title: High yield purification and physico-chemical properties of full-length recombinant allelic variants of sheep prion protein linked to scrapie susceptibility.
abstract: Sheep susceptibility to scrapie is governed by polymorphisms at two major sites, codons 136 and 171, of the prp gene. To get more insight into the prion protein (PrP) sequence-linked basis of differential scrapie susceptibility, a high yield one-step method for the purification (over 99% final purity) of the full-length recombinant sheep PrP was developed, based on the affinity of the conserved octapeptide repeats for transition-metal cations. Thermal and chemical denaturation experiments and limited proteolysis studies were performed on the natural variants (A136R171, V136Q171 and A136Q171) and a recombinant PrP mutated at position 136 (V136R171). Results revealed the influence of mutations in positions 136 and 171 on the folding thermodynamic parameters and on the conformation of the C-terminal domain. Together, our results show that the VQ cellular protein linked to higher scrapie susceptibility is intrinsically more compact and/or stable than the resistance-linked AR counterpart. This might lead to a lower in vivo clearance rate of VQ and a consequently higher probability of occurrence of pathological events.
authors.affiliation: Institut National de la Recherche Agronomique (INRA) Unité 806/EA2703, Muséum National d'Histoire Naturelle, Institut de Biologie Physico-Chimique, Paris, France.
authors.forename: H
authors.initials: H
authors.lastname: Rezaei
authors.forename: D
authors.initials: D
authors.lastname: Marc
authors.forename: Y
authors.initials: Y
authors.lastname: Choiset
authors.forename: M
authors.initials: M
authors.lastname: Takahashi
authors.forename: G
authors.initials: G
authors.lastname: Hui Bon Hoa
authors.forename: T
authors.initials: T
authors.lastname: Haertlé
authors.forename: J
authors.initials: J
authors.lastname: Grosclaude
authors.forename: P
authors.initials: P
authors.lastname: Debey
mesh_terms: D000483:Alleles; D000818:Animals; D015153:Blotting, Western; D002942:Circular Dichroism; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D020022:Genetic Predisposition to Disease; D006863:Hydrogen-Ion Concentration; D008957:Models, Genetic; D016296:Mutagenesis; D011110:Polymorphism, Genetic; D011328:Prions / Q000235:genetics* / Q000302:isolation & purification*; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D012608:Scrapie / Q000235:genetics / Q000378:metabolism*; D012756:Sheep; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
chemical_list: D018076:DNA, Complementary; D011328:Prions; D011994:Recombinant Proteins; D019286:Endopeptidase K