scholarly journals Differences between microsomal and mitochondrial-matrix palmitoyl-coenzyme A hydrolase, and palmitoyl-l-carnitine hydrolase from rat liver

1979 ◽  
Vol 181 (1) ◽  
pp. 119-125 ◽  
Author(s):  
R K Berge ◽  
B Døssland
Keyword(s):  
Rat Liver ◽  
Microsomal Fraction ◽  
Temperature Stability ◽  
Fatty Acyl ◽  
Mitochondrial Matrix ◽  
Microsomal Enzymes ◽  
Coa Hydrolase ◽  
Hydrolysis Of ◽  
Temperature Optima ◽  
Matrix Enzyme

Palmitoyl-CoA hydrolase (EC 3.1.2.2) and palmitoyl-L-carnitine hydrolase (EC 3.1.1.28) activities from rat liver were investigated. 1. Microsomal and mitochondrial-matrix palmitoyl-CoA hydrolase activities had similar pH and temperature optima, although the activities showed different temperature stability. They were inhibited by Pb2+ and Zn2+. The palmitoyl-CoA hydrolase activities in microsomal fraction and mitochondrial matrix were differently affected by the addition of Mg2+, Ca2+, Co2+, K+ and Na+ to the reaction mixture. ATP, ADP and NAD+ stimulated the microsomal activity and inhibited the mitochondrial-matrix enzyme. The activity of both the microsomal and mitochondrial-matrix hydrolase enzymes was specific for long-chain fatty acyl-CoA esters (C12-C18), with the highest activity for palmitoyl-CoA. The apparent Km for palmitoyl-CoA was 47 microM for the microsomal enzyme and 17 microM for the mitochondrial-matrix enzyme. 2. The palmitoyl-CoA hydrolase and palmitoyl-L-carnitine hydrolase activities of microsomal fraction had similar pH optima and were stimulated by dithiothreitol, but were affected differently by the addition of Pb2+, Mg2+, Ca2+, Mn2+ and cysteine. The two enzymes had different temperature-sensitivities. 3. The data strongly suggest that palmitoyl-CoA hydrolase and palmitoyl-L-carnitine hydrolase are separate microsomal enzymes, and that the hydrolysis of palmitoyl-CoA in the microsomal fraction and mitochondria matrix was catalysed by two different enzymes.

scholarly journals A highly basic N-terminal extension of the mitochondrial matrix enzyme ornithine transcarbamylase from rat liver

FEBS Letters ◽  
1984 ◽  
Vol 177 (1) ◽  
pp. 41-46 ◽  
Author(s):  
Peter Mclntyre ◽  
Lynda Graf ◽  
Julian Mercer ◽  
Gregory Peterson ◽  
Peter Hudson ◽  
...  
Keyword(s):  
Rat Liver ◽  
Ornithine Transcarbamylase ◽  
Mitochondrial Matrix ◽  
Matrix Enzyme

scholarly journals Inhibition in vitro of acyl-CoA dehydrogenases by 2-mercaptoacetate in rat liver mitochondria

1983 ◽  
Vol 215 (3) ◽  
pp. 457-464 ◽  
Author(s):  
F Bauché ◽  
D Sabourault ◽  
Y Giudicelli ◽  
J Nordmann ◽  
R Nordmann
Keyword(s):  
Rat Liver ◽  
Conformational Changes ◽  
Reaction Medium ◽  
Liver Mitochondria ◽  
Fatty Acyl ◽  
Rat Liver Mitochondria ◽  
Mitochondrial Matrix ◽  
Main Effect ◽  
Acyl Coa Dehydrogenases

