Struct. Invest. of Hsp70 and TGFb by Solution Studies and High-Perform. Comput.

结构。

• 批准号: 8529559
• 负责人: Emre H. Brookes
• 金额: $ 12.39万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8529559
• 关键词: Affect; Binding; Biological; Biological Assay; Cell Differentiation process; Cell Proliferation; Cell membrane; Complement; Complex; Computer Analysis; Computer software; Computing Methodologies; Data; Data Analyses; Development; Diffusion; Extracellular Matrix Proteins; Grant; Investigation; Knockout Mice; Knowledge; Malignant Neoplasms; Mammals; Mediating; Methodology; Methods; Modeling; Molecular; Molecular Chaperones; Molecular Conformation; Neoplasm Metastasis; Neurodegenerative Disorders; Neutrons; Nucleotides; Peptides; Performance; Phenotype; Physiological; Play; Process; Protein Isoforms; Proteins; Radial; Reaction; Relative (related person); Resolution; Roentgen Rays; Role; Signal Transduction; Simulate; Solutions; Solvents; Structure; System; Techniques; Tertiary Protein Structure; Testing; Tissues; Validation; Variant; Viscosity; analytical ultracentrifugation; base; biological systems; heat-shock proteins 110; human disease; in vivo; macromolecular assembly; macromolecule; molecular dynamics; novel; public health relevance; receptor; research study; response; sedimentation coefficient; software development; therapeutic development; three dimensional structure; tumor growth

DESCRIPTION (provided by applicant): This grant proposes the development of a novel methodology, based on computational approaches and experiments performed under physiological solution conditions, to complement high-resolution methods for the structure determination of biological macromolecules and their assemblies, and the application of this methodology to two relevant cases, Hsp70 and TGF¿. Starting with high-resolution structures, the developed software will generate alternative conformations using molecular dynamics. The conformations will be grouped into equivalence classes (ECs). Small angle X-ray and/or neutron scattering profiles, radius of gyration and hydrodynamic molecular parameters, such as the sedimentation coefficient, translational and rotational diffusion coefficients, and intrinsic viscosity, will be computed for each EC. The ECs will be globally fitted to experimental data, resulting in a distribution of contributing ECs. This information will provide important structural and dynamic detail obtained under physiological conditions and help to validate corresponding data from other high-resolution techniques. The proposed analysis strategy is based on well-established first principles and has universal applicability to any biological systems asking similar questions. We will test our software on two systems where relevant open questions remain. Hsp70 chaperones are 2-domain proteins that mediate a large number of protein processing and folding reactions. Upon binding ATP or ADP, Hsp70s undergo conformational changes that result in release or binding of their protein substrates. These processes are important in neurodegenerative disease and cancer. No crystal structure of an Hsp70 in an ATP state has ever been determined. A structure of the related protein Hsp110 bound to ATP has been determined and has been proposed to model for the Hsp70:ATP conformation, but this has not been tested directly. Controversy exists regarding the interpretation of the crystal structures of Hsp70s. The conformational state of Hsp70 in its nucleotide-free, ADP, and ATP states, and with and without bound peptide substrates will be analyzed under solution conditions where conformations will not be influenced by crystal packing forces or solvent components that are not physiological. TGF¿s are proteins which regulate cell proliferation, cell differentiation, and expression of extracellular matrix proteins. Three isoforms have arisen in mammals which play prominent roles in human disease, especially cancer. TGF¿1 and -¿3 can have opposing roles in tumor growth and metastasis. Understanding the origins of these differences in activity requires a detailed understanding of their structural dynamics. Dimeric TGF¿ binds with receptors TRII and TRI to signal across cell membranes, and can exist in an open or closed state. The open and closed ternary states TGF¿:TRII:TRI form differing signaling complexes. The proposed analysis method will quantify the extent which TGF¿1 and -¿3 exist in their ternary complexes with their receptors in an open state. For both cases, our method will provide additional detail by quantifying multiple conformations existing in solution rather than insisting on a single state.

描述（由申请人提供）：该资助提出了一种新方法的开发，基于计算方法和在生理溶液条件下进行的实验，以补充生物大分子及其组装体结构测定的高分辨率方法，并将该方法应用于两个相关案例，Hsp 70和TGF？。从高分辨率结构开始，开发的软件将使用分子动力学生成替代构象。构象将被分组为等价类（EC）。将计算每个EC的小角X射线和/或中子散射曲线、回转半径和流体动力学分子参数，如沉降系数、平移和旋转扩散系数以及特性粘度。EC将被全局拟合到实验数据，从而得到贡献EC的分布。这些信息将提供在生理条件下获得的重要结构和动态细节，并有助于验证来自其他高分辨率技术的相应数据。所提出的分析策略是基于完善的第一原则，并具有普遍适用性的任何生物系统提出类似的问题。我们将在两个系统上测试我们的软件，其中仍然存在相关的开放问题。Hsp 70分子伴侣是一种双结构域蛋白，介导大量蛋白质加工和折叠反应。在结合ATP或ADP时，Hsp 70经历构象变化，导致其蛋白质底物的释放或结合。这些过程在神经退行性疾病和癌症中很重要。ATP状态下的Hsp 70的晶体结构还没有被确定。与ATP结合的相关蛋白Hsp 110的结构已被确定，并已被提议为Hsp 70：ATP构象的模型，但这还没有被直接测试。关于热休克蛋白70的晶体结构的解释存在争议。将在溶液条件下分析Hsp 70在其无核苷酸、ADP和ATP状态以及有和无结合肽底物的构象状态，其中构象不受晶体堆积力或非生理性溶剂组分的影响。TGF β是调节细胞增殖、细胞分化和细胞外基质蛋白表达的蛋白质。在哺乳动物中出现了三种亚型，它们在人类疾病，特别是癌症中起着重要作用。TGF β 1和TGF β 3在肿瘤生长和转移中可能具有相反的作用。要了解这些活动差异的起源，需要详细了解它们的结构动力学。二聚体TGF β与受体TRII和TRI结合以跨细胞膜传递信号，并且可以以开放或闭合状态存在。开放和封闭的三元态TGF β：TRII：TRI形成不同的信号复合物。所提出的分析方法将量化TGF β 1和TGF β 3在开放状态下与其受体的三元复合物中存在的程度。对于这两种情况，我们的方法将通过量化溶液中存在的多种构象而不是坚持单一状态来提供额外的细节。