Role of branched-chain 2-oxo acid dehydrogenase and pyruvate dehydrogenase in 2-oxobutyrate metabolism.
Role of branched-chain 2-oxo acid dehydrogenase and pyruvate dehydrogenase in 2-oxobutyrate metabolism.
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支链2-含氧酸脱氢酶和丙酮酸脱氢酶在2-氧代丁酸代谢中的作用。
DOI：
10.1042/bj2340295
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发表时间：
1986
期刊：
影响因子：
--
通讯作者：
Harris,RA
中科院分区：
文献类型：
--
作者：
Paxton,R;Scislowski,PW;Davis,EJ;Harris,RA
纯化的支链 2-氧代酸脱氢酶 (BCODH) 和丙酮酸脱氢酶 (PDH) 对于 2-氧代丁酸的表观 Km 值 (μM) 分别为 26 和 114，相对 Vmax。 （3-甲基-2-氧代丁酸和丙酮酸的 Vmax.%）分别为 38% 和 45%。大鼠肝脏、肾脏、心脏和骨骼肌线粒体提取物中两种复合物的磷酸化状态均显示出影响 2-氧代丁酸的氧化脱羧作用。将 BCODH 抑制性抗体和 PDH（3-氟丙酮酸）抑制剂与线粒体提取物一起使用，以确定两种复合物对 2-氧代丁酸氧化脱羧的相对贡献。根据上面给出的动力学常数和两种复合物的活性计算出的线粒体提取物中 2-氧代丁酸脱羧速率与测量速率相同。大鼠肝脏线粒体提取物的羟基磷灰石色谱仅显示两个 2-氧代丁酸氧化脱羧峰，其中一个峰与 PDH 相关，另一个峰与 BCODH 相关。对各种 2-含氧酸的竞争研究表明，与心脏或骨骼肌提取物相比，肝脏线粒体提取物具有不同的抑制模式。我们得出结论，两种线粒体内复合物均负责 2-氧代丁酸的氧化脱羧。然而，根据动力学分析、两种复合物的活性或磷酸化状态、竞争研究以及丙酮酸、2-氧代丁酸和支链 2-含氧酸的表观生理浓度，BCODH 可能是更重要的复合物，特别是在肝脏中。
Purified branched-chain 2-oxo acid dehydrogenase (BCODH) and pyruvate dehydrogenase (PDH) had apparent Km values (microM) for 2-oxobutyrate of 26 and 114, with a relative Vmax. (% of Vmax. for 3-methyl-2-oxobutyrate and pyruvate) of 38 and 45% respectively. The phosphorylation state of both complexes in extracts of mitochondria from rat liver, kidney, heart and skeletal muscle was shown to influence oxidative decarboxylation of 2-oxobutyrate. Inhibitory antibodies to BCODH and an inhibitor of PDH (3-fluoropyruvate) were used with mitochondrial extracts to determine the relative contribution of both complexes to oxidative decarboxylation of 2-oxobutyrate. Calculated rates of 2-oxobutyrate decarboxylation in mitochondrial extracts, based on the kinetic constants given above and the activities of both complexes, were the same as the measured rates. Hydroxyapatite chromatography of extracts of mitochondria from rat liver revealed only two peaks of oxidative decarboxylation of 2-oxobutyrate, with one peak associated with PDH and the other with BCODH. Competition studies with various 2-oxo acids revealed a different inhibition pattern with mitochondrial extracts from liver compared with those from heart or skeletal muscle. We conclude that both intramitochondrial complexes are responsible for oxidative decarboxylation of 2-oxobutyrate. However, the BCODH is probably the more important complex, particularly in liver, on the basis of kinetic analyses, activity or phosphorylation state of both complexes, competition studies, and the apparent physiological concentration of pyruvate, 2-oxobutyrate and the branched-chain 2-oxo acids.