title: Infrared microspectroscopy detects protein misfolding cyclic amplification (PMCA)-induced conformational alterations in hamster scrapie progeny seeds.
abstract: The self-replicative conformation of misfolded prion proteins (PrP) is considered a major determinant for the seeding activity, infectiousness, and strain characteristics of prions in different host species. Prion-associated seeding activity, which converts cellular prion protein (PrP(C)) into Proteinase K-resistant, infectious PrP particles (PrP(TSE)), can be monitored in vitro by protein misfolding cyclic amplification (PMCA). Thus, PMCA has been established as a valuable analytical tool in prion research. Currently, however, it is under discussion whether prion strain characteristics are preserved during PMCA when parent seeds are amplified in PrP(C) substrate from the identical host species. Here, we report on the comparative structural analysis of parent and progeny (PMCA-derived) PrP seeds by an improved approach of sensitive infrared microspectroscopy. Infrared microspectroscopy revealed that PMCA of native hamster 263K scrapie seeds in hamster PrP(C) substrate caused conformational alterations in progeny seeds that were accompanied by an altered resistance to Proteinase K, higher sedimentation velocities in gradient ultracentrifugations, and a longer incubation time in animal bioassays. When these progeny seeds were propagated in hamsters, misfolded PrP from brain extracts of these animals showed mixed spectroscopic and biochemical properties from both parental and progeny seeds. Thus, strain modifications of 263K prions induced by PMCA seem to have been partially reversed when PMCA products were reinoculated into the original host species.
authors.affiliation: From FG 14-AG 5: Unconventional Pathogens and Their Inactivation, Applied Infection Control and Hospital Hygiene, Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany and.
authors.forename: Martin L
authors.initials: ML
authors.lastname: Daus
authors.forename: Katja
authors.initials: K
authors.lastname: Wagenführ
authors.forename: Achim
authors.initials: A
authors.lastname: Thomzig
authors.forename: Susann
authors.initials: S
authors.lastname: Boerner
authors.forename: Peter
authors.initials: P
authors.lastname: Hermann
authors.forename: Antje
authors.initials: A
authors.lastname: Hermelink
authors.forename: Michael
authors.initials: M
authors.lastname: Beekes
authors.forename: Peter
authors.initials: P
authors.lastname: Lasch
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D006224:Cricetinae; D019286:Endopeptidase K; D008647:Mesocricetus; D018625:Microscopy, Atomic Force; D018622:PrP 27-30 Protein / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D018620:PrPSc Proteins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D011487:Protein Conformation; D017510:Protein Folding; D055550:Protein Stability; D012608:Scrapie / Q000378:metabolism / Q000635:transmission; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods
chemical_list: D018620:PrPSc Proteins; D018622:PrP 27-30 Protein; D019286:Endopeptidase K
keywords: Amyloid; Biophysics; Infrared Microspectroscopy (IR-MSP); Neurodegenerative Diseases; Prion Strains; Prion Structure; Prions; Protein Misfolding; Protein Misfolding Cyclic Amplification (PMCA); Seeding Activity