Anti-bovine prion protein RNA aptamer containing tandem GGA repeat interacts both with recombinant bovine prion protein and its beta isoform with high affinity

作者：Murakami Kazuyoshi; Nishikawa Fumiko; Noda Ken; Yokoyama Takashi; Nishikawa Satoshi^*

来源：Prion, 2008, 2(2): 73-80.

• RNA aptamer
• prion protein
• SELEX
• GGA repeat
• G-quadruplex

Iorder to obtain RNA aptamers against bovine prion protein (bPrP), we carried out in vitro selection from RNA pools containing a 55-nucleotide randomized region to target recombinant bPrP. Most of obtained aptamers contained conserved GGA tandem repeats (GGA) 4 and aptamer #1 (apt #1) showed a high affinity for both bPrP and its beta isoform (bPrP-beta). The sequence of apt #1 suggested that it would have a G-quadruplex structure, which was confirmed using CD spectra in titration with KCl. A mutagenic study of this conserved region, and competitive assays, showed that the conserved (GGA) 4 sequence is important for specific binding to bPrP and bPrP-beta. Following 5'-biotinylation, aptamer #1 specifically detected PrPc in bovine brain homogenate in a Northwestern blotting assay. Protein deletion mutant analysis showed that the bPrP aptamer binds within 25-131 of the bPrP sequence. Interestingly, the minimized aptamer #1 (17 nt) showed greater binding to bPrP and bPrP-beta as compared to apt #1. This minimized form of aptamer #1 may therefore be useful in the detection of bPrP, diagnosis of prion disease, enrichment of bPr and ultimately in gaining a better understanding of prion diseases.

• 出版日期2008-6