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Updated: May 6, 2026
Protein Misfolding Cyclic Amplification of Prions
Published on: November 7, 2012
Infrared microspectroscopy detects protein misfolding cyclic amplification (PMCA)-induced conformational alterations
Martin L Daus1, Katja Wagenführ, Achim Thomzig
1From FG 14-AG 5: Unconventional Pathogens and Their Inactivation, Applied Infection Control and Hospital Hygiene, Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany and.
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.
Insights
Prion protein (PrP) misfolding during in vitro amplification (PMCA) alters prion strain characteristics. These modifications were partially reversed when amplified prions were reintroduced into the original host species, impacting prion infectivity.
Area of Science:
- Neuroscience
- Biochemistry
- Molecular Biology
Background:
- Misfolded prion protein (PrP) conformation dictates prion seeding activity, infectiousness, and strain characteristics.
- Protein Misfolding Cyclic Amplification (PMCA) is a key in vitro method for monitoring prion seeding activity.
- The preservation of prion strain characteristics during PMCA is currently debated.
Purpose of the Study:
- To comparatively analyze the structural characteristics of parent and progeny prion seeds using infrared microspectroscopy.
- To investigate whether prion strain characteristics are maintained during PMCA amplification within the same host species.
Main Methods:
- Utilized an improved infrared microspectroscopy technique for sensitive structural analysis of prion seeds.
- Conducted comparative analysis of parent (263K scrapie) and progeny (PMCA-derived) hamster prion seeds.
- Assessed Proteinase K resistance, sedimentation velocities, and animal bioassays to evaluate progeny seed properties.
Main Results:
- Infrared microspectroscopy revealed conformational alterations in PMCA-derived progeny prion seeds.
- Progeny seeds exhibited altered Proteinase K resistance and higher sedimentation velocities compared to parent seeds.
- Animal bioassays showed that reinoculation of progeny seeds into hamsters resulted in PrP with mixed parental and progeny properties, indicating partial reversal of strain modification.
Conclusions:
- PMCA can induce conformational changes and alter strain characteristics of prions.
- Strain modifications induced by PMCA are not entirely stable and can be partially reversed upon host reinoculation.
- These findings have implications for the use of PMCA as an analytical tool and understanding prion strain dynamics.
