CRYSTAL STRUCTURE DETERMINATION OF COLLAGEN-RELATED STRUCTURES

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 中心，不一定是研究者的机构。 IV型胶原是所有后生动物基底膜的主要结构蛋白组分。它含有一个大的C-末端球状结构域（NC 1结构域），通过该结构域，三条链以链特异性方式缔合形成三螺旋原聚体。这也在细胞外基质中IV型胶原的超分子组装中起主要作用。IV型胶原的组织特异性原聚体类型存在于哺乳动物中，α 1. α 1. α 2原聚体普遍存在，α 3. α 4. α 5原聚体存在于肾脏中。我们实验室在这个项目中的长期兴趣是确定六聚体形式的NC 1结构域的晶体结构以及它们的单体组分，以了解链特异性组装IV型胶原。这些结构将对理解IV型胶原在组织发育中的作用具有重要意义，更具体地说，肾脏。至少有两种肾脏疾病，即Alport综合征和Goodpasture综合征，与IV型胶原蛋白直接相关。[α 3. α 4. α 5]2和α 3单体的分子结构在理解肺出血肾炎综合征发病机制的分子基础方面尤其重要。这是一种自身免疫性疾病，其中抗体特异性靶向单体中的α 3 NC 1结构域，但当它是[α 3. α 4. α 5]2六聚体的一部分时不靶向。我们已经使用在SSRL收集的Br-MAD数据确定了NC 1六聚体的第一晶体结构[α 1 α 1 α 2]2。我们还收集了α 2和α 3单体晶体的数据，这些晶体的结构无法通过使用六聚体的单体模型的分子置换方法来解决。接下来，我们将尝试使用Se或Br作为异常散射体通过MAD来解决这些结构。 我们还获得了[α 3. α 4. α 5]2 NC 1六聚体的弱衍射晶体，其不能使用内部来源表征。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Type IV collagen is a major structural protein component of basement membrane of all metazoan. It contains a large C-terminal globular domain (NC1 domain) by which three chains associate to form triple helical protomers in a chain specific manner. This also plays a major role in the supramolecular assembly of collagen type IV in the extracellular matrix. Tissue specific protomer types of collagen IV exist in mammals, with alpha1.alpha1.alpha2 protomer being ubiquitous and alpha3.alpha4.alpha5 protomer in kidney. The long standing interest of our lab in this project is to determine the crystal structures of NC1 domains in hexameric forms as well as their monomeric components to understand the chain specific assembly type IV collagen. These structures will have strong implications in understanding the role of type IV collagen in tissue development, more specifically, kidney. At least two kidney diseases, namely Alport¿s syndrome and Goodpasture¿s syndrome, are directly linked to type IV collagen. The molecular structures of [alpha3.alpha4.alpha5]2 and alpha3 monomer are especially crucial in understanding the molecular basis for the pathogenesis of Goodpasture syndrome. This is an autoimmune disease in which the antibodies specifically target alpha3 NC1 domain in the monomer, but not when it is a part of [alpha3.alpha4.alpha5]2 hexamer. We have determined the first crystal structure of NC1 hexamer, [alpha1.alpha1.alpha2]2 using Br-MAD data collected at SSRL. We have also collected data on alpha2 and alpha3 monomer crystals, whose structures could not be solved by molecular replacement method using the monomer models from the hexamer. Next, we would attempt to solve these structures by MAD using Se or Br as anomalous scatterer. We have also obtained weakly diffracting crystals of [alpha3.alpha4.alpha5]2 NC1 hexamer, which could not be characterized using the in-house source.