DETERMINANTS OF STABILITY AND INTEGRIN BINDING WITHIN COLLAGEN IV NETWORKS

在目前的资助期间，有三项发现作为目前更新的基础： a）鉴定新的交联，其涉及两个翻译后修饰， B）鉴定控制胶原蛋白IV网络的非胶原蛋白1（NC 1）识别结构域; 胶原蛋白IV网络的组装;和c）鉴定胶原蛋白IV中整合素的新结合位点 整合素α 1 <$1和α 2 <$1在调节肾脏IV型胶原稳态中的关键作用 肾小球在健康和疾病。特别重要的是，识别域现在为我们提供了 一种新的策略，工程师，第一次，组装三螺旋异源三聚体的研究， IV型胶原网络中的新型整联蛋白结合位点。这个更新项目的中心假设是： 在胶原IV网络中，存在稳定这些多价配体的结构的交联位点，并且 赋予α 1 <$1和α 2 <$1整联蛋白结合的不同位点。在这份更新申请中，我们将阐明 两种肾小球IV型胶原蛋白网络在分子和原子水平上的关键相互作用， 整合素为了验证这一假设，我们提出了以下目标：目标1）推导出化学 新型羟赖氨酸-甲硫氨酸交联IV型胶原的结构及形成机制 网络.我们假设甲硫氨酸和羟赖氨酸残基通过共价键连接， 锍结构，并且羟基化由赖氨酰羟化酶的一种同种型催化。目标2） 在a1 a1 a2网络中定位和表征新的a1 <$1和a2 <$1整合素结合位点。我们 假设IV型胶原α 1 α 1 α 2网络具有α 1 β 1和α 2 β 1整联蛋白不同结合位点。 目的3）定位和表征整合素α 3 α 4 α 5网络中的结合位点。我们假设 整合素A1 <$1和A2 <$1在A3 A4 A5网络中的不同结合位点可介导特异性肾小球 功能协调发展的 我们预计，这项研究将产生新的见解的结构基础，胶原蛋白IV 网络结合整联蛋白A1 <$1和A2 <$1。这一关键事件对于维持 肾小球功能这些研究需要整合素、胶原蛋白、细胞和细胞因子之间的协同努力。 结构生物学家

Three discoveries were made during the current grant that serve as a basis for the present renewal: a) the identification of novel crosslinks which involve two post-translational modifications that stabilize collagen IV networks; b) identification of non-collagenous 1 (NC1) recognition domains governing the assembly of collagen IV networks; and c) identification of novel binding sites for integrins in collagen IV networks and a critical role for a1¿1 and a2¿1 integrins in modulating collagen IV homeostasis in kidney glomerulus in health and disease. Of particular importance, the recognition domains now provide us with a novel strategy to engineer, for the first time, the assembly of triple helical heterotrimers for studies of novel integrin binding sites in collagen IV networks. The central hypothesis of this renewal project is: On collagen IV networks, there are crosslink sites that stabilize the structure of these multivalent ligands, and distinct sites that confer binding of a1¿1 and a2¿1 integrins. In this renewal application, we will elucidate critical interactions at the molecular and atomic level of the two glomerular collagen IV networks and integrins. To test this hypothesis, we propose the following aims: Aim 1) To deduce the chemical structure and the mechanism of formation of the novel hydroxylysine-methionine crosslink of collagen IV networks. We hypothesize that methionine and hydroxylysine residues are covalently connected through a sulfonium structure, and that the hydroxylation is catalyzed by one isoform of lysyl hydroxylase. Aim 2) To map and characterize the novel a1¿1 and a2¿1 integrin binding sites in the a1a1a2 network. We hypothesize that a1a1a2 network of collagen IV possess distinct binding sites for alpl and a2p1 integrin. Aim 3) To map and characterize integrin binding sites in the a3a4a5 network. We hypothesize that distinct binding sites for a1¿1 and a2¿1 integrin within a3a4a5 network could mediate specific glomerular functions. We anticipate that this study will generate novel insights into the structural basis whereby collagen IV networks bind integrins a1¿1 and a2¿1. This critical event is fundamental for the maintenance of glomerular function. These studies require a collaborative effort among integrin, collagen, cell, and structural biologists.