Induction of a Novel Long‐Chain Acyl‐CoA Hydrolase in Rat Liver by Administration of Peroxisome Proliferators

Abstract

The activity of long-chain acyl-CoA hydrolase in rat liver was increased by the administration of peroxisome proliferators, such as ethyl p-chlorophenoxyisobutyrate, di(2-ethy1hexyl)phthalate or acetylsalicylic acid. The induced activity was mainly confined 'in the soluble fluid after the subcellular fractionation. The enzyme was purified nearly to homogeneity from livers of rats treated with di(2-ethy1hexyl)phthalate. The specific activity of the final preparation was 247 pmol palmitoyl-CoA hydrolyzed min-' mg protein-'. The molecular weight of the native enzyme was estimated to be 150000 by gel filtration and that of the subunits was 41 000 by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The activity of the enzyme was not increased but inhibited by bovine serum albumin or Triton X-100. The molecular. and catalytic properties of the enzyme suggest that the induced enzyme was different from mitochondria1 and microsomal long-chain acyl-CoA hydrolases in liver.Long-chain fatty acyl-CoA hydrolases are present in various mammalian tissues [I -81. In rat liver, the enzymes are mainly localized in mitochondria and microsomes [2,3]. These enzymes in both particulates have been purified and characterized [4,5]. During the course of our study on the effect of di(2-ethylhexyl)phthalate, which is a potent inducer of peroxisomal /%oxidation system [9], on activities of acylCoA dehydrogenases in rat liver mitochondria, we noticed that the reducing activity of 2,6-dichloroindophenol in the presence of palmitoyl-CoA was markedly induced by the administration of di(2-ethylhexy1)phthalate. The increased rate of the dye reduction was due to the elevated activity of long-chain acyl-CoA hydrolase.In the present study, the induced hydrolase was found in the soluble fraction after the subcellular fractionation of rat livers. The enzyme was purified to a nearly homogeneous preparation and found to be a new enzyme with respect to its molecular and catalytic properties. MATERIALS AND METHODS MaterialsAcetyl-CoA and butyryl-CoA were prepared from the corresponding acid anhydrides. Other acyl-CoA derivatives were synthesized by the mixed anhydride method [lo] and purified as described previously [ l l , 121. CoA, bovine serum albumin and cytochrome c were from Sigma. Enzymes and coenzymes were purchased froin Boehringer. Analytical grades of fatty acids and other reagents were commercial products.Enzymes. Acyl-CoA oxidase (EC 1.3.-.-); adenylate kinase (EC 2.7.4.3); dlcohol dehydrogenase (EC 1.1.1.1); aldolase (EC 4.1.2.13); catalase (EC 1.11.1.6); citrate synthase (EC 4.1.3.7); glutamate dehydrogenase (EC 1.4.1.3); horseradish peroxidase (EC 1.11.1.7); lactate dehydrogendse (EC 1.1.1.27); long-chain acyl-CoA hydrolase (EC 3.1.2.2); malate dehydrogenase (EC 1.1.1.37) ; NADPH-cytochrome c reductase (EC 1.6.2.4); phosphorylase (EC 2.4.1.1). Animals and Subcellular FractionationMale Wistar rats (200-250 g) were fed a powdered diet (Oriental M, Tokyo) containing 2 (w/w) di(2-ethylhexy1)-phthalate for 2-4 weeks, when they were us...

Morphological Changes of Mitochondria in the Hepatocytes Induced by Administration of a Large Amount of Di (2-ethylhexyl) phthalate (DEHP) to Cynomolgus Monkeys (Macaca fascicularis)

“…Di (2-ethylhexyl) phthalate (DEHP), a plasticizer, has been reported to induce an increase in rat peroxisomes 1 ; however, no such increase was induced in cynomolgus monkeys (Macaca fascicularis) when DEPH was administered at 500 mg/kg/day for 14 or 21 days 2,3 . In addition, marmoset peroxisomes were not increased by administration of DEHP at 2,500 mg/kg/day for 13 weeks 4 .…”

Structural basis for recruitment of tandem hotdog domains in acyl-CoA thioesterase 7 and its role in inflammation

“…The absorbance at 232 nm (13) was monitored immediately after adding the substrate and followed for 3 min at 20-s intervals. The molar absorption coefficient, 232 (4,250 M Ϫ1 cm Ϫ1 ) was used to calculate cleavage of the thioester bond (36). GraFit was used to plot the data and calculate maximum velocity and Michaelis-Menten constants.…”

Peroxisome Proliferators Differentially Regulate Long‐chain Acyl‐CoA Thioesterases in Rat Liver

“…The fatty-acid synthesis-terminating enzymes acting on fatty acyl thioesters associated to a pantetheine moiety of the acyl-carrierprotein domain of fatty acid synthase (thioesterase I and thioesterase 11) have been most thoroughly studied . Several long-chain acyl-CoA thioesterases are responsible for the activity in rat liver and some of them have been purified [14,. The enzyme isolated from microsomes appears to be clearly different from other known thioesterases and its close relationship to members of the rat liver microsomal carboxylesterase multi-gene family has been established [24,251.…”