Dynein light chain as a dimerization hub for natively disordered proteins

动力蛋白轻链作为天然无序蛋白质的二聚化中心

• 批准号: 8137075
• 负责人: ELISAR J BARBAR
• 金额: $ 34.23万
• 依托单位: OREGON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8137075
• 关键词: Affect; Apoptosis; Binding; Binding Sites; Biochemical; Biological; C-terminal; Calorimetry; Cell physiology; Cells; Circular Dichroism; Complement; Complex; Cytoskeleton; Deglutition; Development; Dimerization; Disease; Drosophila genus; Dynein ATPase; Enzymes; Fluorescence Resonance Energy Transfer; Fluorescence Spectroscopy; Foundations; Health; Homeostasis; In Vitro; Intracellular Transport; Lead; Length; Light; Link; Masks; Messenger RNA; Microtubules; Modeling; Molecular; Molecular Motors; Motor; Multiple Partners; Mutation; N-terminal; Nurses; Organism; Phenotype; Physiological Processes; Play; Property; Proteins; Proteolysis; Publishing; Regulation; Role; Solid; Solutions; Stretching; Structure; System; Techniques; Testing; Thermodynamics; Titrations; Variant; Viral Pathogenesis; WD Repeat; Work; X-Ray Crystallography; base; design; dimer; dynein light chain; in vivo; insight; interest; mutant; nephrogenesis; novel; protein complex; protein function; protein protein interaction; research study; structural biology; success

DESCRIPTION (provided by applicant): Dynein light chain LC8 is a highly conserved, essential component of the microtubule-based molecular motor dynein. Its interactions with a number of non-dynein proteins led to the widely held view that it functions as a cargo adaptor. However, structural and thermodynamics work has recently shown that LC8 cannot serve as a cargo adaptor and has led to the paradigm-shifting proposition that LC8 is not primarily a dynein subunit, but is an essential component of diverse protein complexes that play roles in a variety of cellular systems. The far-reaching and unifying hypothesis of this proposal is that LC8 is a hub protein and functions as a dimerization facilitator in dynein and in all other complexes in which it participates, promoting the dimerization and ordering of the proteins with which it interacts. Work in this proposal will elucidate binding thermodynamics and structural characterization of the two best-known proteins that interact with LC8. Aim 1 focuses on the interactions of LC8 with the intermediate chain (IC) in dynein assembly and aim 2 focuses on the interactions of LC8 with Swallow, a protein involved in localization of bicoid mRNA during Drosophila development, and the best-studied example of an LC8-interacting protein not associated with dynein. For both aims the mechanism and biological significance of LC8-promoted partner dimerization will be examined by both in vitro biophysical studies of representative protein segments and, when possible, full-length constructs, and in vivo assessment of phenotypes of cells and organisms expressing designed variants of IC and Swallow. These studies will provide a solid foundation for understanding the roles that LC8 plays in the variety of cellular systems in which it participates, and will also promote progress in the LC8 field in general as other LC8 binding partners are physiologically important. These studies also have broader applicability, because LC8 can serve as a model for other hub proteins that interact with a large number of disordered partners. In addition these studies will provide novel insights into the structural biology of dynein motor function, complementing the cell biological approaches that predominate the field of intracellular transport. Dr. Barbar's track record of pioneering work on structure-function relations of LC8, her role as developer of the novel hypothesis that LC8 is a dimerization hub for natively disordered proteins, her success in both producing useful protein constructs and handling these complex and partially disordered proteins, and her expertise in the battery of biophysical and biochemical solution techniques required to probe these questions, make her uniquely suited to lead this effort. PUBLIC HEALTH RELEVANCE: In protein-protein interaction networks, a small proportion of proteins termed "hubs" interact with many diverse partners. These studies will elucidate the mechanism of interaction of dynein light chain LC8, which we propose is a hub protein, and its role as a dimerization engine in the assembly and regulation of the microtubule-based motor complex dynein, and in assembly of non-dynein complexes that are involved in various physiological processes.

描述（由申请人提供）：Dynein轻链LC8是基于微管的分子运动动力蛋白的高度保守，必不可少的组成部分。它与许多非二氧蛋白蛋白的相互作用导致人们广泛认为它是货物适配器。然而，结构和热力学工作最近表明，LC8不能用作货物适配器，并导致范式转移的主张，即LC8主要不是Dynein亚基，而是多种蛋白质复合物的重要组成部分，在多种细胞系统中起着作用。该提议的深远和统一的假设是LC8是一个集线器蛋白，在动力蛋白以及其参与的所有其他复合物中起着二聚化的促进剂的作用，促进了与之相互作用的蛋白质的二聚化和顺序。该提案中的工作将阐明与LC8相互作用的两种最著名蛋白质的结合热力学和结构表征。 AIM 1专注于Dynein组装中LC8与中间链（IC）的相互作用，AIM 2的重点是LC8与燕子的相互作用，燕子发育过程中涉及双聚体mRNA的蛋白质，以及在LC8相互作用的蛋白质中最有研究的示例。对于目标，LC8促进伴侣二聚化的机制和生物学意义都将通过代表性蛋白质段的体外生物物理研究以及在可能的情况下进行全长构建体以及表达IC和Swallow的表达细胞表表表征的体内评估的体内评估。这些研究将为理解LC8在参与的各种细胞系统中所起的作用提供稳固的基础，并将一般来说，因为其他LC8结合伙伴在生理上很重要，也将促进LC8领域的进步。这些研究也具有更广泛的适用性，因为LC8可以作为与大量无序伴侣相互作用的其他枢纽蛋白的模型。此外，这些研究将提供对动力蛋白运动功能的结构生物学的新见解，并补充占细胞内转运领域的细胞生物学方法。 Dr. Barbar's track record of pioneering work on structure-function relations of LC8, her role as developer of the novel hypothesis that LC8 is a dimerization hub for natively disordered proteins, her success in both producing useful protein constructs and handling these complex and partially disordered proteins, and her expertise in the battery of biophysical and biochemical solution techniques required to probe these questions, make her uniquely suited to lead this 努力。公共卫生相关性：在蛋白质 - 蛋白质相互作用网络中，一小部分称为“枢纽”的蛋白质与许多不同的伴侣相互作用。这些研究将阐明动力蛋白轻链LC8相互作用的机理，我们提出的是枢纽蛋白，并且它在基于微管的运动型综合动力蛋白的组装和调节中作为二聚发动机的作用，以及与非二氧蛋白复合物组装中有关各种物理学过程的作用。