These results are discussed in relation to the control of glycerolipid synthesis.

scholarly journals ALTERATIONS IN BIOCHEMICAL COMPOSITION AND RIBONUCLEIC ACID METABOLISM INDUCED IN RAT LIVER BY CORTISONE

1956 ◽  
Vol 2 (3) ◽  
pp. 331-350 ◽  
Author(s):  
Charles Upton Lowe ◽  
Royden N. Rand
Keyword(s):  
Rat Liver ◽  
Biochemical Composition ◽  
Microsomal Fraction ◽  
Acid Metabolism ◽  
Chemical Constituents ◽  
Differential Centrifugation ◽  
Sucrose Solutions ◽  
The Relationship

An investigation of the effect of cortisone administration upon the chemical composition of intracellular particulates of rat liver has been made. Livers were homogenized in 0.25 M sucrose solutions and submitted to differential centrifugation. Five fractions were prepared: mitochondria (Mit), microsomes (Mi), ultracentrifugable (U), non-sedimentable (S), and nuclear (Nuc). Measurement was made of total and polymerized RNA, nitrogen, lipide P, and uptake of P32 by the RNA of each fraction. The following observations were made:— Cortisone administration caused a fall in concentration in all measured constituents except glycogen. On a per liver basis, however, total liver RNA was unchanged in amount; nitrogen content of Mi fell and that of S increased; the lipide P of Mit and Mi also decreased. The biochemical composition of a statistical mitochondrion was significantly altered; in contrast, the microsomal fraction decreased in amount, but the relationship between the chemical constituents was unchanged. When polymerized RNA was sought by a process involving precipitation from ethanol at 20°C., none was found in the Mit of cortisone livers and the amount in Mi was much less than found in the normal. When, however, precipitation was conducted at 4°C., yields of polymerized RNA in all fractions after cortisone were equal to or greater than those found in the normal. Furthermore, incubation of mixtures of homogenates from normal and cortisone livers resulted in loss of warm precipitable RNA. These data strongly suggest the presence of an enzyme in cortisone livers which upon incubation with normal livers made preparation of polymerized RNA virtually impossible by use of the warm method. This agent, thought to operate in vivo and in vitro, was not present in significant amounts in normal livers, since incubation in this instance had no effect upon the amount of polymerized RNA. Mit from cortisone livers obtained by the cold technique had a significantly decreased rate of incorporation of P32 even though the yield of RNA from this fraction was increased. To reconcile these observations, it was proposed that under the influence of cortisone a variant of normal RNA is synthesized or normal RNA is converted to this variant. This "new" RNA has new solubility properties, a new rate of incorporation of P32, and conceivably it cannot act as a template for normal protein synthesis.

scholarly journals Drugs affecting the synthesis of glycerides and phospholipids in rat liver. The effects of clofibrate, halofenate, fenfluramine, amphetamine, cinchocaine, chlorpromazine, demethylimipramine, mepyramine and some of their derivatives

1975 ◽  
Vol 148 (3) ◽  
pp. 461-469 ◽  
Author(s):  
D N Brindley ◽  
M Bowley
Keyword(s):  
Rat Liver ◽  
Inhibitory Effect ◽  
Diacylglycerol Acyltransferase ◽  
Major Effect ◽  
Glycerol Phosphate ◽  
Liver Slices ◽  
Phosphatidate Phosphohydrolase ◽  
Marked Accumulation ◽  
Glycerolipid Synthesis ◽  
Phosphatidylcholine Synthesis

The effects on glycerolipid synthesis of a series of compounds including many drugs were investigated in cell-free preparations and slices of rat liver. p-Chlorobenzoate, p-chlorophenoxyisobutyrate, halofenate, D-amphetamine, adrenaline, procaine and N-[2-(4-chloro-3-sulphamoylbenzoyloxy)ethyl]norfenfluramine had little inhibitory effect on any of the systems investigated. Two amphiphilic anions, clofenapate and 2-(p-chlorophenyl)-2-(m-trifluoromethylphenoxy)acetate, both inhibited glycerol phosphate acyltransferase and diacylglycerol acyltransferase at approx. 1.6 and 0.7 mm respectively. Clofenapate (1 mm) also inhibited the incorporation of glycerol into lipids by rat liver slices without altering the relative proportions of the different lipids synthesized. The amphilic amines, mepyramine, fenfluramine, norfenfluramine, hydroxyethylnorfenfluramine, N-(2-benzoyloxyethyl)norfenfluramine, cinchocaine, chlorpromazine and demethylimipramine inhibited phosphatidate phosphohydrolase by 50% at concentrations between 0.2 and 0.9 mm. The last four compounds inhibited glycerol phosphate acyltransferase by 50% at concentrations between 1 and 2.6 mm. None of the amines examined appeared to be an effective inhibitor of diacylglycerol acyltransferase. Norfenfluramine, hydroxyethylnorfenfluramine and N-(2-benzoyloxyethyl)norfenfluramine produced less inhibition of glycerol incorporation into total lipids than was observed with equimolar clofenapate. The major effect of these amines in liver slices was to inhibit triacylglycerol and phosphatidylcholine synthesis and to produce a marked accumulation of phosphatidate. The results are discussed in terms of the control of glycerolipid synthesis. They partly explain the observed effects of the various drugs on lipid metabolism. The possible use of these compounds as biochemical tools with which to investigate the reactions of glycerolipid synthesis is considered.

scholarly journals The relationship between palmitoyl-coenzyme A synthetase activity and esterification of sn-glycerol 3-phosphate by the microsomal fraction of guinea-pig intestinal mucosa

