Structural basis for activation of fluorogenic dyes by an RNA aptamer lacking a G-quadruplex motif

Sandip A Shelke; Yaming Shao; Artur Laski; Deepak Koirala; Benjamin P Weissman; James R Fuller; Xiaohong Tan; Tudor P Constantin; Alan S Waggoner; Marcel P Bruchez; Bruce A Armitage; Joseph A Piccirilli

doi:10.1038/s41467-018-06942-3

Structural basis for activation of fluorogenic dyes by an RNA aptamer lacking a G-quadruplex motif

Nat Commun. 2018 Oct 31;9(1):4542. doi: 10.1038/s41467-018-06942-3.

Authors

Sandip A Shelke¹, Yaming Shao¹, Artur Laski¹, Deepak Koirala¹, Benjamin P Weissman², James R Fuller¹, Xiaohong Tan^{3

4}, Tudor P Constantin⁴, Alan S Waggoner^{5

6}, Marcel P Bruchez^{4

5

6}, Bruce A Armitage^{3

4}, Joseph A Piccirilli^{7

8}

Affiliations

¹ Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL, 60637, USA.
² Department of Chemistry, The University of Chicago, Chicago, IL, 60637, USA.
³ Center for Nucleic Acids Science and Technology, Carnegie Mellon University, Pittsburgh, PA, 15213, USA.
⁴ Department of Chemistry, Carnegie Mellon University, Pittsburgh, PA, 15213, USA.
⁵ Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA, 15213, USA.
⁶ Molecular Biosensor and Imaging Center, Carnegie Mellon University, Pittsburgh, PA, 15213, USA.
⁷ Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL, 60637, USA. jpicciri@uchicago.edu.
⁸ Department of Chemistry, The University of Chicago, Chicago, IL, 60637, USA. jpicciri@uchicago.edu.

Abstract

The DIR2s RNA aptamer, a second-generation, in-vitro selected binder to dimethylindole red (DIR), activates the fluorescence of cyanine dyes, DIR and oxazole thiazole blue (OTB), allowing detection of two well-resolved emission colors. Using Fab BL3-6 and its cognate hairpin as a crystallization module, we solved the crystal structures of both the apo and OTB-SO₃ bound forms of DIR2s at 2.0 Å and 1.8 Å resolution, respectively. DIR2s adopts a compact, tuning fork-like architecture comprised of a helix and two short stem-loops oriented in parallel to create the ligand binding site through tertiary interactions. The OTB-SO₃ fluorophore binds in a planar conformation to a claw-like structure formed by a purine base-triple, which provides a stacking platform for OTB-SO_3, and an unpaired nucleotide, which partially caps the binding site from the top. The absence of a G-quartet or base tetrad makes the DIR2s aptamer unique among fluorogenic RNAs with known 3D structure.

Publication types

Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Aptamers, Nucleotide / chemistry*
Binding Sites
Crystallography, X-Ray
Fluorescent Dyes / chemistry*
G-Quadruplexes*
Immunoglobulin Fab Fragments / chemistry
Nucleotide Motifs*

Substances

Aptamers, Nucleotide
Fluorescent Dyes
Immunoglobulin Fab Fragments

Grants and funding

R01 GM102489/GM/NIGMS NIH HHS/United States