Mapping the affinity landscape of Thrombin-binding aptamers on 2΄F-ANA/DNA chimeric G-Quadruplex microarrays

Nucleic Acids Res. 2017 Feb 28;45(4):1619-1632. doi: 10.1093/nar/gkw1357.


In situ fabricated nucleic acids microarrays are versatile and very high-throughput platforms for aptamer optimization and discovery, but the chemical space that can be probed against a given target has largely been confined to DNA, while RNA and non-natural nucleic acid microarrays are still an essentially uncharted territory. 2΄-Fluoroarabinonucleic acid (2΄F-ANA) is a prime candidate for such use in microarrays. Indeed, 2΄F-ANA chemistry is readily amenable to photolithographic microarray synthesis and its potential in high affinity aptamers has been recently discovered. We thus synthesized the first microarrays containing 2΄F-ANA and 2΄F-ANA/DNA chimeric sequences to fully map the binding affinity landscape of the TBA1 thrombin-binding G-quadruplex aptamer containing all 32 768 possible DNA-to-2΄F-ANA mutations. The resulting microarray was screened against thrombin to identify a series of promising 2΄F-ANA-modified aptamer candidates with Kds significantly lower than that of the unmodified control and which were found to adopt highly stable, antiparallel-folded G-quadruplex structures. The solution structure of the TBA1 aptamer modified with 2΄F-ANA at position T3 shows that fluorine substitution preorganizes the dinucleotide loop into the proper conformation for interaction with thrombin. Overall, our work strengthens the potential of 2΄F-ANA in aptamer research and further expands non-genomic applications of nucleic acids microarrays.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aptamers, Nucleotide / chemistry*
  • Arabinonucleotides / chemistry*
  • Base Sequence
  • DNA / chemistry*
  • G-Quadruplexes*
  • Magnetic Resonance Spectroscopy
  • Models, Molecular
  • Oligonucleotide Array Sequence Analysis
  • Organophosphorus Compounds / chemistry


  • 2'-deoxy-2'-fluoro-beta-D-arabinonucleic acid
  • Aptamers, Nucleotide
  • Arabinonucleotides
  • Organophosphorus Compounds
  • phosphoramidite
  • thrombin aptamer
  • DNA