Systematic Approach to DNA Aptamer Design Using Amino Acid-Nucleic Acid Hybrids (ANHs) Targeting Thrombin

Chemical modifications of innate DNA/RNA aptamers facilitate the improvement of their function. Herein, we report our modular strategy to manipulate a thrombin-binding DNA aptamer (TBA) to improve its anticoagulation activity and binding affinity. A set of amino acid conjugates, termed amino acid-nucleic acid hybrids or ANHs, was synthesized and incorporated into a TBA loop sequences. We found that substitutions with hydrophobic amino acids in the loop region possessed significantly enhanced antithrombin activity, up to 3-fold higher than the native TBA. We investigated the correlations between thrombin-binding affinity and the features of our amino-acid conjugates using experimental techniques including circular dichroism spectroscopy, surface plasmon resonance assay, and molecular modeling. The present study demonstrates a systematic approach to aptamer design based on amino-acid characteristics, allowing the development of advanced aptamers.

Automated summary

Chemical modifications of innate DNA/RNA aptamers can improve their function. This study demonstrates a modular strategy to manipulate a thrombin-binding DNA aptamer by incorporating amino acid conjugates to enhance anticoagulation activity and binding affinity. The research findings reveal that substitutions with hydrophobic amino acids in the loop region can significantly increase antithrombin activity.