CRYSTAL STRUCTURE DETERMINATION OF COLLAGEN-RELATED STRUCTURES

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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.