Identification of the peroxisomal β-oxidation enzymes involved in the degradation of long-chain dicarboxylic acids

Dicarboxylic acids (DCAs) are -oxidation products of monocarboxylic acids. After activation by a dicarboxylyl-CoA synthetase, the dicarboxylyl-CoA esters are shortened via -oxidation. Although it has been studied extensively where this -oxidation process takes place, the intracellular site of DCA oxidation has remained controversial. Making use of fibroblasts from patients with defined mitochondrial and peroxisomal fatty acid oxidation defects, we show in this paper that peroxisomes, and not mitochondria, are involved in the -oxidation of C16DCA. Additional studies in fibroblasts from patients with X-linked adrenoleukodystrophy, straight-chain acyl-CoA oxidase (SCOX) deficiency, d -bifunctional protein (DBP) deficiency, and rhizomelic chondrodysplasia punctata type 1, together with direct enzyme measurements with human recombinant l -bifunctional protein (LBP) and DBP expressed in a fox2 deletion mutant of Saccharomyces cerevisiae , show that the main enzymes involved in -oxidation of C16DCA are SCOX, both LBP and DBP, and sterol carrier protein X, possibly together with the classic 3-ketoacyl-CoA thiolase. This is the first indication of a specific function for LBP, which has remained elusive until now. —Ferdinandusse, S., S. Denis, C. W. T. van Roermund, R. J. A. Wanders, and G. Dacremont. Identification of the peroxisomal -oxidation enzymes involved in the degradation of long-chain dicarboxylic acids. J. Lipid Res. 2004. 45: 1104–1111. Supplementary key words straight-chain acyl-CoA oxidase • l -bifunctional protein • d -bifunctional protein Dicarboxylic acids (DCAs) are -oxidation products of monocarboxylic acids. Fatty acids are first converted into -hydroxymonocarboxylic acids by a microsomal cytochrome P450 and then further oxidized to -ketomonocarboxylic acids and finally DCAs through the sequential action of cytosolic long-chain alcohol and aldehyde dehydrogenases. After activation by a dicarboxylyl-CoA synthetase present in microsomes, at least in rat liver (1), the dicarboxylyl-CoA esters are shortened via -oxidation ( Fig. 1 ). Where this -oxidation of dicarboxylyl-CoA esters takes place, in peroxisomes or mitochondria, has been studied extensively in rats. However, no unequivocal answer has been found for this question. In vivo studies in rats have shown that both mitochondria and peroxisomes are involved in the catabolism of DCAs (2, 3), and several in vitro studies with isolated organelles have indicated that both mitochondria and peroxisomes are capable of oxidizing DCAs (4–6). However, the contribution of the two organelles differed among these studies, with some authors concluding that the oxidation of DCAs is mainly peroxisomal (6, 7) and others concluding that this is an exclusively peroxisomal process (3, 8). The role of mitochondria seems to be substrate and tissue dependent (5) and is probably determined by the carnitine dicarboxylyltransferase activity (4–6, 8), because intact mitochondria are active with dicarboxylylcarnitine esters and, in addition, mitochondria permeabilized with digitonin are capable of oxidizing dicarboxylylCoA esters (3, 5). The finding of dicarboxylic aciduria in patients with a fatty acid oxidation disorder does not make it clear either whether DCAs are oxidized in peroxisomes or mitochondria. Both patients with a peroxisomal fatty acid oxidation Abbreviations: ALDP, adrenoleukodystrophy protein; BCOX, branched-chain acyl-CoA oxidase; CACT, carnitine acylcarnitine translocase; CPT, carnitine palmitoyltransferase; DBP, d -bifunctional protein; DCA, dicarboxylic acid; LBP, l -bifunctional protein; MTP, mitochondrial trifunctional protein; PBD, peroxisome biogenesis disorder; RCDP, rhizomelic chondrodysplasia punctata; SCOX, straight-chain acyl-CoA oxidase; SCPx, sterol carrier protein X; VLCAD, very longchain acyl-CoA dehydrogenase; XALD, X-linked adrenoleukodystrophy. 1 To whom correspondence should be addressed. e-mail: [email protected] Manuscript received 16 December 2003 and in revised form 19 March 2004. Published, JLR Papers in Press, April 1, 2004. DOI 10.1194/jlr.M300512-JLR200 by gest, on A uust 8, 2017 w w w .j.org D ow nladed fom