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Studies of protein folding have had a major impact on the understanding of protein function and the control of human and animal disease. Not only have a number of diseases been attributed to protein folding defects, but the biophysical characterizations of the folding properties suggest therapeutic strategies for treatment. For example, the deltaF508 mutation of the cystic fibrosis transmembrane conductance regulator (CFTR) has been shown to be a functional chloride channel, but structural alterations prevent its release from the ER. Thus, developing ways to facilitate removal from the ER has important clinical applications in the treatment of cystic fibrosis. Research into the thermodynamic and kinetic properties of the transition of prion protein leads to an understanding of the various forms of transmissible spongiform encephalopathies (TSEs) and can be generalized to other protein aggregation and deposition disorders such as Alzheimer's disease, systemic amyloidosis, and type II diabetes. The pathogenic and infectious form of prion protein, PrPSC, is able to aggregate and form amyloid fibrils, which are very stable and resistant to most disinfecting processes and common proteases. Bovine spongiform encephalopathy (BSE) is linked to the human disease variant Creutzfeldt-Jakob disease (vCJD), and the cost of BSE to European Union member states is estimated at more than 90 billion euros. Because the US beef industry is a $76 billion industry (in 2008), the effects of BSE are potentially devastating to the industry as well as potential health risks to humans.
NON-TECHNICAL SUMMARY: Bovine spongiform encephalopathy (BSE) is linked to the human disease variant Creutzfeldt-Jakob disease (vCJD), and the cost of BSE to European Union member states is estimated at more than 90 billion euros. Because the US beef industry is a $76 billion industry (in 2008), the effects of BSE are potentially devastating to the industry as well as potential health risks to humans. Work in our laboratory has the potential to affect therapeutic strategies for a number of protein aggregation diseases, from Alzheimer?s to Creutzfeldt-Jakob disease to bovine spongiform encephalopathy, because the dysregulation of protein aggregation is a common factor to these diseases. The identification of small molecules that bind to the interfaces of the protein oligomers, resulting in their dissolution is at the forefront of our research.
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APPROACH: Under specific conditions, PrPSC in BSE brain tissue was found degradable by a bacterial keratinase and some other proteases. Since the disease-causing prion is infectious and dangerous to work with, model or surrogate proteins are examined in this project that are safe. We have examined three proteins in our studies: yeast Sup35NM, Apaf-1 CARD, and procaspase-3. The three proteins assemble into higher order structures that can be used for modeling protein assembly. Methods used include fluorescence and circular dichroism spectroscopies, x-ray crystallography, and cell biological assays for protein aggregation and activity. Thermodynamic and kinetic studies are used to examine assembly and aggregation pathways for the proteins.