PP1:NIPP1 HOLOENZYME

PP1:NIPP1全酶

• 批准号: 8363375
• 负责人: Rebecca Page
• 金额: $ 0.4万
• 依托单位: BROOKHAVEN SCIENCE ASSOC-BROOKHAVEN LAB
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363375
• 关键词: Active Sites; Binding; Binding Sites; Catalytic Domain; Cell Cycle Regulation; Cell division; Cell physiology; Complex; Crystallization; Crystallography; DNA Repair; Data; Enzymes; Eukaryota; Event; Funding; Gene Silencing; Grant; Holoenzymes; Light; Molecular Conformation; National Center for Research Resources; Needles; Nuclear; Nuclear Protein; Principal Investigator; Protein Serine/Threonine Phosphatase; Protein phosphatase; Proteins; RNA Splicing; Regulation; Research; Research Infrastructure; Resources; Sampling; Site; Source; Surface; Synchrotrons; Toxin; United States National Institutes of Health; Work; cost; inhibitor/antagonist; insight; interest; prevent; research study; small molecule; two-dimensional

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Phosphoprotein Phosphatase 1, PP1, is a ubiquitous serine/threonine phosphatase (330 residues, ~ 38 kDa) expressed in all eukaryotes that regulates a wide array of cellular processes. The promiscuity of PP1 is greatly reduced by interaction with several inhibitory and targeting proteins. By interacting with residues of the active site of PP1, inhibitory proteins prevent to access catalytic residues of the enzyme. The targeting proteins relocate PP1 to a particular cellular milieu and interact with the surface of the catalytic core in such a way that only certain binding sites are exposed, whereas, others are sites are sterically occluded. Current crystallographic studies of PP1 in complex with small molecule toxins, inhibitory proteins and targeting proteins have revealed that the surface and conformation of the catalytic core remains invariant; however, all of the proteins studied to date have shown markeedly different themes in their interactions with the PP1. To shed new insights on PP1 regulation by targeting proteins, we have initiated work with one such protein, NIPP (Nuclear Inhibitor of PP1). NIPP targets PP1 after interaction with various proteins implicated in four different cellular processes (cell division, gene silencing, DNA repair and RNA splicing). Since targeting of NIPP1 to PP1 is a major event in cell cycle regulation; thus, we are interested in understanding this relationship at the atomic level. After exhaustive biophysical characterization of the monomeric form of the PP1-binding domain of NIPP, we have moved to studying the holoenzyme complex; to this end, we have determined the Kd with ITC (low nM), used data from complex NMR experiments to optimize samples for crystallization trails and attained two dimensional needle-like crystals.

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其他NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 磷蛋白磷酸酶1（PP 1）是一种普遍存在的丝氨酸/苏氨酸磷酸酶（330个残基，~ 38 kDa），在所有真核生物中表达，调节广泛的细胞过程。 PP 1的混杂性通过与几种抑制性和靶向蛋白的相互作用而大大降低。 通过与PP 1活性位点的残基相互作用，抑制性蛋白阻止接近酶的催化残基。 靶向蛋白将PP 1重新定位到特定的细胞环境中，并以仅暴露某些结合位点的方式与催化核心的表面相互作用，而其他位点则在空间上被封闭。 PP 1与小分子毒素、抑制蛋白和靶向蛋白复合物的当前晶体学研究已经揭示了催化核心的表面和构象保持不变;然而，迄今为止研究的所有蛋白质在与PP 1的相互作用中显示出显著不同的主题。 为了通过靶向蛋白质对PP 1调节提供新的见解，我们已经开始了对一种这样的蛋白质NIPP（PP 1核抑制剂）的研究。 NIPP在与涉及四种不同细胞过程（细胞分裂、基因沉默、DNA修复和RNA剪接）的各种蛋白质相互作用后靶向PP 1。 由于NIPP 1靶向PP 1是细胞周期调控中的一个重要事件，因此，我们有兴趣在原子水平上了解这种关系。 经过详尽的生物物理特性的单体形式的PP 1结合结构域的NIPP，我们已经转移到研究全酶复合物;为此，我们已经确定了Kd与ITC（低nM），使用复杂的NMR实验的数据，以优化样品的结晶试验，并获得二维针状晶体。