Integrin alphaIIbbeta3 Structure, Activation, and Ligand Binding

整合素 alphaIIbbeta3 结构、激活和配体结合

• 批准号: 9086391
• 负责人: Barry Coller
• 金额: $ 62.18万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1976
• 资助国家: 美国
• 起止时间: 1976-09-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9086391
• 关键词: 3-Dimensional; Accounting; Address; Adhesions; Algorithms; Applications Grants; Aspirin; Binding; Binding Sites; Biological Assay; Biology; Blood Platelets; Blood Vessels; C-terminal; Cardiovascular Diseases; Caring; Cell Line; Cells; Chemicals; Clinical; Complementary DNA; Computer Simulation; Computing Methodologies; Cryoelectron Microscopy; Cytoplasmic Tail; DNA; DNA Double Strand Break; DNA Repair; Data; Detergents; Development; Diagnosis; Disease; Docking; Drug Targeting; Electron Microscopy; Family; Fibrinogen; Genes; Glycoproteins; Goals; Grant; Head; Health; Hemostatic function; Hospitals; Human; Inflammation; Integrins; Knowledge; Lead; Ligand Binding; Ligands; Lipid Bilayers; Mass Spectrum Analysis; Mediating; Medical; Methods; Modeling; Modification; Molecular; Molecular Conformation; Monitor; Monoclonal Antibodies; Mus; Mutate; Mutation; Myocardial Infarction; National Heart, Lung, and Blood Institute; Natural Immunity; Negative Staining; Neoplasm Metastasis; Pathway interactions; Patients; Pattern; Pharmaceutical Chemistry; Phase; Phenotype; Physiology; Plasma; Platelet Glycoprotein GPIIb-IIIa Complex; Platelet Glycoproteins; Platelet aggregation; Play; Prenatal Diagnosis; Proteins; RGD (sequence); Radial; Resolution; Role; Science; Signal Transduction; Stroke; Structure; Surface; Talin; Techniques; Technology; Testing; Therapeutic; Thrombasthenia; Thrombocytopenia; Thrombosis; Translating; Translations; Wound Healing; X-Ray Crystallography; abciximab; base; carrier testing; combinatorial chemistry; design; filamin; flexibility; glycocalicin; high throughput screening; improved; inhibitor/antagonist; insight; molecular dynamics; mutant; nanodisk; novel; novel therapeutics; prevent; programs; receptor; reconstruction; screening; virtual; zinc finger nuclease

DESCRIPTION (provided by applicant): Platelets play a central role in both hemostasis and thrombosis, and contribute to a wide range of related phenomena, including inflammation and metastasis formation. The ultimate goal of this grant proposal is to understanding the way in which platelets interact with the blood vessel wall and with other platelets via the glycoprotein receptors on their surface, and to use that knowledge to improve human health. Despite advances in analyzing the structure and function of the αIIbβ3 receptor, which is crucial for normal hemostasis and a validated target of antithrombotic therapy, major gaps remain in understanding ligand binding and its impact on platelet physiology. Moreover, there is a need for improved potent antiplatelet therapies that cn be administered in the pre-hospital phase of myocardial infarction. The data obtained from these studies will inform attempts to develop novel inhibitors of αIIbβ3 that have therapeutic advantages over existing agents. In Specific Aim 1 we propose to improve our understanding of ligand binding by: a) Using functional ligand binding data and new crystal structures of αIIbβ3 as inputs to state-of-the art computational methods to identify interactions between the fibrinogen γ-module and αIIbβ3 in addition to those made by the fibrinogen γ-chain C-terminal dodecapeptide (γC-12), b) Using electron cryomicroscopy (cryo-EM) and negative stain EM in conjunction with random conical tilt reconstructions, molecular dynamics (MD)-based flexible fitting and steered MD to obtain atomic resolution 3-dimensional (3D) representations of intact αIIbβ3 in a nanodisc lipid bilayer in the absence of detergent. The inactive receptor, the receptor activated by talin head-domain (THD) in the absence of ligand, and THD-activated receptor in the presence of fibrinogen will each be studied. c) Using zinc finger nuclease-mediated gene editing to evaluate mutations of β3 in murine platelets rather than in cell lines that lack the platelet's signaling machinery, focusing initially on a mutation we hypothesize will enhance the binding of filamin to the β3 cytoplasmic tail and thus diminish platelet sensitivity to activation. d) Using enhanced MD techniques to characterize αIIbβ3 activation pathways and testing the hypothesis that the new αIIbβ3 antagonists identified in the past grant period (RUC-1, RUC-2, MSSM-1, MSSM-2) stabilize a closed, inactive conformation, thus accounting for their reduced ability to activate αIIbβ3 compared to αIIbβ3 antagonists patterned on the Arg-Gly-Asp (RGD) cell recognition sequence. In Specific Aim 2 we propose to identify new compounds that will provide insights into αIIbβ3 structure-function and may have therapeutic potential by characterizing the 57 compounds (out of 126,000 tested) that most potently inhibit αIIbβ3-mediated platelet adhesion and aggregation. We will also perform a new screen to selectively identify compounds that target ancillary fibrinogen binding sites since these may lead to new therapeutics that are safer and more efficacious.

描述（由申请人提供）：血小板在止血和血栓形成中发挥核心作用，并有助于广泛的相关现象，包括炎症和转移的形成。这项拨款提案的最终目标是了解血小板通过其表面的糖蛋白受体与血管壁和其他血小板相互作用的方式，并利用这些知识来改善人类健康。αIIbβ3受体对正常止血至关重要，是抗血栓治疗的有效靶点，尽管在分析αIIbβ3受体的结构和功能方面取得了进展，但在了解配体结合及其对血小板生理的影响方面仍存在重大空白。此外，还需要改进有效的抗血小板治疗方法，以便在心肌梗死院前阶段给予治疗。从