Protein Interactions and Conformational Changes in Fibronectin Fibril Formation

纤连蛋白原纤维形成中的蛋白质相互作用和构象变化

• 批准号: 9333423
• 负责人: Klara Briknarova
• 金额: $ 28.4万
• 依托单位: UNIVERSITY OF MONTANA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9333423
• 关键词: Affinity; Age related macular degeneration; Angiogenesis Inhibition; Angiogenesis Inhibitors; Architecture; Atherosclerosis; Binding; Binding Sites; Biological Models; Blood Vessels; Cell Adhesion; Cells; Chronic; Complex; Cysteine; Data; Deposition; Development; Diabetic Retinopathy; Disease; Disulfides; Energy Transfer; Extracellular Matrix; Extracellular Matrix Proteins; Fibronectins; Generations; Goals; Growth; In Vitro; Investigation; Label; Lesion; Light; Malignant Neoplasms; Maps; Microscopic; Molecular; Molecular Conformation; Molecular Models; NMR Spectroscopy; Neoplasm Metastasis; Pathologic; Pathologic Neovascularization; Pathologic Processes; Physiological Processes; Play; Process; Proteins; Proteolysis; Recruitment Activity; Regulation; Relaxation; Reproduction; Resolution; Rheumatoid Arthritis; Role; Signal Transduction; Site; Site-Directed Mutagenesis; Structural Protein; Structure; Vascular blood supply; Wound Healing; X-Ray Crystallography; angiogenesis; base; beta pleated sheet; crosslink; fibrillogenesis; in vivo; inhibitor/antagonist; insight; migration; molecular modeling; molecular rearrangement; mouse model; mutant; neoplastic cell; new growth; novel anticancer drug; public health relevance; single molecule; tumor; tumor growth

 DESCRIPTION (provided by applicant): The generation of a lethal tumor mass requires that the tumor cells recruit and sustain their own blood supply. In the absence of blood supply, tumors can persist as dormant microscopic lesions but they do not expand beyond the size of 1-2 mm. Hence, inhibition of new blood vessel growth (angiogenesis) offers great promise to eradicate tumors or to convert cancer to a chronic manageable disease. In addition, inhibition of angiogenesis offers great promise to treat a range of other diseases that depend on angiogenesis, including age-related macular degeneration, diabetic retinopathy, rheumatoid arthritis and atherosclerosis. The focus of this application is anastellin, a fragment of the extracellular matrix protein fibronectin that inhibits angiogenesis, tumor growth and metastasis in mouse models. Anastellin requires endogenous fibronectin for its in vivo anti- angiogenic activity, and it binds to fibronectin in vitro and converts the soluble protein to insoluble fibril. Our long term goal is to elucidate how the interaction of anastellin with fibronectin leads to formation of fibrillar aggregates, and in general how the structure of fibronectin determines its function. The goal of this application is to identify the molecular basis of the interaction betwee anastellin and fibronectin. We propose to investigate the structure and dynamics of the complexes between anastellin and its target fibronectin type III (FN3) domains by disulfide crosslinking, nuclear magnetic resonance (NMR) spectroscopy and X-ray crystallography. In addition, we propose to investigate the conformation of the anastellin binding region in fibronectin using paramagnetic relaxation enhancement (PRE) and single molecule Förster resonance energy transfer (FRET). Identification of structural features that are responsible for the activity of anastellin will provide detailed understanding of how this inhibitor of angiogenesi acts at the molecular level and may enable development of new anticancer drugs. The proposed structural studies will also provide insight into the molecular interactions and rearrangements that are involved in conversion of soluble fibronectin to the fibrillar matrix form.

 描述（由申请人提供）：致死肿瘤块的产生需要肿瘤细胞募集并维持自身的血液供应。在没有血液供应的情况下，肿瘤可以作为休眠的微观病变持续存在，但它们不会扩展超过1-2 mm的大小。因此，抑制新血管生长（血管生成）为根除肿瘤或将癌症转化为慢性可控制疾病提供了很大的希望。此外，抑制血管生成为治疗一系列依赖于血管生成的其他疾病提供了巨大的希望，包括年龄相关性黄斑变性、糖尿病视网膜病变、类风湿性关节炎和动脉粥样硬化。本申请的重点是anastellin，其是细胞外基质蛋白纤连蛋白的片段，其在小鼠模型中抑制血管生成、肿瘤生长和转移。Anastellin需要内源性纤连蛋白用于其体内抗血管生成活性，并且其在体外结合纤连蛋白并将可溶性蛋白质转化为不溶性原纤维。我们的长期目标是阐明anastellin与纤连蛋白的相互作用如何导致纤维状聚集体的形成，以及纤连蛋白的结构如何决定其功能。本申请的目的是鉴定anastellin和纤连蛋白之间相互作用的分子基础。我们建议通过二硫键交联，核磁共振（NMR）光谱和X-射线晶体学研究anastellin和其靶向纤连蛋白III型（FN 3）结构域之间的复合物的结构和动力学。此外，我们建议使用顺磁弛豫增强（PRE）和单分子Förster共振能量转移（FRET）研究纤连蛋白中anastellin结合区的构象。负责anastellin活性的结构特征的鉴定将提供对这种血管生成抑制剂如何在分子水平上起作用的详细理解，并可能使新的抗癌药物的开发成为可能。拟议的结构研究也将提供深入了解的分子相互作用和重排，参与可溶性纤连蛋白的纤维基质形式的转换。