The transfer of a phosphate from ATP to a protein substrate, a modification known as protein phosphorylation, is catalyzed by protein kinases. Protein kinases play a crucial role in virtually every cellular activity. Recent studies of atypical protein kinases have highlighted the structural similarity of the kinase superfamily despite notable differences in primary amino acid sequence. Here, using a bioinformatics screen, we searched for putative protein kinases in the intracellular bacterial pathogen Legionella pneumophila and identified the type 4 secretion system effector Lpg2603 as a remote member of the protein kinase superfamily. Employing an array of biochemical and structural biology approaches, including in vitro kinase assays and isothermal titration calorimetry, we show that Lpg2603 is an active protein kinase with several atypical structural features. Importantly, we found that the eukaryote-specific host signaling molecule inositol hexakisphosphate (IP6) is required for Lpg2603 kinase activity. Crystal structures of Lpg2603 in the apo-form and when bound to IP6 revealed an active-site rearrangement that allows for ATP binding and catalysis. Our results on the structure and activity of Lpg2603 reveal a unique mode of regulation of a protein kinase, provide the first example of a bacterial kinase that requires IP6 for its activation, and may aid future work on the function of this effector during Legionella pathogenesis.

中文翻译：

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军团菌效应激酶被宿主肌醇六磷酸酯激活。

磷酸激酶将磷酸从ATP转移至蛋白质底物（一种称为蛋白质磷酸化的修饰）。蛋白激酶在几乎所有细胞活动中都起着至关重要的作用。尽管在一级氨基酸序列上存在显着差异，但对非典型蛋白激酶的最新研究强调了激酶超家族的结构相似性。在这里，我们使用生物信息学的筛选方法，在细胞内细菌病原体肺炎军团菌中寻找推定的蛋白激酶，并将4型分泌系统效应物Lpg2603鉴定为蛋白激酶超家族的一个远程成员。采用了一系列生物化学和结构生物学方法，包括体外激酶测定和等温滴定量热法，我们显示Lpg2603是具有几种非典型结构特征的活性蛋白激酶。重要的是，我们发现Lpg2603激酶活性需要真核生物特异性宿主信号分子肌醇六磷酸（IP6）。Lpg2603的apo型和绑定到IP6的晶体结构揭示了一个活性位点重排，允许ATP结合和催化。我们对Lpg2603的结构和活性的研究结果揭示了蛋白激酶的独特调节模式，提供了需要IP6激活其细菌激酶的第一个实例，并且可能有助于在军团菌发病过程中对该效应子功能的进一步研究。Lpg2603的apo型和绑定到IP6的晶体结构揭示了一个活性位点重排，允许ATP结合和催化。我们对Lpg2603的结构和活性的研究结果揭示了蛋白激酶的独特调节模式，提供了需要IP6激活其细菌激酶的第一个实例，并且可能有助于将来在军团菌发病过程中对该效应子的功能进行研究。Lpg2603的apo型和与IP6结合时的晶体结构揭示了一个活性位点重排，允许ATP结合和催化。我们对Lpg2603的结构和活性的研究结果揭示了蛋白激酶的独特调节模式，提供了需要IP6激活其细菌激酶的第一个实例，并且可能有助于将来在军团菌发病过程中对该效应子的功能进行研究。