In vitro reconstitution of Sgk3 activation by phosphatidylinositol 3-phosphate.
In vitro reconstitution of Sgk3 activation by phosphatidylinositol 3-phosphate.
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DOI：
10.1016/j.jbc.2021.100919
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发表时间：
2021-08
期刊：
影响因子：
--
通讯作者：
Leonard TA
中科院分区：
文献类型：
--
作者：
Pokorny D;Truebestein L;Fleming KD;Burke JE;Leonard TA
血清和糖皮质激素调节的激酶3（Sgk3）是一种丝氨酸/苏氨酸蛋白激酶，其通过I类磷脂酰肌醇3-激酶（PI3K）信号传导在生长因子信号传导的下游由磷脂酰肌醇3-磷酸（PI3P）活化，并通过III类PI3K/Vps34介导的内体上的PI3P产生活化。最近，Sgk 3活性上调与许多人类癌症有关;然而，Sgk 3激活的确切机制尚不清楚。在这里，我们使用了广泛的细胞生物学，生物化学和生物物理技术，包括氢氘交换质谱，研究PI3P激活Sgk3的机制。我们发现，Sgk3是由磷酸化和变构激活的组合。我们表明，结合的Sgk3 PI3P通过其调节phox同源性（PX）域诱导大的构象变化，与其激活Sgk3和PI3P结合口袋的PX域的Sgk3被隔离在其非活性构象。最后，我们在体外通过Vps34介导的磷脂酰肌醇脂质体上的PI3P合成重建Sgk3激活。除了确定PI3P激活Sgk3的机制外，我们的发现还开辟了变构抑制剂开发的潜在治疗途径，以靶向癌症中的Sgk3。
Serum- and glucocorticoid-regulated kinase 3 (Sgk3) is a serine/threonine protein kinase activated by the phospholipid phosphatidylinositol 3-phosphate (PI3P) downstream of growth factor signaling via class I phosphatidylinositol 3-kinase (PI3K) signaling and by class III PI3K/Vps34-mediated PI3P production on endosomes. Upregulation of Sgk3 activity has recently been linked to a number of human cancers; however, the precise mechanism of activation of Sgk3 is unknown. Here, we use a wide range of cell biological, biochemical, and biophysical techniques, including hydrogen–deuterium exchange mass spectrometry, to investigate the mechanism of activation of Sgk3 by PI3P. We show that Sgk3 is regulated by a combination of phosphorylation and allosteric activation. We demonstrate that binding of Sgk3 to PI3P via its regulatory phox homology (PX) domain induces large conformational changes in Sgk3 associated with its activation and that the PI3P-binding pocket of the PX domain of Sgk3 is sequestered in its inactive conformation. Finally, we reconstitute Sgk3 activation via Vps34-mediated PI3P synthesis on phosphatidylinositol liposomes in vitro. In addition to identifying the mechanism of Sgk3 activation by PI3P, our findings open up potential therapeutic avenues in allosteric inhibitor development to target Sgk3 in cancer.
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影响因子：
16
作者：
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通讯作者：
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影响因子：
50.3
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DOI：
10.1107/s0907444906029799
发表时间：
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影响因子：
14.8
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DOI：
10.1073/pnas.1109879108
发表时间：
2011-11-15
影响因子：
11.1
作者：
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通讯作者：
Feldman, Arthur M.