Mechanism of action of A-769662, a valuable tool for activation of AMP-activated protein kinase.
Mechanism of action of A-769662, a valuable tool for activation of AMP-activated protein kinase.
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DOI：
10.1074/jbc.m706536200
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发表时间：
2007-11-09
期刊：
影响因子：
--
通讯作者：
Sakamoto K
中科院分区：
文献类型：
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作者：
Göransson O;McBride A;Hawley SA;Ross FA;Shpiro N;Foretz M;Viollet B;Hardie DG;Sakamoto K
我们研究了 A-769662（一种新的 AMP 激活蛋白激酶 (AMPK) 激活剂）的机制。与其他药理学激活剂不同，它通过模仿 AMP 的两种效应，即变构激活和去磷酸化抑制，直接激活天然大鼠 AMPK。它对分离的 α 亚基激酶结构域（有或没有相关的自抑制结构域）、糖原与 β 亚基糖原结合结构域的相互作用或 AMP 与 γ 亚基的分离的 Bateman 结构域的结合没有影响。将 A-769662 添加到小鼠胚胎成纤维细胞 (MEF) 或原代小鼠肝细胞中会刺激乙酰辅酶 A 羧化酶 (ACC) 的磷酸化，这种作用在 AMPK-α1−/−α2−/− 细胞中完全消除，但在 TAK1−/− MEF 中不会。 LKB1−/− 小鼠肌肉中响应 A-769662 的 AMPK 和 ACC 磷酸化也被消除。然而，在缺乏 LKB1 但表达替代上游激酶 CaMKKβ 的 HeLa 细胞中，响应 A-769662 的 AMPK 和 ACC 磷酸化仍然发生。这些结果表明，在完整细胞中，A-769662 的作用独立于所使用的上游激酶。
We have studied the mechanism of A-769662, a new activator of AMP-activated protein kinase (AMPK). Unlike other pharmacological activators it directly activates native rat AMPK by mimicking both effects of AMP, i.e. allosteric activation and inhibition of dephosphorylation. It has no effect on the isolated α subunit kinase domain with or without the associated auto-inhibitory domain, on interaction of glycogen with the β subunit glycogen-binding domain, or on binding of AMP to the isolated Bateman domains of the γ subunit. Addition of A-769662 to mouse embryo fibroblasts (MEFs) or primary mouse hepatocytes stimulates phosphorylation of acetyl-CoA carboxylase (ACC), effects that are completely abolished in AMPK-α1−/−α2−/− cells, but not in TAK1−/− MEFs. Phosphorylation of AMPK and ACC in response to A-769662 is also abolished in LKB1−/− mouse muscle. However in HeLa cells, which lack LKB1 but express the alternate upstream kinase CaMKKβ, phosphorylation of AMPK and ACC in response to A-769662 still occurs. These results show that in intact cells the effects of A-769662 are independent of the upstream kinase utilized.