Direct observation of prion-like propagation of protein misfolding templated by pathogenic mutants

Many neurodegenerative diseases feature misfolded proteins that propagate via templated conversion of natively folded molecules. However, crucial questions about how such prion-like conversion occurs and what drives it remain unsolved, partly because technical challenges have prevented direct observation of conversion for any protein. We observed prion-like conversion in single molecules of superoxide dismutase-1 (SOD1), whose misfolding is linked to amyotrophic lateral sclerosis. Tethering pathogenic misfolded SOD1 mutants to wild-type molecules held in optical tweezers, we found that the mutants vastly increased misfolding of the wild-type molecule, inducing multiple misfolded isoforms. Crucially, the pattern of misfolding was the same in the mutant and converted wild-type domains and varied when the misfolded mutant was changed, reflecting the templating effect expected for prion-like conversion. Ensemble measurements showed decreased enzymatic activity in tethered heterodimers as conversion progressed, mirroring the single-molecule results. Antibodies sensitive to disease-specific epitopes bound to the converted protein, implying that conversion produced disease-relevant misfolded conformers. Protein misfolding can spread from one molecule to another in infectious prion diseases. The propagation of protein misfolding has been directly observed in single protein molecules. These results showed that pathogenic mutants of the protein superoxide dismutase-1 (SOD1), which causes familial amyotrophic lateral sclerosis, imprint their misfolding onto native wild-type molecules.