Structural studies of PARK14

PARK14基因的突变与一系列神经疾病密切相关， 包括帕金森氏病(PD)和婴儿神经轴索营养不良症(INAD) 机械装置。PARK14基因的产物是一种细胞内不依赖钙的磷脂酶 (PLA2G6或iPLA2β)，它与许多细胞通路有关。这种蛋白质有一种独特的 多结构域的结构和活性在多个水平上受到调节。我们提出要解决晶体结构的问题 以促进对iPLA2β在大脑和其他组织中功能的机制理解 器官。酶的原子分辨结构对于理解其活性机制是至关重要的， 它的调节及其在生理和病理状态下的作用。它将为未来的发展奠定基础 开发治疗神经和心血管疾病的新治疗方法以及 糖尿病、癌症和肌肉营养不良。 IPLA2β调节膜性质，产生生物活性脂质信使，如花生四烯酸 和溶血磷脂，并调节多种细胞类型的储存操作的钙内流。钙调素(CaM) 在钙存在的情况下抑制iPLA2β酶的活性。有几个辅助因素逆转了这种抑制作用。数不胜数 还提出了其他调节机制，包括寡聚、ATP结合和相互作用 与其他辅因子和蛋白质结合。IPLA2β和iPLA2影响的大量和多样化的信号通路 其大分子相互作用的复杂性使其在细胞事件中的功能和作用的定义变得复杂 它在神经系统疾病中发挥作用。PARK14突变存在于蛋白质的所有结构域中。研究 这些突变提供了一个独特的机会将调节元件与iPLA2β的催化活性联系起来 特定的信号机制和功能。关于表面构象的结构信息 活性部位、膜和蛋白质识别界面的表位对于这些是必不可少的 学习。这项提议的目标是获得这种结构性知识，以显著推进 了解iPLA2的β功能。 我们已经结晶了全长的iPLA2β，现在建议提高iPLA2的衍射质量 并获得晶相信息来解算该酶的晶体结构。调查结果： 拟议的研究，以及从生化分析、细胞系统和动物模型获得的数据 将推动整个领域向前发展，并将对理解功能 IPLA2β在脑和其他器官中的作用，包括心脏和胰腺。

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.