Gudi R, Bowker-Kinley M M, Kedishvili N Y, Zhao Y, Popov K M
Department of Biochemistry and Molecular Biology, Indiana University, School of Medicine, Indianapolis 46202-5122, USA.
J Biol Chem. 1995 Dec 1;270(48):28989-94. doi: 10.1074/jbc.270.48.28989.
Recent evidence from this laboratory indicates that at least two isoenzymic forms of pyruvate dehydrogenase kinase (PDK1 and PDK2) may be involved in the regulation of enzymatic activity of mammalian pyruvate dehydrogenase complex by phosphorylation (Popov, K.M., Kedishvili, N.Y., Zhao, Y., Gudi, R., and Harris, R.A. (1994) J. Biol. Chem. 269, 29720-29724). The present study was undertaken to further explore the diversity of the pyruvate dehydrogenase kinase gene family. Here we report the deduced amino acid sequences of three isoenzymic forms of PDK found in humans. In terms of their primary structures, two isoenzymes identified in humans correspond to rat PDK1 and PDK2, whereas a third gene (PDK3) encodes for a new isoenzyme that shares 68% and 67% of amino acid identities with PDK1 and PDK2, respectively. PDK3 cDNA expressed in Eschierichia coli directs the synthesis of a polypeptide with a molecular mass of approximately 45,000 Da that possesses catalytic activity toward kinase-depleted pyruvate dehydrogenase. PDK3 appears to have the highest specific activity among the three isoenzymes tested as recombinant proteins. Tissue distribution of all three isoenzymes of human PDK was characterized by Northern blot analysis. The highest amount of PDK2 mRNA was found in heart and skeletal muscle, the lowest amount in placenta and lung. Brain, kidney, pancreas, and liver expressed an intermediate amount of PDK2 (brain > kidney = pancreas > liver). The tissue distribution of PDK1 mRNA differs markedly from PDK2. The message for PDK1 was expressed predominantly in heart with only modest levels of expression in other tissues (skeletal muscle > liver > pancreas > brain > placenta = lung > kidney). In contrast to PDk1 and PDK2, which are expressed in all tissues tested, the message for PDK3 was found almost exclusively in heart and skeletal muscle, indicating that PDK3 may serve specialized functions characteristic of muscle tissues. In all tissues tested thus far, the level of expression of PDK2 mRNA was essentially higher than that of PDK1 and PDK3, consistent with the idea that PDK2 is a major isoenzyme responsible for regulation of pyruvate dehydrogenase in human tissues.
该实验室最近的证据表明，丙酮酸脱氢酶激酶至少有两种同工酶形式（PDK1和PDK2）可能参与通过磷酸化作用对哺乳动物丙酮酸脱氢酶复合体酶活性的调节（波波夫，K.M.，凯迪什维利，N.Y.，赵，Y.，古迪，R.，和哈里斯，R.A.（1994年）《生物化学杂志》269，29720 - 29724）。本研究旨在进一步探索丙酮酸脱氢酶激酶基因家族的多样性。在此我们报告在人类中发现的三种丙酮酸脱氢酶激酶同工酶形式的推导氨基酸序列。就其一级结构而言，在人类中鉴定出的两种同工酶分别对应大鼠的PDK1和PDK2，而第三个基因（PDK3）编码一种新的同工酶，它与PDK1和PDK2的氨基酸同一性分别为68%和67%。在大肠杆菌中表达的PDK3 cDNA指导合成一种分子量约为45,000 Da的多肽，该多肽对激酶缺陷型丙酮酸脱氢酶具有催化活性。作为重组蛋白，PDK3在测试的三种同工酶中似乎具有最高的比活性。通过Northern印迹分析对人PDK的所有三种同工酶的组织分布进行了表征。发现PDK2 mRNA在心脏和骨骼肌中含量最高，在胎盘和肺中含量最低。脑、肾、胰腺和肝脏表达中等量的PDK2（脑＞肾 = 胰腺＞肝脏）。PDK1 mRNA的组织分布与PDK2明显不同。PDK1的信息主要在心脏中表达，在其他组织中的表达水平适中（骨骼肌＞肝脏＞胰腺＞脑＞胎盘 = 肺＞肾）。与在所有测试组织中都有表达的PDK1和PDK2不同，PDK3的信息几乎只在心脏和骨骼肌中发现，这表明PDK3可能具有肌肉组织特有的特殊功能。在迄今为止测试的所有组织中，PDK2 mRNA的表达水平基本上高于PDK1和PDK3，这与PDK2是负责调节人类组织中丙酮酸脱氢酶的主要同工酶这一观点一致。
