title: RNA aptamers specifically interact with the prion protein PrP.
abstract: We have isolated RNA aptamers which are directed against the recombinant Syrian golden hamster prion protein rPrP23-231 (rPrPc) fused to glutathione S-transferase (GST). The aptamers did not recognize the fusion partner GST or the fusion protein GST::rPrP90-231 (rPrP27-30), which lacks 67 amino acids from the PrP N terminus. The aptamer-interacting region of PrPc was mapped to the N-terminal amino acids 23 to 52. Sequence analyses suggest that the RNA aptamers may fold into G-quartet-containing structural elements. Replacement of the G residues in the G quartet scaffold with uridine residues destroyed binding to PrP completely, strongly suggesting that the G quartet motif is essential for PrP recognition. Individual RNA aptamers interact specifically with prion protein in brain homogenates from wild-type mice (C57BL/6), hamsters (Syrian golden), and cattle as shown by supershifts obtained in the presence of anti-PrP antibodies. No interaction was observed with brain homogenates from PrP knockout mice (prn-p(0/0)). Specificity of the aptamer-PrP interaction was further confirmed by binding assays with antisense aptamer RNA or a mutant aptamer in which the guanosine residues in the G tetrad scaffold were replaced by uridine residues. The aptamers did not recognize PrP27-30 in brain homogenates from scrapie-infected mice. RNA aptamers may provide a first milestone in the development of a diagnostic assay for the detection of transmissible spongiform encephalopathies.
authors.affiliation: Laboratorium für Molekulare Biologie-Genzentrum-Institut für Biochemie der LMU München, Munich, Germany. Weiss@lmb.uni-muenchen.de
authors.forename: S
authors.initials: S
authors.lastname: Weiss
authors.forename: D
authors.initials: D
authors.lastname: Proske
authors.forename: M
authors.initials: M
authors.lastname: Neumann
authors.forename: M H
authors.initials: MH
authors.lastname: Groschup
authors.forename: H A
authors.initials: HA
authors.lastname: Kretzschmar
authors.forename: M
authors.initials: M
authors.lastname: Famulok
authors.forename: E L
authors.initials: EL
authors.lastname: Winnacker
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D006224:Cricetinae; D005982:Glutathione Transferase; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009843:Oligoribonucleotides / Q000737:chemistry*; D018622:PrP 27-30 Protein / Q000737:chemistry*; D016601:RNA-Binding Proteins / Q000737:chemistry*; D011993:Recombinant Fusion Proteins; D013045:Species Specificity; D013329:Structure-Activity Relationship
chemical_list: D009843:Oligoribonucleotides; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D018622:PrP 27-30 Protein; D005982:Glutathione Transferase