Structure-Function of Type IV Collagen NC1 Domains

描述(申请人提供)：肾小球基底膜的结构 在影响肾脏的三种值得注意的疾病中，膜(GBM)会发生变化： 古德帕斯普综合征(抗肾小球基底膜肾炎)、阿尔波特综合征(遗传性 肾炎)和糖尿病。至少有两家，古德帕斯图尔和阿尔波特 综合征与IV型胶原有关，IV型胶原是GBM的主要成分。 IV型胶原是一个由六个α链组成的家族，α1-α6，发现于 各种基底膜(BMS)。三条阿尔法链形成一个胶原蛋白三元组 螺旋原基，每个链都含有一个非胶原性(Nc1)球状结构域 在C-终端。原始体的链化学计量比随 组织的起源，(Alpha1)2.alpha2是普遍存在的形式和 Alpha3、Alpha4、Alpha5为肾小球和肺泡特异。两个三人组 螺旋原基结合在Nc1区域形成尾对尾二聚体。这个 Nc1结构域被推定为包含这两个三元组的结构决定因素 螺旋的形成和六聚体的组装。Alpha3 Nc1结构域含有表位 古德帕斯图尔综合征患者的自身抗体。基因突变 编码三个阿尔法链中的任何一个的基因会导致 Alpha3.alpha4.alpha5在基底核中的网络，这是Alport综合征的原因。 重组α2、α3和α6 NC1结构域显示出抗血管生成和 抗肿瘤特性。在这项研究中，我们提出了以下具体目标 确定NC1域的结构-功能关系： 目的1：确定NC1六聚体的第一晶体结构， ((字母1)2.字母2)2 目的2：结晶并确定GBM NC1六聚体的结构， (字母3.alpha4.alpha5)2 目的3：确定Goodstraure抗原α3 Nc1的结构 单体 目的4：解决血管生成因子α2和α6的结构 抑制剂。 这些目标的实现需要高分子材料的应用。 结晶学、计算生物学、蛋白质化学和分子 生物学。

DESCRIPTION (provided by applicant): The structure of glomerular basement membrane (GBM) is altered in three notable diseases that affect the kidneys: Goodpasture syndrome (anti-GBM nephritis), Alport syndrome (hereditary nephritis), and diabetes mellitus. At least two of them, Goodpasture and Alport syndromes, have been linked to type IV collagen, a major constituent of GBM. Type IV collagen is a family of six alpha chains, alpha1-alpha6, found in various basement membranes (BMs). Three alpha chains form a collagen triple helical protomer and each chain contains a noncollagenous (NC1) globular domain at the C-terminus. The chain stoichiometry of the protomer varies with the origin of the tissue, (alpha1)2.alpha2 being the ubiquitous form and alpha3.alpha4.alpha5 being specific to glomeruli and alveoli. Two triple helical protomers associate at the NC1 region to form tail-to-tail dimer. The NC1 domains are presumed to contain the structural determinants for both triple helix formation and hexamer assembly. The alpha3 NC1 domain harbors the epitope for autoantibodies in patients with Goodpasture syndrome. Mutations in the genes encoding any of the three alpha chains result in the loss of alpha3.alpha4.alpha5 network in the GBM, which is the cause of Alport syndrome. Recombinant alpha2, alpha3, and alpha6 NC1 domains show anti-angiogenic and anti-tumor properties. In this study, we propose the following specific aims to determine the structure-function relationships of NC1 domains: Aim 1: To determine the first crystal structure of a NC1 hexamer, ((alpha1)2.alpha2)2 Aim 2: To crystallize and determine the structure of GBM NC1 hexamer, (alpha3.alpha4.alpha5)2 Aim 3: To determine the structure of the Goodpasture antigen, alpha3 NC1 monomer Aim 4: To solve the structures of alpha2 and alpha6 monomers, the angiogenic inhibitors. The accomplishment of these aims requires the application of macromolecular crystallography, computational biology, protein chemistry and molecular biology.