BONE MATRIX COLLAGEN ASSEMBLY--ROLE ON TYPE V COLLAGEN

骨基质胶原蛋白组装体 - V 型胶原蛋白的作用

• 批准号: 2082154
• 负责人: CHRISTOPHER NIYIBIZI
• 金额: $ 10.02万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-03-01 至 1998-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2082154
• 关键词: biosynthesis; bone density; cell type; collagen; crosslink; extracellular matrix; human subject; mechanical pressure; osteogenesis imperfecta; peptides; polymerization; protein degradation; protein sequence

The long term objective of the project is to understand the bone matrix collagen assembly, specifically the role of type V collagen in bone matrix. There is a growing body of evidence indicating that collagen- collagen interactions may play an important role in controlling fiber diameter and also in providing fibril network with reinforcement, thereby specifying the mechanical properties of different tissues. Although a lot of information has been gathered on type V collagen, its function in bone or other tissues remains poorly understood. Three approaches will be explored for understanding the role of type V collagen in bone matrix. The first will be to determine the potential of type V collagen to copolymerize with type 1 collagen by analysis of the nature of intermolecular cross-linking in bone type V collagen. Secondly, to determine the role of the retained extension peptides on the tissue form of type V collagen, and thirdly to assess the mode of type V collagen degradation. All the intermolecular cross-linking sites in bone type V collagen involving sodium borohydride reducible cross-links and also those involving the mature 3-hydroxypyridinolines will be determined. The reducible cross-links in type V collagen will be determined after reacting the collagen with tritiated sodium borohydride to label the cross-linking bonds. Cross-linked peptides involving 3-hydroxypyridinium residues in type V collagen will be followed by their fluorescence, purified and subjected to protein microsequencing analysis. The role of the extension peptides retained on the tissue form of type V collagen will be assessed by generating the extension peptides from the alpha chains by bacterial collagenase digestion and then producing monospecific antibodies against the extension peptides. The generated antibodies will then be used to examine the role of these extension peptides in bone collagen fibrils. The mode of type V collagen degradation by type V collagenase will be determined by incubating native type V collagen with type V collagenase. The initial sites of attack within the type V collagen will be determined by aminoterminal sequence analysis of the generated fragments. Understanding the bone matrix collagen assembly may help to explain how increased ratio of type V to type I collagen in osteogenesis imperfecta patients may impair structural integrity of the bone tissue and contribute to mechanical weakness.

该项目的长期目标是了解骨基质 胶原蛋白组装，特别是V型胶原蛋白在骨骼中的作用 矩阵。越来越多的证据表明，胶原蛋白- 胶原蛋白的相互作用可能在控制纤维方面发挥重要作用 并且还在提供具有增强的纤维网络中，从而 指定不同组织的机械性能。虽然有很多 已经收集了关于V型胶原的信息，它在骨骼中的功能 或其他组织仍然知之甚少。 我们将探索三种方法来理解类型V的作用 骨基质中的胶原。第一个将是确定 通过分析V型胶原与1型胶原的共聚 骨V型胶原分子间交联性的研究。第二， 确定保留的延伸肽对组织的作用 V型胶原的形态，以及V型胶原的模式 退化。V型骨中所有分子间的交联点 含有硼氢化钠可还原交联剂的胶原蛋白 涉及成熟的3-羟基吡啶类化合物将被确定。这个 反应后将确定V型胶原中可还原的交联物 用氚化硼氢化钠标记的胶原蛋白 债券。含3-羟基吡啶残基的交联肽 V型胶原之后将用它们的荧光，纯化和 进行蛋白质微测序分析。外延的作用 V型胶原组织形式上保留的多肽将被评估 通过细菌从阿尔法链中产生延伸肽 胶原酶消化，然后产生单特异性抗体 延伸肽。产生的抗体将被用来 研究这些延伸肽在骨胶原纤维中的作用。 V型胶原酶降解V型胶原的方式为 通过将天然的V型胶原与V型胶原酶孵育而确定。 V型胶原的初始攻击部位将被确定 通过对产生的片段进行氨基末端序列分析。 了解骨基质胶原蛋白的组装可能有助于解释 成骨不全患者V/I型胶原比值升高 患者可能会损害骨组织的结构完整性，并对 机械上的弱点。