Elucidation of Prion Protein Conformational Changes Associated With Infectivity by Fluorescence Spectroscopy
Annual rept. 15 May 2003-14 May 2004
MONTANA UNIV MISSOULA
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Prion diseases are fatal neurodegenerative diseases of mammals. They are characterized by the conversion of normal prion protein PrP to a misfolded conformational state that accumulates as plaques in the brain. The diagnosis of prion diseases relies on the ability to differentiate between normal PrP and its misfolded, infectious form. This is difficult to accomplish by traditional testing methods, since it requires discerning between conformational states of a protein that is present in both normal and diseased tissue, rather than identifying the appearance of a new protein associated with infection. We wish to design a reporter PrP substrate that may be monitored by fluorescence spectroscopy. After the conversion of normal-PrP to its infectious state, some amino acid residues of PrP will undergo a change in their local solvent environment. We propose to identify these residues by monitoring the fluorescence emission spectrum of a series of mutant 7-AzaTrp-substituted PrP proteins. The 7-AzaTrp fluorescence emission spectrum is both unique compared with normal Trp and exquisitely sensitive to its local environment. This could lead to the development of a rapid, sensitive, and inexpensive technique to detect infectious PrP, based on its ability to bind 7-AzaTrp-substituted PrP, and convert it to the misfolded form.