Using isotopically-coded hydrogen peroxide as a surface modification reagent for the structural characterization of...

Area of Science:

• Proteomics
• Structural Biology
• Neuroscience

Background:

• Prion diseases result from the conversion of cellular prion protein (PrP(C)) into aggregated forms (PrP(ß) and PrP(Sc)).
• Understanding the structural changes during this conversion is crucial for developing therapeutic strategies.

Purpose of the Study:

• To develop and apply a novel method for differential characterization of PrP(C) and PrP(ß) forms.
• To identify specific regions of PrP(C) involved in the formation of PrP(ß) aggregates.

Main Methods:

• Utilized isotopically-coded hydrogen peroxide (H2(16)O2 and H2(18)O2) for surface modification of prion protein.
• Employed liquid chromatography-mass spectrometry (LC-MS) for quantitative analysis of oxidized residues.
• Compared oxidation patterns between native PrP(C) and aggregated PrP(ß) forms.

Main Results:

• Successfully differentiated PrP(C) and PrP(ß) based on differential oxidation of methionine and tryptophan residues.
• Identified specific amino acid residues with altered surface exposure during conversion to the β-oligomeric form.
• Detected conformational changes, including a potential flip of the β1-H1-β2 loop, during PrP aggregation.

Conclusions:

• Isotopically-coded hydrogen peroxide surface modification coupled with mass spectrometry provides a quantitative method to compare protein conformations.
• The study elucidates structural regions of the prion protein that undergo conformational transitions during the formation of disease-associated aggregates.
• Findings contribute to understanding the molecular mechanisms underlying prion diseases.