CRYSTAL STRUCTURE DETERMINATION OF COLLAGEN-RELATED STRUCTURES

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 IV型胶原是所有后生动物基底膜的主要结构蛋白成分。它含有一个大的C-末端球状结构域(NC1域)，通过它，三条链以链特异的方式结合形成三个螺旋原基。这也在细胞外基质中IV型胶原的超分子组装中发挥着重要作用。在哺乳动物中存在组织特异性的IV型胶原原，α1α1α2原蛋白在肾脏中普遍存在，α3α4α5原蛋白在肾脏中普遍存在。我们实验室长期以来对这个项目的兴趣是确定六聚体形式的NC1结构域的晶体结构以及它们的单体组成，以了解链特异性组装的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，更具体地说，在肾脏中具有重要意义。至少有两种肾脏疾病，即阿波特S综合征和好牧场S综合征，与IV型胶原直接相关。[alpha3，alpha4，alpha5]2和alpha3单体的分子结构对于了解古德巴德综合征发病的分子基础尤为重要。这是一种自身免疫性疾病，抗体特异性地针对单体中的α3NC1域，但当它是[alpha3.alpha4.alpha5]2六聚体的一部分时不是。我们用SSRL收集的BR-MAD数据确定了NC1六聚体[alpha1.alpha1.alpha2]2的第一个晶体结构。我们还收集了α2和α3单体晶体的数据，它们的结构不能用六角体的单体模型用分子置换方法来求解。下一步，我们将尝试用Se或Br作为反常散射体来解决这些结构。我们还得到了[α3α4α5]2NCl六面体的弱衍射晶体，这是用内部源所不能表征的。

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.