title: Confomational analysis of soluble oligomers of GFP tagged prion protein by fluorescence fluctuation spectroscopy.
abstract: The conversion of prion protein (PrP) from the monomeric cellular isoform to the oligomeric pathological isoform is a crucial event in the pathogenesis of prion diseases. To investigate oligomer formation of PrP, enhanced green fluorescent protein (EGFP)-tagged PrP (EGFP-PrP) without the glycosylphosphatidylinositol (GPI) anchor was prepared and the oligomerization of EGFP-PrP induced by sodium dodecyl sulphate (SDS) was monitored by fluorescence correlation spectroscopy (FCS). The FCS analysis indicated that soluble oligomers were formed at 0.011% SDS. Furthermore, the combination of fluorescence cross-correlation spectroscopy (FCCS) and a panel of anti-PrP monoclonal antibodies (mAbs) revealed the conformational changes in PrP. Our studies provide a method to analyze conformational changes of proteins in solution.
authors.affiliation: Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido University, Sapporo 001-0021, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Sakata
authors.forename: Motohiro
authors.initials: M
authors.lastname: Horiuchi
authors.forename: Izumi
authors.initials: I
authors.lastname: Takahashi
authors.forename: Masataka
authors.initials: M
authors.lastname: Kinjo
mesh_terms: D049452:Green Fluorescent Proteins / Q000737:chemistry*; D011328:Prions / Q000737:chemistry* / Q000648:ultrastructure*; D011487:Protein Conformation; D012995:Solubility; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D011328:Prions; D049452:Green Fluorescent Proteins