Molecular determinants of KA1 domain-mediated autoinhibition and phospholipid activation of MARK1 kinase.
Molecular determinants of KA1 domain-mediated autoinhibition and phospholipid activation of MARK1 kinase.
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DOI：
10.1042/bcj20160792
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发表时间：
2017-02-01
期刊：
影响因子：
--
通讯作者：
Ferguson KM
中科院分区：
文献类型：
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作者：
Emptage RP;Lemmon MA;Ferguson KM
蛋白激酶通常由蛋白质模块之间的分子内自抑制相互作用调节，当这些模块与其他“激活”的蛋白质或膜结合的靶标结合时，这种相互作用被逆转。一组激酶，MAP/微管亲和力调节激酶(MARKs)在其C端含有一个鲜为人知的调节模块，KA1(激酶相关-1)结构域。Mark1的KA1结构域和从酵母到人类的几种相关激酶已被证明与含有阴离子磷脂的膜结合，也有多肽配体的报道。据报道，删除或突变C端的KA1结构域可以激活发现该结构域的激酶，这也表明了分子内的自我抑制作用。在这里，我们证明了人类Mark1的KA1结构域与Mark1激活域相互作用并抑制。利用定点突变，我们确定了这种自抑制活性所需的KA1结构域中的残基，并发现参与自抑制和阴离子磷脂结合的残基是相同的。我们还证明了，当与含有阴离子磷脂的小泡结合时，“迷你”Mark1就会被激活，但前提是蛋白质通过第二个信号靶向这些小泡。这些研究为理解Mark1及其近亲如何在膜表面需要一个以上的信号来控制其在正确的位置和时间的激活提供了机制基础。MARK家族激酶与阿尔茨海默氏症、癌症和自闭症等多种疾病状态有关，因此推进我们对其调节机制的理解可能最终具有治疗价值。
Protein kinases are frequently regulated by intramolecular autoinhibitory interactions between protein modules that are reversed when these modules bind other ‘activating’ protein or membrane-bound targets. One group of kinases, the MAP/microtubule affinity-regulating kinases (MARKs) contain a poorly understood regulatory module, the KA1 (kinase associated-1) domain, at their C-terminus. KA1 domains from MARK1 and several related kinases from yeast to humans have been shown to bind membranes containing anionic phospholipids, and peptide ligands have also been reported. Deleting or mutating the C-terminal KA1 domain has been reported to activate the kinase in which it is found — also suggesting an intramolecular autoinhibitory role. Here, we show that the KA1 domain of human MARK1 interacts with, and inhibits, the MARK1 kinase domain. Using site-directed mutagenesis, we identify residues in the KA1 domain required for this auto-inhibitory activity, and find that residues involved in autoinhibition and in anionic phospholipid binding are the same. We also demonstrate that a ‘mini’ MARK1 becomes activated upon association with vesicles containing anionic phospholipids, but only if the protein is targeted to these vesicles by a second signal. These studies provide a mechanistic basis for understanding how MARK1 and its relatives may require more than one signal at the membrane surface to control their activation at the correct location and time. MARK family kinases have been implicated in a plethora of disease states including Alzheimer’s, cancer, and autism, so advancing our understanding of their regulatory mechanisms may ultimately have therapeutic value.