Recognition of the inner lipoyl-bearing domain of dihydrolipoyl transacetylase and of the blood glucose-lowering compound AZD7545 by pyruvate dehydrogenase kinase 2.
Recognition of the inner lipoyl-bearing domain of dihydrolipoyl transacetylase and of the blood glucose-lowering compound AZD7545 by pyruvate dehydrogenase kinase 2.
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丙酮酸脱氢酶激酶 2 识别二氢硫辛酰转乙酰酶和降血糖化合物 AZD7545 的内部硫辛酰基结构域。
DOI：
10.1021/bi700650k
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发表时间：
2007
期刊：
影响因子：
2.9
通讯作者：
Popov,KirillM
中科院分区：
文献类型：
--
作者：
Tuganova,Alina;Klyuyeva,Alla;Popov,KirillM
丙酮酸脱氢酶激酶2（PDHK2）是一种独特的线粒体蛋白激酶，调节丙酮酸脱氢酶多酶复合物（PDC）的活性。PDHK2是PDC的组成部分，与PDC的二氢硫辛酰基转乙酰酶组分（E2）的内部硫辛酰基承载结构域（L2）紧密结合。据报道，这种结合使激酶活性增加高达10倍。尽管E2在维持PDHK2在PDC结合状态下的功能性方面发挥了重要作用，但负责PDHK2识别L2和E2依赖性PDHK2活化的分子机制在很大程度上是未知的。在这项研究中，我们使用了分子建模和定点诱变的组合，以确定PDHK2和L2之间的相互作用和E2激活PDHK2所必需的氨基酸残基。基于定点诱变的结果，似乎位于其R结构域（P22、L23、F28、F31、F44、L45和L160）和所谓的“横臂”结构（K368、R372和K391）中的许多PDHK 2残基在确定PDHK 2和L2之间的相互作用的强度中是关键的。对于被PDHK 2识别所必需的L2残基包括L140、K173、I176、E179，以及较小程度的D164、D172和A174。重要的是，某些PDHK 2残基与L2形成界面，即，K17、P22、F31、F44、R372和K391对于维持E2结合状态下增强的PDHK2活性也是关键的。最后，有证据表明，降糖化合物AZD 7545破坏了PDHK 2和L2之间的相互作用，从而抑制了PDHK 2的活性。
Pyruvate dehydrogenase kinase 2 (PDHK2) is a unique mitochondrial protein kinase that regulates the activity of the pyruvate dehydrogenase multienzyme complex (PDC). PDHK2 is an integral component of PDC tightly bound to the inner lipoyl-bearing domains (L2) of the dihydrolipoyl transacetylase component (E2) of PDC. This association has been reported to bring about an up to 10-fold increase in kinase activity. Despite the central role played by E2 in the maintenance of PDHK2 functionality in the PDC-bound state, the molecular mechanisms responsible for the recognition of L2 by PDHK2 and for the E2-dependent PDHK2 activation are largely unknown. In this study, we used a combination of molecular modeling and site-directed mutagenesis to identify the amino acid residues essential for the interaction between PDHK2 and L2 and for the activation of PDHK2 by E2. On the basis of the results of site-directed mutagenesis, it appears that a number of PDHK2 residues located in its R domain (P22, L23, F28, F31, F44, L45, and L160) and in the so-called “cross arm” structure (K368, R372, and K391) are critical in determining the strength of the interaction between PDHK2 and L2. The residues of L2 essential for recognition by PDHK2 include L140, K173, I176, E179, and to a lesser extent D164, D172, and A174. Importantly, certain PDHK2 residues forming interfaces with L2, i.e., K17, P22, F31, F44, R372, and K391, are also critical for the maintenance of enhanced PDHK2 activity in the E2-bound state. Finally, evidence that the blood glucose-lowering compound AZD7545 disrupts the interactions between PDHK2 and L2 and thereby inhibits PDHK2 activity is presented.