Characterization of 2′-fluoro-RNA aptamers that bind preferentially to disease-associated conformations of prion protein and inhibit conversion

Rhie, Alexandre and Kirby, Louise and Sayer, Natalie and Wellesley, Rosanna and Disterer, Petra and Sylvester, Ian and Gill, Andrew C. and Hope, James and James, William and Tahiri-Alaoui, Abdessamad (2003) Characterization of 2′-fluoro-RNA aptamers that bind preferentially to disease-associated conformations of prion protein and inhibit conversion. Journal of Biological Chemistry, 278 (41). pp. 39697-39705. ISSN 0021-9258

Full content URL: http://doi.org/10.1074/jbc.M305297200

Documents
rhie39697.pdf
[img]
[Download]
[img]
Preview
PDF
rhie39697.pdf - Whole Document

390kB
Item Type:Article
Item Status:Live Archive

Abstract

We have isolated artificial ligands or aptamers for infectious prions in order to investigate conformational aspects of prion pathogenesis. The aptamers are 2'-fluoro-modified RNA produced by in vitro selection from a large, randomized library. One of these ligands (aptamer SAF-93) had more than 10-fold higher affinity for PrPSc than for recombinant PrPC and inhibited the accumulation of PrPres in near physiological cell-free conversion assay. To understand the molecular basis of these properties and to distinguish specific from nonspecific aptamer-PrP interactions, we studied deletion mutants of bovine PrP in denatured, alpha-helix-rich and beta-sheet-rich forms. We provide evidence that, like scrapie-associated fibrils (SAF), the beta-oligomer of PrP bound to SAF-93 with at least 10-fold higher affinity than did the alpha-form. This differential affinity could be explained by the existence of two binding sites within the PrP molecule. Site 1 lies within residues 23-110 in the unstructured N terminus and is a nonspecific RNA binding site found in all forms of PrP. The region between residue 90 and 110 forms a hinge region that is occluded in the alpha-rich form of PrP but becomes exposed in the denatured form of PrP. Site 2 lies in the region C-terminal of residue 110. This site is beta-sheet conformation-specific and is not recognized by control RNAs. Taken together, these data provide for the first time a specific ligand for a disease conformation-associated site in a region of PrP critical for conformational conversion. This aptamer could provide tools for the further analysis of the processes of PrP misfolding during prion disease and leads for the development of diagnostic and therapeutic approaches to TSEs.

Keywords:creutzfeldt jakob disease, NMR structure, transmissible spongiform encephalopathies, prion, RNA aptamer
Subjects:C Biological Sciences > C760 Biomolecular Science
Divisions:College of Science
ID Code:29573
Deposited On:16 Mar 2018 16:55

Repository Staff Only: item control page