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Default Protocol for aerosol-free recombinant production and NMR analysis of prion proteins

Protocol for aerosol-free recombinant production and NMR analysis of prion proteins

Abstract

The central hallmark of prion diseases is the misfolding of cellular prion protein (PrPC) into a disease-associated aggregated isoform known as scrapie prion protein (PrPSc). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded and aggregated states. Recent studies reporting on de novo generation of prions from recombinant PrP and infection of animals using prion aerosols suggest that adjustment of current biosafety measures may be necessary, particularly given the relatively high protein concentrations required for NMR applications that favor aggregation. We here present a protocol for the production of recombinant PrP under biosafety level 2 conditions that avoids entirely exposure of the experimenter to aerosols that might contain harmful PrP aggregates. In addition, we introduce an NMR sample tube setup that allows for safe handling of PrP samples at the spectrometer that usually is not part of a dedicated biosafety level 2 laboratory.



Source: Journal of Biomolecular NMR
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