Structural studies of PARK14

PARK14的结构研究

• 批准号: 9180460
• 负责人: SERGEY KOROLEV
• 金额: $ 22.73万
• 依托单位: SAINT LOUIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9180460
• 关键词: Active Sites; Affect; Animal Model; Animals; Ankyrin Repeat; Arachidonic Acids; Binding; Binding Sites; Biochemical; Brain; Calcium; Calmodulin; Cardiovascular Diseases; Catalytic Domain; Cellular Assay; Complex; Crystallization; Data; Development; Diabetes Mellitus; Disease; Epitopes; Event; Foundations; Future; Genes; Goals; Grant; Heart; Heavy Metals; Homologous Gene; In Vitro; Individual; Inherited; Knowledge; Laboratories; Length; Link; Lipids; Lysophospholipids; Malignant Neoplasms; Maps; Membrane; Membrane Proteins; Methods; Modeling; Molecular; Molecular Conformation; Muscular Dystrophies; Mutation; Nerve Degeneration; Organ; PLA2G6 gene; Pancreas; Parkinson Disease; Pathway interactions; Phase; Phospholipase; Physiological; Play; Probability; Property; Proteins; Regulation; Regulatory Element; Resolution; Role; Seitelberger' s Disease; Selenium; Selenomethionine; Signal Pathway; Signal Transduction; Structure; Surface; System; Techniques; Tertiary Protein Structure; Testing; Tissues; base; cell type; cofactor; design; electron density; enzyme activity; enzyme mechanism; enzyme structure; improved; in vivo; insight; mutant; nervous system disorder; novel; novel therapeutic intervention; protein function; protein structure

Mutations in the PARK14 gene are strongly associated with a spectrum of neurological disorders, including Parkinson's disease (PD) and infantile neuroaxonal dystrophy (INAD) through currently unknown mechanisms. The product of the PARK14 gene is an intracellular calcium-independent phospholipase (PLA2G6 or iPLA2β), which has been implicated in numerous cellular pathways. The protein has a unique multi-domain structure and its activity is regulated at several levels. We propose to solve the crystal structure of the protein in order to advance the mechanistic understanding of iPLA2β function in the brain and other organs. An atomic resolution structure of the enzyme is critical for understanding the mechanism of its activity, its regulation and its function in physiological and pathological states. It will provide the foundation for future development of novel therapeutic approaches to treating neurological and cardiovascular diseases as well as diabetes, cancer and muscular dystrophy. iPLA2β modulates membrane properties, produces bioactive lipid messengers such as arachidonic acid and lysophospholipids, and regulates store-operated calcium entry in multiple cell types. Calmodulin (CaM) inhibits iPLA2β enzyme activity in the presence of calcium. Several cofactors reverse the inhibition. Numerous additional regulatory mechanisms have been suggested including oligomerization, ATP binding and interaction with other cofactors and proteins. The large number and variety of signaling pathways affected by iPLA2β and the complexity of its macromolecular interactions complicate defining its function in cellular events and the role it plays in neurological disorders. PARK14 mutations are found in all structural domains of the protein. Studies of these mutations provide a unique opportunity to link regulatory elements and the catalytic activity of iPLA2β to specific signaling mechanisms and functions. Structural information about the conformation of surface epitopes, of the active site and membrane and protein recognition interfaces will be indispensible for these studies. The goal of this proposal is to obtain this structural knowledge to significantly advance the understanding of iPLA2β function. We have crystallized the full-length iPLA2β and now propose to improve the diffraction quality of the crystals and to obtain phasing information to solve the crystal structure of the enzyme. Results from the proposed studies, together with data obtained from biochemical assays, cellular systems and animal models from our group and others will move the entire field forward and will be critical for understanding the functional role of iPLA2β in the brain and other organs including heart and pancreas.

PARK14基因中的突变与一系列神经系统疾病密切相关， 包括当前未知 机制。 PARK14基因的产物是细胞内钙独立的磷脂酶 （PLA2G6或IPLA2β），在许多细胞途径中隐含。该蛋白具有独特的 多域结构及其活性在多个级别进行调节。我们建议解决晶体结构 蛋白质的蛋白质，以提高对大脑和其他IPLA2β功能的机械理解 器官。酶的原子分辨率结构对于理解其活性机制至关重要， 它的调节及其在物理和病理状态中的功能。它将为未来提供基础 开发新的治疗方法来治疗神经系统和心血管疾病以及 糖尿病，癌症和肌肉营养不良。 IPLA2β调节膜特性，产生生物活性脂质使者，例如花生四烯酸 和溶物磷脂，并调节多种细胞类型的商店经营的钙进入。钙调蛋白（CAM） 在存在钙的情况下抑制IPLA2β酶活性。几个辅助因子反转了抑制作用。很多的 已经提出了其他调节机制，包括寡聚，ATP结合和相互作用 与其他辅助因子和蛋白质。受IPLA2β和 其大分子相互作用的复杂性使定义其在细胞事件中的功能和作用复杂 它在神经系统疾病中发挥作用。在蛋白质的所有结构结构域中发现PARK14突变。研究 这些突变为连接调节元件和IPLA2β的催化活性提供了独特的机会 特定的信号机制和功能。有关表面构象的结构信息 活性位点以及膜和蛋白质识别界面的表位是必不可少的 研究。该提议的目的是获得这种结构知识，以显着促进 了解IPLA2β功能。 我们已经结晶了全长IPLA2β，现在提出了提高提高衍射质量的建议 晶体并获得分阶段信息以求解酶的晶体结构。结果 拟议的研究，以及从生化测定，细胞系统和动物模型获得的数据 来自我们的小组和其他人将向前推进整个领域，对于理解功能至关重要 IPLA2β在大脑和其他器官中的作用，包括心脏和胰腺。