title: Integrity of helix 2-helix 3 domain of the PrP protein is not mandatory for prion replication.
abstract: The process of prion conversion is not yet well understood at the molecular level. The regions critical for the conformational change of PrP remain mostly debated and the extent of sequence change acceptable for prion conversion is poorly documented. To achieve progress on these issues, we applied a reverse genetic approach using the Rov cell system. This allowed us to test the susceptibility of a number of insertion mutants to conversion into prion in the absence of wild-type PrP molecules. We were able to propagate several prions with 8 to 16 extra amino acids, including a polyglycine stretch and His or FLAG tags, inserted in the middle of the protease-resistant fragment. These results demonstrate the possibility to increase the length of the loop between helices H2 and H3 up to 4-fold, without preventing prion replication. They also indicate that this loop probably remains unstructured in PrP(Sc). We also showed that bona fide prions can be produced following insertion of octapeptides in the two C-terminal turns of H2. These insertions do not interfere with the overall fold of the H2-H3 domain indicating that the highly conserved sequence of the terminal part of H2 is not critical for the conversion. Altogether these data showed that the amplitude of modifications acceptable for prion conversion in the core of the globular domain of PrP is much greater than one might have assumed. These observations should help to refine structural models of PrP(Sc) and elucidate the conformational changes underlying prions generation.
authors.affiliation: INRA, UR892 Virologie Immunologie Moléculaires, F-78350 Jouy-en-Josas, France.
authors.forename: Khalid
authors.initials: K
authors.lastname: Salamat
authors.forename: Mohammed
authors.initials: M
authors.lastname: Moudjou
authors.forename: Jérôme
authors.initials: J
authors.lastname: Chapuis
authors.forename: Laetitia
authors.initials: L
authors.lastname: Herzog
authors.forename: Emilie
authors.initials: E
authors.lastname: Jaumain
authors.forename: Vincent
authors.initials: V
authors.lastname: Béringue
authors.forename: Human
authors.initials: H
authors.lastname: Rezaei
authors.forename: Annalisa
authors.initials: A
authors.lastname: Pastore
authors.forename: Hubert
authors.initials: H
authors.lastname: Laude
authors.forename: Michel
authors.initials: M
authors.lastname: Dron
mesh_terms: D000818:Animals; D002460:Cell Line; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D016254:Mutagenesis, Insertional; D011328:Prions / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D011328:Prions