| First Author | Gaspersic J | Year | 2010 |
| Journal | FEBS J | Volume | 277 |
| Issue | 9 | Pages | 2038-50 |
| PubMed ID | 20345906 | Mgi Jnum | J:200371 |
| Mgi Id | MGI:5508565 | Doi | 10.1111/j.1742-4658.2010.07619.x |
| Citation | Gaspersic J, et al. (2010) Tetracysteine-tagged prion protein allows discrimination between the native and converted forms. FEBS J 277(9):2038-50 |
| abstractText | The conformational conversion of prion protein (PrP) from a native conformation to the amyloid form is a hallmark of transmissible spongiform encephalopathies. Conversion is usually monitored by fluorescent dyes, which bind generic amyloids and are less suited for living cell imaging. We report a new method for the synthesis of membrane-permeable and membrane-impermeable biarsenical reagents, which are then used to monitor murine PrP (mPrP) misfolding. We introduced tetracysteine (TC) tags into three different positions of mPrP, which folded into a native-like structure. Whereas mPrPs with a TC tag inserted at the N-terminus or C-terminus supported fibril formation, insertion into the helix 2-helix 3 loop inhibited conversion. We devised a quantitative protease-free method to determine the fraction of converted PrP, based on the ability of the fluorescein arsenical helix binder reagent to differentiate between the monomeric and fibrilized form of TC-tagged PrP, and showed that TC-tagged mPrP could be detected on transfected cells, thereby expanding the potential use of this method for the detection and study of conformational diseases. |
