RESUMEN
In this report, we describe the first known patient with a deficiency of sterol carrier protein X (SCPx), a peroxisomal enzyme with thiolase activity, which is required for the breakdown of branched-chain fatty acids. The patient presented with torticollis and dystonic head tremor as well as slight cerebellar signs with intention tremor, nystagmus, hyposmia, and azoospermia. Magnetic resonance imaging showed leukencephalopathy and involvement of the thalamus and pons. Metabolite analyses of plasma revealed an accumulation of the branched-chain fatty acid pristanic acid, and abnormal bile alcohol glucuronides were excreted in urine. In cultured skin fibroblasts, the thiolytic activity of SCPx was deficient, and no SCPx protein could be detected by western blotting. Mutation analysis revealed a homozygous 1-nucleotide insertion, 545_546insA, leading to a frameshift and premature stop codon (I184fsX7).
Asunto(s)
Proteínas Portadoras/genética , Demencia Vascular/diagnóstico , Distonía/diagnóstico , Polineuropatías/diagnóstico , Tortícolis/diagnóstico , Adulto , Proteínas Portadoras/sangre , Codón sin Sentido , Demencia Vascular/genética , Distonía/genética , Ácidos Grasos/sangre , Mutación del Sistema de Lectura , Glucurónidos/orina , Humanos , Imagen por Resonancia Magnética , Masculino , Polineuropatías/genética , Puente/patología , Síndrome , Tálamo/patología , Tortícolis/genéticaRESUMEN
Phytanoyl-CoA hydroxylase (PhyH) catalyzes the conversion of phytanoyl-CoA to 2-hydroxyphytanoyl-CoA, which is the first step in the phytanic acid alpha-oxidation pathway. Recently, several studies have shown that in humans, phytanic acid alpha-oxidation is localized in peroxisomes. In rat, however, the alpha-oxidation pathway has been reported to be mitochondrial. In order to clarify this differential subcellular distribution, we have studied the rat PhyH protein. We have purified PhyH from rat liver to apparent homogeneity as judged by SDS-PAGE. Sequence analysis of two PhyH peptide fragments allowed cloning of the rat PHYH cDNA encoding a 38. 6 kDa protein. The deduced amino acid sequence revealed strong homology to human PhyH including the presence of a peroxisome targeting signal type 2 (PTS2). Heterologous expression of rat PHYH in Saccharomyces cerevisiae yielded a 38.6 kDa protein whereas the PhyH purified from rat liver had a molecular mass of 35 kDa. This indicates that PhyH is probably processed in rat by proteolytic removal of a leader sequence containing the PTS2. This type of processing has been reported in several other peroxisomal proteins that contain a PTS2. Subcellular localization studies using equilibrium density centrifugation showed that PhyH is indeed a peroxisomal protein in rat. The finding that PhyH is peroxisomal in both rat and humans provides strong evidence against the concept of a differential subcellular localization of phytanic acid alpha-oxidation in rat and human.
Asunto(s)
Hígado/enzimología , Oxigenasas de Función Mixta/aislamiento & purificación , Secuencia de Aminoácidos , Animales , Radioisótopos de Carbono , Centrifugación por Gradiente de Densidad , Clonación Molecular , ADN Complementario/biosíntesis , Humanos , Ácidos Cetoglutáricos/metabolismo , Masculino , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Datos de Secuencia Molecular , Oxidación-Reducción , Peroxisomas/enzimología , Ácido Fitánico/metabolismo , Ratas , Ratas Wistar , Saccharomyces cerevisiae/genética , Homología de Secuencia de Aminoácido , LevadurasRESUMEN
To study the putative role of human carnitine octanoyltransferase (COT) in the beta-oxidation of branched-chain fatty acids, we identified and cloned the cDNA encoding human COT and expressed it in the yeast Saccharomyces cerevisiae. Enzyme activity measurements showed that COT efficiently converts one of the end products of the peroxisomal beta-oxidation of pristanic acid, 4, 8-dimethylnonanoyl-CoA, to its corresponding carnitine ester. Production of the carnitine ester of this branched/medium-chain acyl-CoA within the peroxisome is required for its transport to the mitochondrion where further beta-oxidation occurs. In contrast, 4, 8-dimethylnonanoyl-CoA is not a substrate for carnitine acetyltransferase, another acyltransferase localized in peroxisomes, which catalyzes the formation of carnitine esters of the other products of pristanic acid beta-oxidation, namely acetyl-CoA and propionyl-CoA. Our results shed new light on the function of COT in fatty acid metabolism and point to a crucial role of COT in the beta-oxidation of branched-chain fatty acids.