High yield purification and physico-chemical properties of full-length recombinant allelic variants of sheep prion protein linked to scrapie susceptibility

Sheep susceptibility to scrapie is governed by polymorphisms at two major s ites, codons 136 and 171, of the prp gene. To get more insight into the pri on protein (PrP) sequence-linked basis of differential scrapie susceptibili ty, a high yield one-step method for the purification (over 99% final purit y) of the full-length recombinant sheep PrP was developed, based on the aff inity of the conserved octapeptide repeats for transition-metal cations. Th ermal and chemical denaturation experiments and limited proteolysis studies were performed on the natural variants (A136R171, V136Q171 and A136Q171) a nd a recombinant PrP mutated at position 136 (V136R171). Results revealed t he influence of mutations in positions 136 and 171 on the folding thermodyn amic parameters and on the conformation of the C-terminal domain. Together, our results show that the VQ cellular protein linked to higher scrapie sus ceptibility is intrinsically more compact and/or stable than the resistance -linked AR counterpart. This might lead to a lower in vivo clearance rate o f VQ and a consequently higher probability of occurrence of pathological ev ents.