In rat liver hypo-osmotically treated mitochondria, 2-mercaptoacetate inhibits respiration induced by palmitoyl-CoA, octanoate or butyryl-CoA only when the reaction medium is supplemented with ATP. Under this condition, NADH-stimulated respiration is not affected. In liver mitochondrial matrix, the presence of ATP is also required to observe a 2-mercaptoacetate-induced inhibition of acyl-CoA dehydrogenases tested with palmitoyl-CoA, butyryl-CoA or isovaleryl-CoA as substrate. As the oxidation of these substrates is also inhibited by the incubation medium resulting from the reaction of 2-mercaptoacetate with acetyl-CoA synthase, with conditions under which 2-mercaptoacetate has no effect, 2-mercaptoacetyl-CoA seems to be the likely inhibitory metabolite responsible for the effects of 2-mercaptoacetate. Kinetic experiments show that the main effect of the 2-mercaptoacetate-active metabolite is to decrease the affinities of fatty acyl-CoA dehydrogenases towards palmitoyl-CoA or butyryl-CoA and of isovaleryl-CoA dehydrogenase towards isovaleryl-CoA. Addition of N-ethylmaleimide to mitochondrial matrix pre-exposed to 2-mercaptoacetate results in the immediate reversion of the inhibitions of palmitoyl-CoA and isovaleryl-CoA dehydrogenations and in a delayed reversion of butyryl-CoA dehydrogenation. These results led us to conclude that (i) the ATP-dependent conversion of 2-mercaptoacetate into an inhibitory metabolite takes place in the liver mitochondrial matrix and (ii) the three fatty acyl-CoA dehydrogenases and isovaleryl-CoA dehydrogenase are mainly competitively inhibited by this compound. Finally, the present study also suggests that the inhibitory metabolite of 2-mercaptoacetate may bind non-specifically to, or induce conformational changes at, the acyl-CoA binding sites of these dehydrogenases.

scholarly journals The synthesis and hydrolysis of long-chain fatty acyl-coenzyme A thioesters by soluble and microsomal fractions from the brain of the developing rat

1976 ◽  
Vol 160 (2) ◽  
pp. 247-251 ◽  
Author(s):  
P J Brophy ◽  
D E Vance
Keyword(s):  
Fatty Acid ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Arachidic Acid ◽  
Fatty Acyl ◽  
Chain Elongation ◽  
Fatty Acid Chain ◽  
Specific Activities ◽  
Coa Hydrolase ◽  
Long Chain

1. The specific activities of long-chain fatty acid-CoA ligase (EC6.2.1.3) and of long-chain fatty acyl-CoA hydrolase (EC3.1.2.2) were measured in soluble and microsomal fractions from rat brain. 2. In the presence of either palmitic acid or stearic acid, the specific activity of the ligase increased during development; the specific activity of this enzyme with arachidic acid or behenic acid was considerably lower. 3. The specific activities of palmitoyl-CoA hydrolase and of stearoyl-CoA hydrolase in the microsomal fraction decreased markedly (75%) between 6 and 20 days after birth; by contrast, the corresponding specific activities in the soluble fraction showed no decline. 4. Stearoyl-CoA hydrolase in the microsomal fraction is inhibited (99%) by bovine serum albumin; this is in contrast with the microsomal fatty acid-chain-elongation system, which is stimulated 3.9-fold by albumin. Inhibition of stearoyl-CoA hydrolase does not stimulate stearoyl-CoA chain elongation. Therefore it does not appear likely that the decline in the specific activity of hydrolase during myelogenesis is responsible for the increased rate of fatty acid chain elongation. 5. It is suggested that the decline in specific activity of the microsomal hydrolase and to a lesser extent the increase in the specific activity of the ligase is directly related to the increased demand for long-chain acyl-CoA esters during myelogenesis as substrates in the biosynthesis of myelin lipids.

scholarly journals Hydrolysis of membrane phospholipids by phospholipases of rat liver lysosomes

1979 ◽  
Vol 182 (2) ◽  
pp. 599-606 ◽  
Author(s):  
Donald E. Richards ◽  
Robin F. Irvine ◽  
Rex M. C. Dawson
Keyword(s):  
Phospholipase C ◽  
Rat Liver ◽  
Lysosomal Enzymes ◽  
Microsomal Fraction ◽  
Water Soluble ◽  
Membrane Phospholipids ◽  
Liver Lysosomes ◽  
Rat Liver Lysosomes ◽  
Hydrolysis Of

