Co-crystal structure of the iMango-III fluorescent RNA aptamer using an X-ray free-electron laser

Acta Crystallogr F Struct Biol Commun. 2019 Aug 1;75(Pt 8):547-551. doi: 10.1107/S2053230X19010136. Epub 2019 Aug 2.

Abstract

Turn-on aptamers are in vitro-selected RNAs that bind to conditionally fluorescent small molecules and enhance their fluorescence. Upon binding TO1-biotin, the iMango-III aptamer achieves the largest fluorescence enhancement reported for turn-on aptamers (over 5000-fold). This aptamer was generated by structure-guided engineering and functional reselection of the parental aptamer Mango-III. Structures of both Mango-III and iMango-III have previously been determined by conventional cryocrystallography using synchrotron X-radiation. Using an X-ray free-electron laser (XFEL), the room-temperature iMango-III-TO1-biotin co-crystal structure has now been determined at 3.0 Å resolution. This structural model, which was refined against a data set of ∼1300 diffraction images (each from a single crystal), is largely consistent with the structures determined from single-crystal data sets collected at 100 K. This constitutes a technical benchmark on the way to XFEL pump-probe experiments on fluorescent RNA-small molecule complexes.

Keywords: RNA structure; X-ray free-electron lasers; XFELS; fluorescence; fluorescence turn-on aptamers; iMango-III; room temperature.

MeSH terms

  • Aptamers, Nucleotide / chemistry*
  • Aptamers, Nucleotide / genetics
  • Aptamers, Nucleotide / metabolism*
  • Crystallography, X-Ray
  • Electrons
  • Fluorescent Dyes / chemistry*
  • Lasers
  • Nucleic Acid Conformation
  • RNA / chemistry*
  • RNA / genetics
  • RNA / metabolism*
  • X-Rays

Substances

  • Aptamers, Nucleotide
  • Fluorescent Dyes
  • RNA

Grant support

This work was funded by National Heart, Lung, and Blood Institute grant . National Cancer Institute grant .