Development of an Efficient G-Quadruplex-Stabilised Thrombin-Binding Aptamer Containing a Three-Carbon Spacer Molecule

Chembiochem. 2017 Apr 18;18(8):755-763. doi: 10.1002/cbic.201600654. Epub 2017 Mar 15.

Abstract

The thrombin-binding aptamer (TBA), which shows anticoagulant properties, is one of the most studied G-quadruplex-forming aptamers. In this study, we investigated the impact of different chemical modifications such as a three-carbon spacer (spacer-C3 ), unlocked nucleic acid (UNA) and 3'-amino-modified UNA (amino-UNA) on the structural dynamics and stability of TBA. All three modifications were incorporated at three different loop positions (T3, T7, T12) of the TBA G-quadruplex structure to result in a series of TBA variants and their stability was studied by thermal denaturation; folding was studied by circular dichroism spectroscopy and thrombin clotting time. The results showed that spacer-C3 introduction at the T7 loop position (TBA-SP7) significantly improved stability and thrombin clotting time while maintaining a similar binding affinity as TBA to thrombin. Detailed molecular modelling experiments provided novel insights into the experimental observations, further supporting the efficacy of TBA-SP7. The results of this study could provide valuable information for future designs of TBA analogues with superior thrombin inhibition properties.

Keywords: G-quadruplex; aptamers; modified DNA; thrombin; unlocked nucleic acid.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aptamers, Nucleotide / chemical synthesis*
  • Aptamers, Nucleotide / chemistry*
  • Blood Coagulation
  • G-Quadruplexes*
  • Models, Chemical
  • Molecular Dynamics Simulation
  • Molecular Structure
  • Stochastic Processes

Substances

  • Aptamers, Nucleotide
  • TBA-SP7 aptamer
  • thrombin aptamer