This is a competing renewal of Merit Grant AR02594, which originally focused on the structure of immunoglubulins and their role in systemic amyloidosis. Although these investigations elucidated important structural data, the hypervariability of immunoglobulins and the lack of reliable animal models lead us to extend our studies to cerebral amyloidoses, and in particular the prion related disorders for which there are better model system. However, the central question we are addressing remains the same: What are the factors producing disease associated, abnormal protein conformation. The amyloidoses and several degenerative conditions are disorders of protein conformation, whereby a normal protein is structurally altered and is associated with the disease state. The prion diseases are the best defined human conformational diseases. Central to the etiology of prion disorders is the conversion of normal prion protein, PrP/C, into its infectious and pathogenic form, PrP/Sc. Current data indicate that PrP/c and PrP/Sc differ only in their conformation, with PrP/Sc have a greater beta-sheet content. However, identification of numerous """"""""strains"""""""" of prions suggest the existence of multiple PrP/Sc conformations; alternatively, undefined co-factors may have a role in infectivity and strain specificity. We have postulated and demonstrated previously that molecular chaperones can bind and alter protein conformation leading to amyloidogenesis.
In specific aim 1, we propose to identify PrP/Sc binding proteins and study their role in infectivity. and their strain-specific conformation properties. The existence of multiple conformations will be assessed by circular dichroism, FT-IR and proteinase K resistance of the isolated strains. The effect of PrP/Sc binding proteins on PrP/c conformation in vitro will be correlated with infectivity in vivo.
In specific aim 2 we propose to construct synthetic peptides or mini chaperones capable of reversing pathologic protein conformation in order to modify PrP/Sc species in vitro and determine the effect of in vivo infectivity. Purified PrP/Sc, pretreated with mini-chaperone peptides specifically designed to bind and alter prior conformation will be assessed for alterations in infectivity and strain behavior. We shall test whether similar peptides can be used for in vivo imaging of PrP/Sc, utilizing [I/125]-labeling followed by autoradiography pr [19F]-labeling in conjunction with magnetic resonance spectroscopy. Our long-term goals are to define the structural features critical for the formation of PrP/Sc, which will have implications for the design rationale of effective pharmacological and diagnostic tools, as well as deepen our understanding of the basic mechanism(s) underlying conformational diseases.
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