Branched Chain Amino Acid Metabolism

描述(申请人提供)：氨基酸除了作为蛋白质组成单元的作用外，还具有重要的营养信号作用。支链氨基酸亮氨酸是一种营养信号，刺激靶组织蛋白质合成增加，它支持我们的假设，即亮氨酸信号与亮氨酸代谢有关。分解代谢的第一步是由支链转氨酶同工酶(BCAT)催化的可逆转氨化反应。我们已经解析了人类线粒体BCAT(HBCATm)的几个结构，并发现它包含一个氧化还原活性的二硫醇/二硫键中心。CXXC中心对活性的氧化还原相关调节表明，半胱氨酸残基是可能在体内发挥作用的过氧化传感机制的一部分。我们发现hBCATm可以与BCKD络合物结合，并且这种相互作用是氧化还原敏感的。HBCATm和线粒体BCKD复合体之间的联系表明，BCAA分解代谢酶形成了称为代谢子的超分子复合体。我们将使用BCAT蛋白作为“诱饵”来验证蛋白质网控制支链氨基酸代谢的假设，以确定支链氨基酸代谢的蛋白质成分。BCAT蛋白网络的确定，特别是神经元特异性胞浆同工酶(BCATC)和新发现的人BCATm的一种选择性剪接形式(HBCATmsp)的确定，可能有助于鉴定这些蛋白的新功能。支链α-酮酸脱氢酶复合体(BCKD)是支链α-酮酸脱氢酶复合体催化BCA氧化的限速步骤。BCKD活性由ELA亚基上特定丝氨酸残基的磷酸化和去磷酸化来调节。我们将确定BCATm在亮氨酸信号中的作用，即亮氨酸或KIC是激活大鼠体内亮氨酸敏感组织中mTOR信号通路的信号。使用一种新开发的快速而简单的测试系统来测量BCKD的急性调节，将在体内和在β细胞模型中确定BCATm的作用以及mTOR信号通路的亮氨酸激活和BCKD复合体(代谢)之间的联系。我们希望，这项提案中提出的想法将为氨基酸代谢领域带来新的思维方式，并将导致令人兴奋的新研究方向。

DESCRIPTION (provided by applicant): In addition to their role as protein building blocks, amino acids have a significant role as nutritional signals. The BCAA leucine is a nutrient signal that stimulates increased protein synthesis in target tissues, and it Js our hypothesis that leucine signaling is linked to leucine metabolism. The first step in the catabolic pathway is reversible transamination catalyzed by the branched chain aminotransferase isozymes (BCAT). We have solved several structures of the human mitochondrial BCAT (hBCATm) and discovered that it contains a redox-active dithiol/disulfide center. The redox-linked regulation of activity by the CXXC center demonstrates that the cysteine residues are part of a peroxide-sensing mechanism that may have an in vivo role. We have discovered that hBCATm can associate with the BCKD complex and that this interaction is redox sensitive. The association between hBCATm and the mitochondrial BCKD complex suggests that BCAA catabolic enzymes form supramolecular complex called a metabolon. We will test the hypothesis that a protein web controls BCAA metabolism using the BCAT proteins as "bait" to identify protein components of the BCAA metabolon. Determination of the protein web for the BCAT, particularly of the neuron-specific cytosolic isozyme (BCATc) and a newly recognized alternatively spliced form of human BCATm (hBCATmsp), is likely to allow identification of new functions for these proteins. The rate-limiting step in BCAAoxidation is catalyzed in mitochondria by the branched-chain alpha-keto acid dehydrogenase enzyme complex (BCKD). BCKD activity is regulated by phosphorylation and dephosphorylation of a specific serine residue on the Ela subunit. We will determine the role of BCATm in leucine signaling, that is, whether leucine or KIC is the signal that activates the mTOR-signaling pathway in leucine-sensitive tissues in vivo in the rat. Using a newly developed rapid and simple test system to measure acute regulation of BCKD, the role of BCATm and link between leucine activation of the mTOR signaling pathway and BCKD complex (metabolism) will be determined in vivo and in a beta-cell model. It is our hope that the ideas put forth in this proposal will bring new ways of thinking into the field of amino acid metabolism and will lead to exciting new research directions.