(1) The hydrolysis of 32P- or myo-[2-3H]inositol-labelled rat liver microsomal phospholipids by rat liver lysosomal enzymes has been studied. (2) The relative rates of hydrolysis of phospholipids at pH4.5 are: sphingomyelin>phosphatidylethanolamine>phosphatidylcholine> phosphatidylinositol. (3) The predominant products of phosphatidylcholine and phosphatidylethanolamine hydrolysis are their corresponding lyso-compounds, indicating a slow rate of total deacylation. (4) Ca2+ inhibits the hydrolysis of all phospholipids, though only appreciably at high (>5mm) concentration. The hydrolysis of sphingomyelin is considerably less sensitive to Ca2+ than that of glycerophospholipids. (5) Analysis of the water-soluble products of phosphatidylinositol hydrolysis (by using myo-[3H]inositol-labelled microsomal fraction as a substrate) produced evidence that more than 95% of the product is phosphoinositol, which was derived by direct cleavage from phosphatidylinositol, rather than by hydrolysis of glycerophosphoinositol. (6) This production of phosphoinositol, allied with negligible lysophosphatidylinositol formation and a detectable accumulation of diacylglycerol, indicates that lysosomes hydrolyse membrane phosphatidylinositol almost exclusively in a phospholipase C-like manner. (7) Comparisons are drawn between the hydrolysis by lysosomal enzymes of membrane substrates and that of pure phospholipid substrates, and also the possible role of phosphatidylinositol-specific lysosomal phospholipase C in cellular phosphatidylinositol catabolism is discussed.

scholarly journals The hydrolysis of phosphatidylinositol by lysosomal enzymes of rat liver and brain

1978 ◽  
Vol 176 (2) ◽  
pp. 475-484 ◽  
Author(s):  
R F Irvine ◽  
N Hemington ◽  
R M C Dawson
Keyword(s):  
Phospholipase C ◽  
Rat Liver ◽  
Gel Filtration ◽  
Phosphatidyl Inositol ◽  
Temperature Stability ◽  
Enzyme Systems ◽  
Phosphatidic Acid Phosphatase ◽  
Bivalent Metals ◽  
Hydrolysis Of ◽  
Marked Specificity

1. Lysosomes from rat liver contain two enzymic systems for hydrolysing phosphatidyl-inositol: a deacylation via lysophosphatidylinositol producing glycerophosphoinositol and non-esterified fatty acid, and a phospholipase C-like cleavage into inositol 1-phosphate and diaclygycerol. 2. The separate enzyme systems involved can be distinguished by gel filtration, differential temperature-stability and the inhibitory action of detergents. 3. The enzyme systems both have pH optima at 4.8 and their attack on a pure phosphatidylinositol substrate is inhibited by many bivalent metals including Ca2+ and Mg2+, and cationic drugs. 4. Whereas the deacylation system will attack other glycerophospholipids, the phospholipase C shows a marked specificity towards phosphatidylinositol, although it will also slowly attach phosphatidylcholine with the liberation of phosphocholine. 5. Gel filtration and temperature-stability distinguish the phospholipase C from lysosomal phosphatidic acid phosphatase, but not from sphingomyelinase. 6. Evidence is presented that an EDTA-insensitive phospholipase C degrading phosphatidylinositol is present in rat brain.

scholarly journals Activation of a peroxisome-proliferating catabolite of cholic acid to its CoA ester

1993 ◽  
Vol 296 (1) ◽  
pp. 265-270 ◽  
Author(s):  
T Nishimaki-Mogami ◽  
A Takahashi ◽  
Y Hayashi
Keyword(s):  
Cholic Acid ◽  
Rat Liver ◽  
Microsomal Fraction ◽  
Liver Microsomes ◽  
Subcellular Fractionation ◽  
Fatty Acyl ◽  
Peroxisome Proliferator ◽  
Rat Liver Microsomes ◽  
Triton X ◽  
Long Chain

