Duplex/quadruplex aptamers that effectively modulate two key coagulation factors: a spectroscopic and calorimetric study

Duplex/quadruplex aptamers that effectively modulate two key coagu lation factors: a spectroscopic and calorimetric study Aptamers are short DNA or RNA oligonucleotides characterized by the ability to bind a molecular target with strong affinity and high selectivity [1]. Due to their three-dimensional structure, aptamers can inhibit or control activity of their targets through recognition of specific binding sites [2]. An extensively analysed class of these aptamers is that consisting of oligonucleotides adopting a G-quadruplex structure. Recently, researchers have focused on the study of aptamers with bimodular architecture, characterized by the presence of two structural motifs within the same chain, a duplex and a quadruplex one. Some of these duplex/quadruplex aptamers are able to modulate the coagulation process by binding coagulation factors, such as thrombin or prothrombin [3-8]. Human α-thrombin is a trypsin-like serine protease that has the unique ability to convert soluble fibrinogen in insoluble fibrin clot [9]. In addition to the active site, this enzyme owns two electropositive regions, exosite I and II, located at opposite sides of its globular shape and playing an essential role in the recognition of substrate, cofactors, and aptamers. Thrombin precursor is prothrombin, a multi-domain protein, in which exosite I is exposed whereas exosite II is well hidden inside the protein structure [10]. Here we present a spectroscopic and calorimetric characterization of structure and stability of different duplex/quadruplex anti-thrombin aptamers, including NU172, the only anticoagulant aptamer currently in clinical trials, in the free and liganded state [6]. The role of the duplex domain in the stabilization of bimodular aptamers and the thermodynamic characterization of the binding of these oligonucleotides at exosite I of thrombin or prothrombin, will be discussed at the Meeting. Bibliografia 1. Rozenblu m, G.T.; Lopez, V.G.; Vitullo, A.D.; Radrizzani, M.;, Expert Opin Drug Discov, 2016. 11, 127-135. 2. Keefe, A.D.; Pai, S.; Ellington, A.;, Nat Rev Drug Discov, 2010. 9, 537-550. 3. Russo Krauss, I.; Merlino, A.; Randazzo, A.; Novellino, E.; Mazzarella, L.; S ica, F.;, Nucleic Acids Res, 2012. 40, 8119-8128. 4. Russo Krauss, I.; Pica, A.; Merlino, A.; Mazzarella, L.; Sica, F.;, Acta Crystallogr D, 2013. 69, 2403-2411. 5. Russo Krauss, I.; Spiridonova, V.; Pica, A.; Napolitano, V.; Sica, F.;, Nucleic Acids Res, 2016. 44, 983-99 1. 6. Russo Krauss, I.; Napolitano, V.; Petraccone, L.; Troisi, R.; Spiridonova, V.; Mattia, C.A.; Sica, F.;, Int J Biol Macromol, 2018. 107, 1697- 1705. 7. Tro isi, R.; Napolitano, V.; Spiridonova, V.; Russo Krauss, I.; Sica F.;, Nucleic Acids Res, 2018. 46, 12177-12185. 8. Zavyalova, E.G.; Legatova, V.A.; Alieva, R.S.; Zalevsky, A.O.; Tashlitsky, V.N.; Arutyunyan, A.M.; Kopylov A.M.;, Biomolecules, 2019. 9, 41. 9. D i Cera, E.;, J Thromb Haemost, 2007. 5 Suppl 1, 196-202. 10. Pozzi, N.; Chen, Z.; Gohara, D.W.; Niu, W.; Heyduk, T.; Di Cera, E.;, J Biol Chem, 2013. 288, 22734-22744.