The overall objective is to establish the mechanisms responsible for the regulation of the oxidation of branched-chain amino acids by mammalian cells. Studies will be conducted with purified branched-chain Alpha-ketoacid dehydrogenase (BCKDH) complex, purified BCKDH kinase, purified BCKDH phosphatase, isolated hepatocytes, isolated myocytes, perfused rat heart, isolated liver and heart mitochondria, and intact animals. The working hypothesis is that covalent modification by phosphorylation-dephosphorylation is an important mechanism for regulation of BCKDH. Studies will be conducted to further characterize the BCKDH complex purified in this laboratory from rabbit liver, to isolate and characterize the phosphatase responsible for dephosphorylation and activation of BCKDH to further investigate with intact animals the effects of various nutritional and hormonal states as well as specific drugs on the activity and phosphorylation state of BCKDH in various tissues, and finally to determine with isolated hepatocytes, myocytes, and mitochondria whether the phosphorylation state of the BCKDH is subject to acute hormonal control. The detailed specific aims include to establish (a) molecular weight and subunit organization of BCKDH; (b) the number and amino acid sequences of the phosphorylation sites in the 47,000 Mr Alpha-subunit; (c) the effectiveness of various Alpha-chloro- and Alpha-ketoacids as inhibitors of BCKDH kinase; (d) the role of Ca++ as an inhibitor of BCKDH kinase; (e) the effects of fasting, obesity, diabetes, and protein deprivation on the phosphorylation state of BCKDH in liver, heart, muscle, kidney, and adipose tissue; and (f) whether the phosphorylation state of BCKDH is subject to hormonal control. These studies should help explain altered branched-chaim amino acid metabolism known to occur in numerous diseases and metabolic conditions, including diabetes, obesity, maple syrup urine disease, phenylketonuria, kwashiorkor, and cancer.
| Shimomura, Yoshiharu; Honda, Takashi; Shiraki, Makoto et al. (2006) Branched-chain amino acid catabolism in exercise and liver disease. J Nutr 136:250S-3S |
| Joshi, Mandar A; Jeoung, Nam Ho; Obayashi, Mariko et al. (2006) Impaired growth and neurological abnormalities in branched-chain alpha-keto acid dehydrogenase kinase-deficient mice. Biochem J 400:153-62 |
| Harris, Robert A; Joshi, Mandar; Jeoung, Nam Ho et al. (2005) Overview of the molecular and biochemical basis of branched-chain amino acid catabolism. J Nutr 135:1527S-30S |
| Harris, Robert A; Joshi, Mandar; Jeoung, Nam Ho (2004) Mechanisms responsible for regulation of branched-chain amino acid catabolism. Biochem Biophys Res Commun 313:391-6 |
| Obayashi, Mariko; Shimomura, Yoshiharu; Nakai, Naoya et al. (2004) Estrogen controls branched-chain amino acid catabolism in female rats. J Nutr 134:2628-33 |
| Kobayashi, Rumi; Murakami, Taro; Obayashi, Mariko et al. (2002) Clofibric acid stimulates branched-chain amino acid catabolism by three mechanisms. Arch Biochem Biophys 407:231-40 |
| Harris, R A; Kobayashi, R; Murakami, T et al. (2001) Regulation of branched-chain alpha-keto acid dehydrogenase kinase expression in rat liver. J Nutr 131:841S-845S |
| Obayashi, M; Sato, Y; Harris, R A et al. (2001) Regulation of the activity of branched-chain 2-oxo acid dehydrogenase (BCODH) complex by binding BCODH kinase. FEBS Lett 491:50-4 |
| Baker, L J; Dorocke, J A; Harris, R A et al. (2001) The crystal structure of yeast thiamin pyrophosphokinase. Structure 9:539-46 |
| Timm, D E; Liu, J; Baker, L J et al. (2001) Crystal structure of thiamin pyrophosphokinase. J Mol Biol 310:195-204 |
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