2′-fluoro-modified pyrimidines enhance affinity of RNA oligonucleotides to HIV-1 reverse transcriptase
- 1Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, USA
- 2Bond Life Sciences Center, University of Missouri, Columbia, Missouri 65211, USA
- 3Department of Veterinary Pathobiology, University of Missouri, Columbia, Missouri 65211, USA
- 4Department of Molecular Microbiology and Immunology, University of Missouri, Columbia, Missouri 65211, USA
- 5Department of Biological Engineering, University of Missouri, Columbia, Missouri 65211, USA
- Corresponding author: burkedh{at}missouri.edu
Abstract
Nucleic acid aptamers can be chemically modified to enhance function, but modifying previously selected aptamers can have nontrivial structural and functional consequences. We present a reselection strategy to evaluate the impact of several modifications on preexisting aptamer pools. RNA aptamer libraries with affinity to HIV-1 reverse transcriptase (RT) were retranscribed with 2′-F, 2′-OMe, or 2′-NH2 pyrimidines and subjected to three additional selection cycles. RT inhibition was observed for representative aptamers from several structural families identified by high-throughput sequencing when transcribed with their corresponding modifications. Thus, reselection identified specialized subsets of aptamers that tolerated chemical modifications from unmodified preenriched libraries. Inhibition was the strongest with the 2′-F-pyrimidine (2′-FY) RNAs, as compared to inhibition by the 2′-OMeY and 2′-NH2Y RNAs. Unexpectedly, a diverse panel of retroviral RTs were strongly inhibited by all 2′-FY-modified transcripts, including sequences that do not inhibit those RTs as unmodified RNA. The magnitude of promiscuous RT inhibition was proportional to mole fraction 2′-FY in the transcript. RT binding affinity by 2′-FY transcripts was more sensitive to salt concentration than binding by unmodified transcripts, indicating that interaction with retroviral RTs is more ionic in character for 2′-FY RNA than for unmodified 2′-OH RNA. These surprising features of 2′-FY-modified RNA may have general implications for applied aptamer technologies.
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Footnotes
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Article is online at http://www.rnajournal.org/cgi/doi/10.1261/rna.077008.120.
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Freely available online through the RNA Open Access option.
- Received June 25, 2020.
- Accepted July 24, 2020.
This article, published in RNA, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.