G-Quadruplex Aptamer-Ligand Characterization

0563712 - BFÚ 2023 RIV CH eng J - Článek v odborném periodiku
Moreira, D. - Leitao, D. - Lopes-Nunes, J. - Santos, T. C. B. - Figueiredo, J. - Miranda, A.I. - Alexandre, D. - Tomaz, C. - Mergny, Jean-Louis - Cruz, C.
G-Quadruplex Aptamer-Ligand Characterization.
Molecules. Roč. 27, č. 20 (2022), č. článku 6781. E-ISSN 1420-3049
Grant CEP: GA MŠMT EF15_003/0000477
Institucionální podpora: RVO:68081707
Klíčová slova: G-quadruplex aptamer * ligands * aptamer-ligand interactions * biophysical techniques
Obor OECD: Biochemistry and molecular biology
Impakt faktor: 4.6, rok: 2022
Způsob publikování: Open access
https://www.mdpi.com/1420-3049/27/20/6781

In this work we explore the structure of a G-rich DNA aptamer termed AT11-L2 (TGGTGGTGGTTGTTGTTGGTGGTGGTGGT, derivative of AT11) by evaluating the formation and stability of G-quadruplex (G4) conformation under different experimental conditions such as KCl concentration, temperature, and upon binding with a variety of G4 ligands (360A, BRACO-19, PDS, PhenDC3, TMPyP4). We also determined whether nucleolin (NCL) can be a target of AT11-L2 G4. Firstly, we assessed by circular dichroism, UV and NMR spectroscopies the formation of G4 by AT11-L2. We observed that, for KCl concentrations of 65 mM or less, AT11-L2 adopts hybrid or multiple topologies. In contrast, a parallel topology predominates for buffer containing 100 mM of KCl. The T-m of AT11-L2 in 100 mM of KCl is 38.9 degrees C, proving the weak stability of this sequence. We also found that upon titration with two molar equivalents of 360A, BRACO-19 and PhenDC3, the G4 is strongly stabilized and its topology is maintained, while the addition of 3.5 molar equivalents of TMPyP4 promotes the disruption of G4. The K-D values between AT11-L2 G4, ligands and NCL were obtained by fluorescence titrations and are in the range of mu M for ligand complexes and nM when adding NCL. In silico studies suggest that four ligands bind to the AT11-L2 G4 structure by stacking interactions, while the RBD1,2 domains of NCL interact preferentially with the thymines of AT11-L2 G4. Finally, AT11-L2 G4 co-localized with NCL in NCL-positive tongue squamous cell carcinoma cell line.
Trvalý link: https://hdl.handle.net/11104/0340583