Structure-Function of Type IV Collagen NC1 Domains

描述（申请人提供）：肾小球基底结构 膜（GBM）在三种影响肾脏的显著疾病中发生改变： Goodpasture综合征（抗GBM肾炎）、Alport综合征（遗传性 肾炎）和糖尿病。其中至少有两个，古德帕斯彻和奥尔波特 综合征，已与IV型胶原蛋白，GBM的主要成分。 IV型胶原蛋白是六条α链的家族，α 1-α 6，发现于 各种基底膜（BM）。三条α链形成胶原蛋白三重体 螺旋状原聚体，每条链含有一个非胶原（NC 1）球状结构域 在C端。原异构体的链化学计量随着分子量的变化而变化。 组织的起源，（α 1）2·α 2是普遍存在的形式， α 3、α 4、α 5对肾小球和肺泡具有特异性。两个三 螺旋原体在NC 1区结合形成尾对尾二聚体。的 NC 1结构域被认为包含两个三联体的结构决定因素， 螺旋形成和六聚体组装。α 3 NC 1结构域具有表位 肺出血肾炎综合征患者的自身抗体突变 编码三种α链中任何一种的基因都会导致 α 3. α 4. α 5网络，这是Alport综合征的原因。 重组α 2、α 3和α 6 NC 1结构域显示抗血管生成和 抗肿瘤特性。在本研究中，我们提出了以下具体目标， 确定NC 1结构域的结构-功能关系： 目的1：确定NC 1六聚体的第一晶体结构， （（alpha1）2.alpha2）2 目的2：对GBM NC 1六聚体进行结晶和结构测定， （α 3. α 4. α 5）2 目的3：确定Goodpasture抗原α 3 NC 1的结构 单体 目的4：为了解决α 2和α 6单体的结构， 抑制剂的 这些目标的实现需要应用高分子材料， 晶体学、计算生物学、蛋白质化学和分子生物学 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.