Generation of Infectious Prions Amenable to Site-Specific Click Chemistry

Prion diseases are a group of fatal neurodegenerative diseases that proceed through the templated conversion of the normal PrP^C protein to a self-propagating and infectious form termed PrP^Sc. This conversion process is central to the disease progression. However, because of difficulties in producing functional PrP^Sc molecules that can be selectively modified with chemical probes, many aspects of PrP^Sc biology cannot be directly studied. To overcome this limitation, we substituted p-azido-l-phenylalanine (AzF), a small click chemistry-reactive amino acid, for tryptophan residue 99 of PrP^C. The W99AzF PrP^C substrate can efficiently and faithfully propagate either infectious or noninfectious PrP^Sc conformers in vitro. Critically, W99AzF PrP^Sc remains amenable to click chemistry by various ligands after the prion conversion process. Through the combination of site-specific substitution, the modularity of click chemistry, and the functional diversity of click labels, a multitude of modified prions can now be produced to ask targeted questions about the biochemical and biological bases of prion infectivity.