Background: Stabilization of G-quadruplex helices by small ligands has attracted growing attention because they
inhibit the activity of the enzyme telomerase, which is overexpressed in> 80% cancer cells. TMPyP4, one of the
most studied G-quadruplex ligands, is used as a model to show that the ligands can exhibit different binding
features with different conformations of a human telomeric specific sequence.
Methods: UV–Vis, FRET melting Assay, Isothermal Titration Calorimetry, Time-resolved Fluorescence lifetime,
T-Jump and Molecular Dynamics.
Results: TMPyP4 yields two different complexes with two Tel22 telomeric conformations in the presence of Na+
or K+. T-Jump kinetic experiments show that the rates of formation and dissociation of these complexes in the
ms time scale differ by one order of magnitude. MD simulations reveal that, in K+ buffer, “hybrid 1” conformation
yields kinetic constants on interaction with TMPyP4 one order lower than “hybrid 2”. The binding
involves π–π stacking with external loop bases.
Conclusions: For the first time we show that for a particular buffer TMPyP4 interacts in a kinetically different
way with the two Tel22 conformations even if the complexes formed are thermodynamically indistinguishable.
General significance: G-quadruplexes, endowed with technological applications and potential impact on regulation
mechanisms, define a new research field. The possibility of building different conformations from same
sequence is a complex issue that confers G-quadruplexes very interesting features. The obtaining of reliable
kinetic data constitutes an efficient tool to determine reaction mechanisms between conformations and small
molecules.
