Record Details

Identification and characterization of a spontaneously aggregating amyloid-forming variant of human PrP((90-231)) through phage-display screening of variants randomized between residues 101 and 112.

DIR@IMTECH: CSIR-Institute of Microbial Technology

View Archive Info
 
 
Field Value
 
Title Identification and characterization of a spontaneously aggregating amyloid-forming variant of human PrP((90-231)) through phage-display screening of variants randomized between residues 101 and 112.
 
Creator Verma, Archana
Sharma, Swati
Ganguly, N K
Majumdar, Siddharta
Guptasarma, Purnananda
Luthra-Guptasarma, Manni
 
Subject QD Chemistry
 
Description The N-terminal 'unstructured' region of the human prion protein [PrP((90-231))] is believed to play a role in its aggregation because mutations in this region are associated with seeding-independent deposition disorders like Gerstmann-Straussler-Scheinker disease (GSS). One way of examining the effects of such mutations is to search combinatorially derived libraries for sequence variants showing a propensity to aggregate and/or the ability to interact with prion molecules folded into a beta-sheet-based conformation (i.e., beta-PrP or PrP(Sc)). We created a library of 1.8x10(7) variants randomized between positions 101 and 112, displayed it on filamentous bacteriophage, and 'spiked' it with a approximately 25% population of phages-bearing wild-type prion (wt-PrP). Screening was performed through four rounds of biopanning and amplification against immobilized beta-PrP, and yielded three beta-PrP-binding populations: wt-PrP (26% representation) and two non-wt-PrP variants ( approximately 10% and approximately 64% representation, respectively). The remarkable enrichment of one non-wt-PrP variant (MutPrP) incorporating residues KPSKPKTNMKHM in place of KGVLTWFSPLWQ, despite its initial representation at a 5 million-fold lower level than wt-PrP, caused us to produce it and discover: (i) that it readily aggregates into thioflavin-T-binding amyloids between pH 6.0 and 9.0, (ii) that it adopts a soluble beta-sheet based monomeric structure at pH 10.0, (iii) that it is less thermally stable and more compact than wt-PrP, and (iv) that it displays significantly greater resistance to proteolysis than wt-PrP. Our results suggest that sequence variations in the 101-112 region can indeed predispose the prion for aggregation.
 
Publisher Elsevier Science
 
Date 2008
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://crdd.osdd.net/open/1076/1/guptasarma2008.pdf
Verma, Archana and Sharma, Swati and Ganguly, N K and Majumdar, Siddharta and Guptasarma, Purnananda and Luthra-Guptasarma, Manni (2008) Identification and characterization of a spontaneously aggregating amyloid-forming variant of human PrP((90-231)) through phage-display screening of variants randomized between residues 101 and 112. The international journal of biochemistry & cell biology, 40 (4). pp. 663-76. ISSN 1357-2725
 
Relation http://www.sciencedirect.com/science/article/pii/S1357272507003214
http://crdd.osdd.net/open/1076/