title: Molecular cloning of rat acss3 and characterization of mammalian propionyl-CoA synthetase in the liver mitochondrial matrix.
abstract: Among the three acyl-CoA synthetase short-chain family members (ACSS), ACSS3 is poorly characterized. To characterize ACSS3, we performed molecular cloning and protein expression of rat acss3 and determined its intracellular localization, tissue distribution, and substrate specificity. Transient expression of rat ACSS3 in HeLa cells resulted in a 10-fold increase of acetyl-CoA synthetase activity compared with that in control cells. The acss3 transcripts are expressed in a wide range of tissues, with the highest levels observed in liver tissue followed by kidney tissue. Subcellular fractionation using liver tissue showed that ACSS3 is localized into the mitochondrial matrix. Among the short-chain fatty acids examined, recombinant ACSS3, purified from Escherichia coli cells transformed with the plasmid containing rat acss3, preferentially utilized propionate with a KM value of 0.19 mM. Knockdown of acss3 in HepG2 cells resulted in a significant decrease of ACSS3 expression level and propionyl-CoA synthetase activity in cell lysates. Levels of ACSS3 in the liver and the activity of propionyl-CoA synthetase in the mitochondria were significantly increased by fasting. These results suggested that ACSS3 is a liver mitochondrial matrix enzyme with high affinity to propionic acid, and its expression level is upregulated under ketogenic conditions.
authors.affiliation: Department of Nutritional Science Faculty of Health and Welfare Science, Okayama Prefectural University, 111 Kuboki, Soja-shi, Okayama 719-1197, Japan.
authors.forename: Yukihiro
authors.initials: Y
authors.lastname: Yoshimura
authors.affiliation: Department of Nutritional Science Faculty of Health and Welfare Science, Okayama Prefectural University, 111 Kuboki, Soja-shi, Okayama 719-1197, Japan.
authors.forename: Aya
authors.initials: A
authors.lastname: Araki
authors.affiliation: Department of Nutritional Science Faculty of Health and Welfare Science, Okayama Prefectural University, 111 Kuboki, Soja-shi, Okayama 719-1197, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Maruta
authors.affiliation: Department of Nutritional Science Faculty of Health and Welfare Science, Okayama Prefectural University, 111 Kuboki, Soja-shi, Okayama 719-1197, Japan.
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Takahashi
authors.affiliation: Department of Nutritional Science Faculty of Health and Welfare Science, Okayama Prefectural University, 111 Kuboki, Soja-shi, Okayama 719-1197, Japan.
authors.forename: Hiromi
authors.initials: H
authors.lastname: Yamashita
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D003066:Coenzyme A Ligases / Q000235:genetics* / Q000378:metabolism*; D006367:HeLa Cells; D056945:Hep G2 Cells; D006801:Humans; D008930:Mitochondria, Liver / Q000201:enzymology* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: C000618751:ACSS3 protein, rat; D003066:Coenzyme A Ligases; C068072:propionate - CoA ligase
keywords: molecular cloning; propionate metabolism; propionyl-CoA synthetase; subcellular fractionation; substrate specificity