Journal
APPLIED SPECTROSCOPY
Volume 61, Issue 10, Pages 1025-1031Publisher
SOC APPLIED SPECTROSCOPY
DOI: 10.1366/000370207782217680
Keywords
Fourier transform infrared spectroscopy; FT-IR spectroscopy; prion protein; protein folding; curve fitting; band decomposition
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The conformational change of the recombinant, murine prion protein (PrP) from an alpha-helical to a beta-sheet enriched state was monitored by time-resolved Fourier transform infrared (FT-IR) spectroscopy. The alpha-to-beta transition is induced by reduction of the single disulfide bond in PrP. This transition is believed to generate the scrapie form PrPSc, the supposed infectious agent of transmissible spongiform encephalopathies. We followed the kinetics of this conformational change using a novel method for amide I band analysis of the infrared (IR) spectra. The amide I analysis provides the secondary structure. The amide I decomposition was calibrated with the three dimensional structure of cellular PrP solved by nuclear magnetic resonance (NMR). The novel secondary structure analysis provides a root mean squared deviation (RMSD) of only 3% as compared to the NMR structure. Reduction of alpha-helical PrP caused the transient accumulation of a partially unfolded intermediate, followed by formation of a state with higher beta-sheet than alpha-helical structure contents. The novel approach allows us to now determine the secondary structure of the P-sheet conformation. This was not determined by either NMR or Xray. The experiments were performed in a double-sealed security cuvette developed for IR analysis of potentially infectious PrP samples outside the biosafety laboratory.
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