Conformational detection of prion protein with biarsenical labeling and FlAsH fluorescence

被引：31

作者：

Coleman, Bradley M.

Nisbet, Rebecca M. ^{[2
,3
]}

Han, Sen ^{[2
,3
]}

Cappai, Roberto ^{[2
,3
]}

Hatters, Danny M. ^{[2
,3
]}

Hill, Andrew F. ^{[1
,2
,3
]}

机构：

[1] Univ Melbourne, Mol Sci & Biotechnol Inst Bio21, Dept Biochem & Mol Biol, Parkville, Vic 3010, Australia

[2] Univ Melbourne, Dept Pathol, Parkville, Vic 3010, Australia

[3] Univ Melbourne, Mental Hlth Res Inst, Parkville, Vic 3010, Australia

来源：

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS | 2009年 / 380卷 / 03期

基金：

澳大利亚国家健康与医学研究理事会;

关键词：

Prion; Protein folding; PrP; Fluorescence; BLOOD-TRANSFUSION; PRP; TRANSMISSION; CONVERSION; BSE; REPLICATION; FEATURES;

D O I：

10.1016/j.bbrc.2009.01.120

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

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.

引用

页码：564 / 568

页数：5

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