Purification of human very-long-chain acyl-coenzyme A dehydrogenase and characterization of its deficiency in seven patients.

Abstract

Mitochondrial very-long-chain acyl-coenzyme A dehydrogenase (VLCAD) was purified from human liver. The molecular masses of the native enzyme and the subunit were estimated to be 154 and 70 kD, respectively. The enzyme was found to catalyze the major part of mitochondrial palmitoylcoenzyme A dehydrogenation in liver, heart, skeletal muscle, and skin fibroblasts (89-97, 86-99, 96-99, and 78-87%, respectively).Skin fibroblasts from 26 patients suspected of having a disorder of mitochondrial a-oxidation were analyzed for VLCAD protein using immunoblotting, and 7 of them contained undetectable or trace levels of the enzyme. The seven deficient fibroblast lines were characterized by measuring acyl-coenzyme A dehydrogenation activities, overall palmitic acid oxidation, and VLCAD protein synthesis using pulse-chase, further confirming the diagnosis of VLCAD deficiency. These results suggested the heterogenous nature of the mutations causing the deficiency in the seven patients.Clinically, all patients with VLCAD deficiency exhibited cardiac disease. At least four of them presented with hypertrophic cardiomyopathy. This frequency (> 57%) was much higher than that observed in patients with other disorders of mitochondrial long-chain fatty acid oxidation that may be accompanied by cardiac disease in infants. (J. Clin. Invest. 1995Invest. . 95:2465Invest. -2473

Acute kidney injury induced by protein-overload nephropathy down-regulates gene expression of hepatic cerebroside sulfotransferase in mice, resulting in reduction of liver and serum sulfatides

“…Whole-tissue lysates (liver and kidney) were prepared from each mouse as described previously [14][15][16][17]. Protein concentrations were measured using the BCA Protein Assay Kit (Pierce Biotechnology, Rockford, IL, USA) [18].…”

Activation of PPAR<i>α</i> by Fatty Acid Accumulation Enhances Fatty Acid Degradation and Sulfatide Synthesis

“…VLCAD and LCAD catalyze reactions that generate long-chain fatty acyl-CoAs such as palmitoylCoA. Notably, VLCAD contributes more than 90% of palmitoyl-CoA dehydrogenation activity in human heart and liver tissue (Aoyama et al 1995b). VLCAD deficiency is associated with the accumulation of fat in multiple organs and tissues, liver dysfunction, cardiomyopathy, cardiomegaly, muscle weakness, and fasting coma with high frequency (Aoyama et al 1993(Aoyama et al , 1995a.…”

“…Notably, VLCAD contributes more than 90% of palmitoyl-CoA dehydrogenation activity in human heart and liver tissue (Aoyama et al 1995b). VLCAD deficiency is associated with the accumulation of fat in multiple organs and tissues, liver dysfunction, cardiomyopathy, cardiomegaly, muscle weakness, and fasting coma with high frequency (Aoyama et al 1993(Aoyama et al , 1995a. Additionally, mutation analyses have identified a genotype-phenotype relationship for VLCAD deficiency (Aoyama et al 1995a;Souri et al 1996;Andresen et al 1999;Gregersen et al 2001).…”

“…VLCAD deficiency is associated with the accumulation of fat in multiple organs and tissues, liver dysfunction, cardiomyopathy, cardiomegaly, muscle weakness, and fasting coma with high frequency (Aoyama et al 1993(Aoyama et al , 1995a. Additionally, mutation analyses have identified a genotype-phenotype relationship for VLCAD deficiency (Aoyama et al 1995a;Souri et al 1996;Andresen et al 1999;Gregersen et al 2001). Mutation analysis of six patients has revealed homozygous loss of 35 amino acids in patients 1 and 6 (Aoyama et al 1995a), homozygous loss of glutamate-130 in patient 2 (Souri et al 1996), point mutation of lysine-382 to glutamine and loss of glutamate-130 in patient 3 (Souri et al 1996), homozygous loss of lysine-299 in patient 4 (Souri et al 1996), and loss of 45 amino acids and point mutation of arginine-613 to tryptophan in patient 5 (Souri et al 1996).…”

“…Additionally, mutation analyses have identified a genotype-phenotype relationship for VLCAD deficiency (Aoyama et al 1995a;Souri et al 1996;Andresen et al 1999;Gregersen et al 2001). Mutation analysis of six patients has revealed homozygous loss of 35 amino acids in patients 1 and 6 (Aoyama et al 1995a), homozygous loss of glutamate-130 in patient 2 (Souri et al 1996), point mutation of lysine-382 to glutamine and loss of glutamate-130 in patient 3 (Souri et al 1996), homozygous loss of lysine-299 in patient 4 (Souri et al 1996), and loss of 45 amino acids and point mutation of arginine-613 to tryptophan in patient 5 (Souri et al 1996). Immunoblot analysis using an anti-human VLCAD antibody demonstrated the absence of mature VLCAD protein in all cell lines (Aoyama et al 1995b).…”