Site Specific Interactions and Collagen Self Assembly

位点特异性相互作用和胶原蛋白自组装

• 批准号: 6459210
• 负责人: ANDRZEJ FERTALA
• 金额: $ 18.9万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-13 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6459210
• 关键词: biosensor device; collagen; computer simulation; densitometry; electron microscopy; endopeptidases; enzyme activity; extracellular matrix; gel filtration chromatography; ion exchange chromatography; light microscopy; molecular assembly /self assembly; monomer; protein binding; protein biosynthesis; protein protein interaction; protein purification; site directed mutagenesis; thermodynamics; tissue /cell culture

The ability of proteins to self-assemble into macromolecules is fundamental to all living organisms. Some of the most complex of these structures are the fibrils formed from the triple helical collagen. The precise manner in which the monomers self-assemble into fibrils has not been defined. The present application is based on our observations that the telopeptides of collagen I bind to a specific region spanning amino acids 776 to 822 of the alpha1(1) chain, and that this interaction is critical for collagen self-assembly. Our recent results of the assembly of collagen I and collagen II mutants, strongly support our hypothesis that site-specific interactions between collagen molecules are critical for the fibril formation. We now use the gene-engineered collagen II and collagen IX to study the mechanism of the formation of homotypic and heterotypic fibrils in detail. The broad, long-term goal of this project is to understand fundamental principles of the formation of collagen fibrils in health and disease. The overall hypothesis is that the specific interaction between collagen monomers during fibril assembly is controlled by the binding of telopeptides to specific regions of the triple- helical domain. This binding is critical for the regulation of collagen fibril formation. Moreover, we hypothesize that assembly of heterotypic fibrils is also controlled by the site-specific formation. Moreover we hypothesize that assembly of heterotypic fibrils is also controlled by the site-specific interactions between different collagen types, and that mutations in fibrillar collagens change the site-specific interaction and, as a result, alter the structure of the extracellular matrix. To test the hypotheses, the following specific aims are proposed: (1) To elucidate a site-specific mechanism of the formation of homotypic and heterotypic collagen fibrils, (2) To determine how the site-specific interactions between collagen molecules regulate morphology of fibrils, kinetics and thermodynamics of collagen self-assembly, and (3) To analyze how mutations in fibrillar collagens affect the site-specific mechanism of collagen assembly. The proposed research will yield new information on collagen self-assembly structure of collagen fibrils. Results of the proposed studies will have a wide application in the areas where collagen fibrils play an important role. Examples include cell differentiations and proliferations, development, bone mineralization, tissue remodeling, fibrosis, tissue engineering, and therapy of the heritable diseases of connective tissue.

蛋白质自组装成大分子的能力是所有生物体的基础。这些结构中最复杂的一些是由三螺旋胶原蛋白形成的原纤维。单体自组装成原纤维的精确方式尚未确定。本申请基于我们的观察，即胶原蛋白I的端肽与跨越α1(1)链的氨基酸776至822的特定区域结合，并且这种相互作用对于胶原蛋白自组装至关重要。我们最近对胶原蛋白 I 和胶原蛋白 II 突变体的组装结果有力地支持了我们的假设，即胶原蛋白分子之间的位点特异性相互作用对于原纤维的形成至关重要。我们现在利用基因工程改造的II型胶原和IX型胶原来详细研究同型和异型原纤维的形成机制。该项目的广泛、长期目标是了解健康和疾病中胶原纤维形成的基本原理。总体假设是，原纤维组装过程中胶原单体之间的特异性相互作用是通过端肽与三螺旋结构域的特定区域的结合来控制的。这种结合对于胶原纤维形成的调节至关重要。此外，我们假设异型原纤维的组装也受到位点特异性形成的控制。此外，我们假设异型原纤维的组装也受到不同胶原类型之间的位点特异性相互作用控制，并且原纤维胶原的突变改变了位点特异性相互作用，从而改变了细胞外基质的结构。为了检验这些假设，提出了以下具体目标：（1）阐明同型和异型胶原原纤维形成的位点特异性机制，（2）确定胶原蛋白分子之间的位点特异性相互作用如何调节原纤维的形态、胶原蛋白自组装的动力学和热力学，以及（3）分析纤维状胶原蛋白的突变如何影响胶原蛋白组装的位点特异性机制。拟议的研究将产生有关胶原原纤维的胶原自组装结构的新信息。拟议研究的结果将在胶原纤维发挥重要作用的领域得到广泛应用。例子包括细胞分化和增殖、发育、骨矿化、组织重塑、纤维化、组织工程和结缔组织遗传性疾病的治疗。