MRC Clinical Sciences Centre, Cellular Stress Group, Hammersmith Hospital Campus, Imperial College, DuCane Road, London, UK.
Cell Metab. 2011 Nov 2;14(5):707-14. doi: 10.1016/j.cmet.2011.09.009. Epub 2011 Oct 20.
The SNF1 protein kinase complex plays an essential role in regulating gene expression in response to the level of extracellular glucose in budding yeast. SNF1 shares structural and functional similarities with mammalian AMP-activated protein kinase. Both kinases are activated by phosphorylation on a threonine residue within the activation loop segment of the catalytic subunit. Here we show that ADP is the long-sought metabolite that activates SNF1 in response to glucose limitation by protecting the enzyme against dephosphorylation by Glc7, its physiologically relevant protein phosphatase. We also show that the regulatory subunit of SNF1 has two ADP binding sites. The tighter site binds AMP, ADP, and ATP competitively with NADH, whereas the weaker site does not bind NADH, but is responsible for mediating the protective effect of ADP on dephosphorylation. Mutagenesis experiments suggest that the general mechanism by which ADP protects against dephosphorylation is strongly conserved between SNF1 and AMPK.
SNF1 蛋白激酶复合物在调节细胞对细胞外葡萄糖水平的反应中的基因表达中起着至关重要的作用。SNF1 在结构和功能上与哺乳动物 AMP 激活的蛋白激酶（AMPK）相似。这两种激酶都通过在催化亚基的激活环片段中的一个苏氨酸残基的磷酸化而被激活。在这里，我们发现 ADP 是通过保护酶免受其生理相关的蛋白磷酸酶 Glc7 的去磷酸化来激活 SNF1 的长期寻找的代谢物，从而响应葡萄糖限制。我们还发现 SNF1 的调节亚基有两个 ADP 结合位点。 tighter 位点与 NADH 竞争性地结合 AMP、ADP 和 ATP，而 weaker 位点不结合 NADH，但负责介导 ADP 对去磷酸化的保护作用。突变实验表明，ADP 防止去磷酸化的一般机制在 SNF1 和 AMPK 之间是高度保守的。