We have shown that a microbial cholic acid catabolite (4R)-4-(2,3,4,6,6a beta,7,8,9,9a alpha,9b beta-decahydro-6a beta-methyl-3-oxo- 1H-cyclopenta[f]quinolin-7 beta-yl)valeric acid (DCQVA), is a potent peroxisome proliferator. In this paper a possible key stage in DCQVA metabolism, the activation of DCQVA to its CoA ester, has been investigated in rat liver microsomes and particulate fractions. The microsomal reaction was dependent on CoA, ATP, DCQVA (0.2-1 mM) and protein content. The reaction was decreased by storage at 4 degrees C, preincubation of microsomes at 37 degrees C for 5 min, or inclusion of Triton X-100 in the reaction mixture. Such treatments also enhanced generation of long-chain fatty acyl-CoAs, as determined by h.p.l.c. analysis. The same effect was caused by exposing the microsomes to phospholipase A2, suggesting that endogenous fatty acids may compete with DCQVA for esterification with CoA. Subcellular fractionation of rat liver demonstrated that the activity of DCQVA-CoA synthesis was localized predominantly in the microsomal fraction, in contrast to long-chain fatty acyl-CoA synthetase, which was distributed among all particulate fractions. Administration of clofibrate of rats did not affect the distribution of DCQVA-CoA synthesis activity. In contrast to a 2-fold induction of long-chain fatty acyl-CoA synthetase by clofibrate treatment, the activity of DCQVA-CoA synthesis in the microsomal fraction decreased by 80%. These results suggest that DCQVA is activated by an enzyme distinct from long-chain fatty acyl-CoA synthetase. The resulting perturbation of fatty acid metabolism may be involved in the mechanism whereby DCQVA causes peroxisome proliferation.

scholarly journals Identity of purified monoacylglycerol lipase, palmitoyl-CoA hydrolase and aspirin-metabolizing carboxylesterase from rat liver microsomal fractions. A comparative study with enzymes purified in different laboratories

1985 ◽  
Vol 232 (2) ◽  
pp. 479-483 ◽  
Author(s):  
R Mentlein ◽  
R K Berge ◽  
E Heymann
Keyword(s):  
Rat Liver ◽  
Microsomal Fraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Monoacylglycerol Lipase ◽  
Enzyme Preparations ◽  
Nitrophenyl Phosphate ◽  
Liver Microsomal Fraction ◽  
Coa Hydrolase ◽  
Rat Liver Cytosol

Two purified carboxylesterases that were isolated from a rat liver microsomal fraction in a Norwegian and a German laboratory were compared. The Norwegian enzyme preparation was classified as palmitoyl-CoA hydrolase (EC 3.1.2.2) in many earlier papers, whereas the German preparation was termed monoacylglycerol lipase (EC 3.1.1.23) or esterase pI 6.2/6.4 (non-specific carboxylesterase, EC 3.1.1.1). Antisera against the two purified enzyme preparations were cross-reactive. The two proteins co-migrate in sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Both enzymes exhibit identical inhibition characteristics with Mg2+, Ca2+ and bis-(4-nitrophenyl) phosphate if assayed with the two substrates palmitoyl-CoA and phenyl butyrate. It is concluded that the two esterase preparations are identical. However, immunoprecipitation and inhibition experiments confirm that this microsomal lipase differs from the palmitoyl-CoA hydrolases of rat liver cytosol and mitochondria.

scholarly journals The action of phosphatidylinositol-specific phospholipases C on membranes

1976 ◽  
Vol 154 (1) ◽  
pp. 203-208 ◽  
Author(s):  
M G Low ◽  
J B Finean
Keyword(s):  
Phospholipase C ◽  
Rat Liver ◽  
Microsomal Fraction ◽  
Phosphatidyl Inositol ◽  
Detectable Effect ◽  
Erythrocyte Ghosts ◽  
Membrane Phospholipids ◽  
Liver Microsomal Fraction ◽  
Hydrolysis Of ◽  
Low Activity

A phospholipase C prepared from lymphocytes readily hydrolysed pure phosphatidyl-inositol but was relatively ineffective against phosphatidylinositol in erythrocyte “ghosts” and rat liver microsomal fraction and also against sonicated lipid extracts from these membranes. In contrast, a phospholipase C prepared from Staphylcoccus aureus readily hydrolysed phosphatidylinositol in sonicated lipid extracts but had only low activity against purified phosphatidylinositol. Unlike the enzyme from lymphocytes, the S. aureus phospholipase C did not require Ca2+ for its activity and was inhibited by cations. The previously reported specificity of this enzyme was confirmed by our observation of hydrolysis of approx. 75% of the phosphatidylinositol in ox, sheep and cat erythrocyte “ghosts” together with no detectable effect on the major erythrocyte membrane phospholipids. The phosphatidylinositol of rat liver microsomal fraction was hydrolysed only to a maximum of 15%. Some preliminary experiments showed that approx. 60% of the phosphatidylinositol of ox or sheep erythrocytes could be hydrolysed without causing substantial haemolysis.

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