Protein Phosphatase 1 Holoenzyme Formation and Subunit Exchange

蛋白磷酸酶 1 全酶形成和亚基交换

基本信息

  • 批准号:
    9985412
  • 负责人:
  • 金额:
    $ 4.39万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2019
  • 资助国家:
    美国
  • 起止时间:
    2019-07-10 至 2023-03-31
  • 项目状态:
    已结题

项目摘要

ABSTRACT Phosphorylation is one of the most ubiquitous, reversible posttranslational modifications in cells. The enzymes responsible for controlling the phosphorylation state of the cell are kinases, which catalyze the transfer of the γ-phosphate moiety of ATP to substrates, and phosphatases, which catalyze the reverse hydrolysis reaction, the removal of the phosphate moiety from phosphorylated substrates. Thus, phosphatases dynamically reverse the effects of kinases. Because phosphorylation is critical for all biological processes from cell growth to differentiation to development, the location and duration of the reciprocal actions of kinases and phosphatases must be exquisitely regulated both temporally and spatially within the cell. Consequently, when this tight regulation is disrupted, dysregulation of phosphorylation signaling ensues and the consequence is most often disease. Here we are investigating the cellular assembly of the serine/threonine protein phosphatase 1 (PP1). The regulatory protein SDS22 and the inhibitor-3 (I-3) have been proposed to be critical for this process, by functioning in a chaperone-like fashion. Using biochemistry and structural biology we show that this model is incorrect. Rather SDS22 and I-3 can individually function as a PP1 inhibitor. However, SDS22 inhibits PP1 via a completely novel mechanism. As we show, SDS22 unexpectedly distorts the PP1 active site so substrates can no longer bind. Thus as long as SDS22 is bound to PP1, PP1 is inactive. I-3 binds metals and thus can transport metals, which are necessary for PP1 activity, to the PP1 active site. Upon metal loading, I-3 is released in a p37-dependent manner from PP1. However, despite I-3 metal-loading, SDS22 continues to maintain PP1 in an inactive state. Thus, SDS22 is the key regulator of PP1 activity in cells. The presented research project uses a powerful integrated approach that combines X-ray crystallography and NMR spectroscopy with biochemical and cell biology experiments to obtain novel insights into the molecular mechanisms used by these regulators to control PP1 activity. We will: 1) determine the structures of the regulators in their free forms, 2) determine the structures of the PP1 dimeric and trimeric holoenzymes and 3) determine how these essential complexes direct and regulate PP1 signaling in cells. We will then leverage these structures to elucidate, at a molecular level, the biological functions and modes of action of these key PP1 holoenzymes. Because PP1 holoenzymes have critical roles in human diseases, the proposed work will provide novel strategies for selectively inhibiting PP1 activity by targeting the PP1 holoenzyme formation and subunit exchange, which is essential for understanding how distinct PPPs contribute to disease.
摘要 磷酸化是细胞中最普遍、最可逆的翻译后修饰之一。这些酶 负责控制细胞的磷酸化状态的是激酶,它催化细胞内 底物的γ-磷酸部分,以及催化逆水解反应的磷酸酶, 从磷酸化底物中去除磷酸盐部分。因此,磷酸酶动态逆转 激动酶的作用。因为磷酸化对所有生物过程都是至关重要的,从细胞生长到 分化到发育,激酶和磷酸酶相互作用的位置和持续时间 必须在细胞内的时间和空间上都进行精细的调节。因此,当这个紧身衣 调节被打乱,磷酸化信号的调节失调,其后果最常见的是 疾病。在这里,我们正在研究丝氨酸/苏氨酸蛋白磷酸酶1(PP1)的细胞组装。 调节蛋白SDS22和抑制物-3(I-3)被认为是这一过程的关键,通过 以一种类似监护人的方式运作。利用生物化学和结构生物学,我们证明了这个模型是 不正确。相反,SDS22和I-3可以单独作为PP1抑制剂发挥作用。然而,SDS22通过抑制PP1 一种全新的机制。如我们所示,SDS22出人意料地扭曲了PP1活性位点,因此底物 不能再捆绑了。因此,只要SDS22与PP1结合,PP1就处于非活性状态。I-3结合金属,因此可以 将PP1活性所必需的金属运输到PP1活性部位。在金属加载时,I-3是 以p37依赖的方式从PP1释放。然而,尽管I-3金属装载,SDS22继续 将PP1保持在非活动状态。因此,SDS22是细胞内PP1活性的关键调节因子。已提交的 研究项目使用了一种强大的集成方法,将X射线结晶学和核磁共振相结合 光谱学与生化和细胞生物学实验相结合,以获得对分子的新见解 这些调节器用来控制PP1活性的机制。我们将:1)确定 自由形式的调节剂,2)确定PP1二聚体和三聚体全酶的结构和3) 确定这些基本复合体如何引导和调节细胞中的PP1信号。然后我们将利用 这些结构从分子水平上阐明了这些键的生物学功能和作用方式 PP1全酶。由于PP1全酶在人类疾病中起着关键作用,拟议中的工作将 通过靶向PP1全酶的形成,提供选择性抑制PP1活性的新策略 亚基交换,这对于理解不同的PPP如何导致疾病至关重要。

项目成果

期刊论文数量(1)
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Wolfgang Peti其他文献

Wolfgang Peti的其他文献

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{{ truncateString('Wolfgang Peti', 18)}}的其他基金

Serine/Threonine Phosphatases in Neurological Diseases
神经系统疾病中的丝氨酸/苏氨酸磷酸酶
  • 批准号:
    10583671
  • 财政年份:
    2023
  • 资助金额:
    $ 4.39万
  • 项目类别:
Protein Phosphatase 1 Holoenzyme Formation
蛋白磷酸酶 1 全酶形成
  • 批准号:
    10441693
  • 财政年份:
    2022
  • 资助金额:
    $ 4.39万
  • 项目类别:
Protein Phosphatase 1 Holoenzyme Formation
蛋白磷酸酶 1 全酶形成
  • 批准号:
    10671729
  • 财政年份:
    2022
  • 资助金额:
    $ 4.39万
  • 项目类别:
Protein Phosphatase 1 Holoenzyme Formation
蛋白磷酸酶 1 全酶形成
  • 批准号:
    10793305
  • 财政年份:
    2022
  • 资助金额:
    $ 4.39万
  • 项目类别:
Shared Tundra screening cryo-EM for New England
新英格兰共享 Tundra 冷冻电镜筛查
  • 批准号:
    10413473
  • 财政年份:
    2022
  • 资助金额:
    $ 4.39万
  • 项目类别:
Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌中β-内酰胺抗性酶的机制和活性
  • 批准号:
    10624757
  • 财政年份:
    2019
  • 资助金额:
    $ 4.39万
  • 项目类别:
Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌中β-内酰胺抗性酶的机制和活性
  • 批准号:
    10391315
  • 财政年份:
    2019
  • 资助金额:
    $ 4.39万
  • 项目类别:
Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌β-内酰胺抗性酶的机制和活性
  • 批准号:
    9927573
  • 财政年份:
    2019
  • 资助金额:
    $ 4.39万
  • 项目类别:
Dynamics & energetics of p38a kinase regulation by ligands
动力学
  • 批准号:
    8608555
  • 财政年份:
    2013
  • 资助金额:
    $ 4.39万
  • 项目类别:
Dynamics & energetics of p38a kinase regulation by ligands
动力学
  • 批准号:
    8436569
  • 财政年份:
    2013
  • 资助金额:
    $ 4.39万
  • 项目类别:

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