STRUCTURAL STUDIES OF PRION CONFORMATIONAL CONVERSION

点击翻译按钮获取中文摘要

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. This proposal is to continue structural studies using small angle x-ray scattering (SAXS) on oligomeric forms of prion proteins which self associate as a result of conformational conversion from an alpha-helical form to a beta-rich form. Previous studies have led to the formulation of a "trimer-of-dimers" atomic scale model for a hexameric form of the mammalian recombinant PrP which can be compared directly with electron microscope data from negatively stained samples of the scrapie form of the protein obtained directly form animal brains. This comparison allows structural bounds to be placed on the differences between the non-infective recombinant form and the infective disease form. We plan to extend these studies by obtaining higher resolution data with the use of the CCD detector, and also to extend the studies to a protein , Sup-35, from yeast which has prion-like properties. and from which the existence of oligomeric intermediates along the folding pathway has been established by light scattering, em and afm work. We also plan to measure oligomeric forms of a protein fragment srcSH3 for which we have a series of alanine-replacement mutants. Following work of Dobson and collaborators, who have demonstrated that formation of amyloid fibrils appears to be a universal for self-association of polypeptides, we have shown that this protein forms fibrils after a lag phase. Study of the dependence of the formation of oligomers on point mutations, should help in validation of structural models for protein self-association such as domain swapping, or suggest new directions for understanding this fundamental phenomenon which is of importance in diseases such as Alzheimer's and Parkinson's.