Intraperoxisomal Localization of Very-Long-Chain Fatty Acyl-CoA Synthetase: Implication in X-Adrenoleukodystrophy

Department of Pediatrics, Medical University of South Carolina, Charleston, South Carolina, 29425
Experimental Cell Research (Impact Factor: 3.37). 03/2000; 254(2):309-320. DOI: 10.1006/excr.1999.4757

ABSTRACT X-Adrenoleukodystrophy (X-ALD) is a demyelinating disorder characterized by the accumulation of saturated very-long-chain (VLC) fatty acids (>C22:0) due to the impaired activity of VLC acyl-CoA synthetase (VLCAS). The gene responsible for X-ALD was found to code for a peroxisomal integral membrane protein (ALDP) that belongs to the ATP binding cassette superfamily of transporters. To understand the function of ALDP and how ALDP and VLCAS interrelate in the peroxisomal β-oxidation of VLC fatty acids we investigated the peroxisomal topology of VLCAS protein. Antibodies raised against a peptide toward the C-terminus of VLCAS as well as against the N-terminus were used to define the intraperoxisomal localization and orientation of VLCAS in peroxisomes. Indirect immunofluorescent and electron microscopic studies show that peroxisomal VLCAS is localized on the matrix side. This finding was supported by protease protection assays and Western blot analysis of isolated peroxisomes. To further address the membrane topology of VLCAS, Western blot analysis of total membranes or integral membranes prepared from microsomes and peroxisomes indicates that VLCAS is a peripheral membrane-associated protein in peroxisomes, but an integral membrane in microsomes. Moreover, peroxisomes isolated from cultured skin fibroblasts from X-ALD patients with a mutation as well as a deletion in ALDP showed a normal amount of VLCAS. The consequence of VLCAS being localized to the luminal side of peroxisomes suggests that ALDP may be involved in stabilizing VLCAS activity, possibly through protein–protein interactions, and that loss or alterations in these interactions may account for the observed loss of peroxisomal VLCAS activity in X-ALD.

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    • "It will be important to study the tissue and cellular pattern of ABCD1 expression to determine where and at which developmental period(s) the function of ALDP is necessary [16] [17]. These studies and a comparison between human and mouse might offer an explanation as to why the same biochemical deficiency can lead to an inflammatory demyelinating disorder in humans but not in mice. "
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