1973 ◽  
Vol 132 (4) ◽  
pp. 707-715 ◽  
Author(s):  
David N. Brindley
Keyword(s):  
Guinea Pig ◽  
Intestinal Mucosa ◽  
Specific Activity ◽  
Synthetase Activity ◽  
Glycerol Phosphate ◽  
Acyltransferase Activity ◽  
Low Concentrations ◽  
System 2 ◽  
The Mean ◽  
Glycerolipid Synthesis

1. With microsomal fractions of guinea-pig intestinal mucosa the mean specific activity of palmitoyl-CoA synthetase was approx. 1.3-fold the esterification of sn-glycerol 3-phosphate with palmitoyl-CoA generated by the endogenous synthetase. The latter activity was approx. 2.5- and 5-fold that when palmitoyl-CoA was generated from palmitoylcarnitine or when it was added directly to the assay system. 2. There were significant correlations (P<0.001) between the specific activities of palmitoyl-CoA synthetase and glycerolipid synthesis from either palmitate or palmitoylcarnitine. 3. The mean molar composition of glycerolipid synthesized from palmitate or palmitoylcarnitine was approx. 18% lysophosphatidate, 75% phosphatidate and 7% neutral lipid. 4. Glycerolipid synthesis from palmitate was inhibited by 80–90% after preincubation of microsomal fractions at 37°C for 40min and was caused by inactivation of palmitoyl-CoA synthetase. 5. Addition of 100–400mm-KCl inhibited palmitoyl-CoA synthetase activity and glycerolipid synthesis from palmitate but stimulated glycerol phosphate acyltransferase activity. 6. Diversion of palmitoyl-CoA synthesized by the endogenous synthetase to palmitoylcarnitine resulted in an almost stoicheiometric decrease in glycerolipid synthesis. 7. Addition of rac-1-monopalmitin promoted utilization of palmitoyl-CoA by the monoglyceride pathway but did not inhibit phosphatidate biosynthesis. 8. With rate-limiting concentrations of CoA and Mg2+ the relative decreases in velocity for palmitoyl-CoA synthetase and glycerolipid synthesis from palmitate were almost identical. However, low concentrations of palmitate and ATP produced greater decreases in synthetase activity than in glycerolipid synthesis. 9. There appears to be a fine balance between the activities of palmitoyl-CoA synthetase and glycerol phosphate acyltransferase, with neither activity being in excess with respect to phosphatidate synthesis.

scholarly journals The effect of exogenous δ-aminolaevulinate on rat liver haem and cytochromes

1972 ◽  
Vol 129 (5) ◽  
pp. 1095-1099 ◽  
Author(s):  
Robert Druyan ◽  
Aldon Kelly
Keyword(s):  
Rat Liver ◽  
Synthetase Activity ◽  
Adult Rats ◽  
Adult Rat ◽  
Haem Biosynthesis ◽  
Rate Limiting ◽  
Mitochondrial Cytochromes ◽  
Hepatic Cytochrome

The activity of δ-aminolaevulinate synthetase is generally regarded as rate-limiting for hepatic haem biosynthesis. It has been suggested that cytochrome synthesis may also be regulated by changes in δ-aminolaevulinate synthetase activity. This hypothesis was studied by injecting product, δ-aminolaevulinate, into adult rats over a 4–240h period. The concentrations of hepatic mitochondrial cytochromes a, b, c and c1 were unchanged by treatment with δ-aminolaevulinate, allylisopropylacetamide or phenobarbital. In control animals, total microsomal haem content equalled the sum of cytochromes b5 plus P-450. After δ-aminolaevulinate administration the total amount of microsomal haem, measured as the pyridine haemochromogen, exceeded these components, indicating the formation of a ‘free’ haem pool. Haem synthesis does not appear rate-limiting for hepatic cytochrome synthesis in the adult rat.

scholarly journals Evidence for the existence of isoenzymes of glycerol phosphate acyltransferase

1979 ◽  
Vol 177 (1) ◽  
pp. 283-288 ◽  
Author(s):  
H G Nimmo
Keyword(s):  
Rat Liver ◽  
Microsomal Fraction ◽  
Thiol Group ◽  
Subcellular Fractionation ◽  
Heat Inactivation ◽  
Mitochondrial Enzyme ◽  
Microsomal Enzyme ◽  
Glycerol Phosphate ◽  
Low Concentrations ◽  
High Concentrations

Subcellular-fractionation studies confirmed previous findings that rat liver glycerol phosphate acyltransferase was located in both mitochondria and the microsomal fraction. Studies of the two activities revealed several differences between them. The mitochondrial enzyme had a lower Km for sn-glycerol 3-phosphate and was more resistant to heat inactivation than was the microsomal enzyme. Some preparations of the mitochondrial enzyme were inhibited by high concentrations of glycerol phosphate. The mitochondrial enzyme was not inactivated by thiol-group reagents, whereas the microsomal enzyme was very rapidly inactivated by these compounds. However, the microsomal enzyme could be specifically protected against this inactivation by low concentrations of palmitoyl-CoA. The results indicate the existence of distinct isoenzymes of glycerol phosphate acyltransferase with different intracellular locations.

Influence of 1,4-benzdiazepin-2-ones on rat liver microsomal fraction and hepatocytes

1987 ◽  
Vol 21 (1) ◽  
pp. 5-8
Author(s):  
T. I. Davidenko ◽  
O. V. Sevast'yanov ◽  
L. N. Yakubovskaya
Keyword(s):  
Rat Liver ◽  
Microsomal Fraction ◽  
Liver Microsomal Fraction
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