BONE MATRIX COLLAGEN ASSEMBLY--ROLE ON TYPE V COLLAGEN

该项目的长期目标是了解骨基质 胶原蛋白组装，特别是 V 型胶原蛋白在骨中的作用 矩阵。 越来越多的证据表明胶原蛋白—— 胶原蛋白相互作用可能在控制纤维方面发挥重要作用 直径并且还提供原纤维网络的增强，从而 指定不同组织的机械特性。 虽然很多 已经收集了有关 V 型胶原蛋白及其在骨骼中的功能的信息 或其他组织仍然知之甚少。 将探索三种方法来理解 V 型的作用 骨基质中的胶原蛋白。 首先是确定潜力 通过分析 V 型胶原蛋白与 1 型胶原蛋白共聚 V 型骨胶原分子间交联的性质。 其次， 确定保留的延伸肽对组织的作用 V型胶原蛋白的形式，第三评估V型胶原蛋白的模式 退化。 V 型骨中所有分子间交联位点 涉及硼氢化钠可还原交联的胶原蛋白以及那些 涉及成熟的3-羟基吡啶啉的情况将被确定。 的 V型胶原蛋白中的可还原交联将在反应后确定 胶原蛋白与氚硼氢化钠标记交联 债券。涉及 3-羟基吡啶鎓残基的交联肽 V 型胶原蛋白将进行荧光检测、纯化和 进行蛋白质微测序分析。 扩展的作用 将评估保留在 V 型胶原组织形式上的肽 通过细菌从 α 链产生延伸肽 胶原酶消化，然后产生单特异性抗体 延伸肽。 生成的抗体将用于 检查这些延伸肽在骨胶原原纤维中的作用。 V型胶原酶降解V型胶原的方式为 通过将天然 V 型胶原蛋白与 V 型胶原酶一起孵育来测定。 V 型胶原蛋白内的初始攻击位点将被确定 通过对生成的片段进行氨基末端序列分析。 了解骨基质胶原蛋白组装可能有助于解释如何 成骨不全症中 V 型与 I 型胶原的比例增加 患者可能会损害骨组织的结构完整性并有助于 机械弱点。

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.