Journal
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Volume 380, Issue 3, Pages 564-568Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.bbrc.2009.01.120
Keywords
Prion; Protein folding; PrP; Fluorescence
Categories
Funding
- NHMRC
- University of Melbourne
Ask authors/readers for more resources
Prion diseases are associated with the misfolding of the host-encoded cellular prion protein (PrPC) into a disease associated form (PrPSc). Recombinant PrP can be refolded into either an alpha-helical rich conformation (alpha-PrP) resembling PrPC or a P-sheet rich. protease resistant form similar to PrPSc. Here, we generated tetracysteine tagged recombinant PrP, folded this into alpha- or beta-PrP and determined the levels of FlAsH fluorescence. Insertion of the tetracysteine tag at three different sites within the 91-111 epitope readily distinguished beta-PrP from alpha-PrP upon FlAsH labeling. Labelling of tetracysteine tagged PrP in the alpha-helical form showed minimal fluorescence, whereas labeling of tagged PrP in the beta-sheet form showed high fluorescence indicating that this region is exposed upon conversion. This highlights a region of PrP that can be implicated in the development of diagnostics and is a novel, protease free mechanism for distinguishing PrPSc from PrPC. This technique may also be applied to any protein that undergoes conformational change and/or misfolding Such as those involved in other neurodegenerative disorders including Alzheimer's, Huntington's and Parkinson's diseases. (C) 2009 Elsevier Inc. All rights reserved.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available