Sedimentation velocity analysis of TMPyP4-induced dimer formation of human telomeric G-quadruplex

Abstract

5,10,15,20-Tetra-(N-methyl-4-pyridyl)porphyrin (TMPyP4), a ligand of G-quadruplex, has shown the ability to stabilize G-quadruplex structures and inhibit the activity of telomerase. Many methods have been used to study the interactions between TMPyP4 and G-quadruplex. However, many issues such as the binding number and the corresponding structural change are still controversial. Here, interactions between TMPyP4 and AGGG(TTAGGG)₃ (Tel22) have been studied by a combination of analytical ultracentrifugation sedimentation velocity (AUC-SV), polyacrylamide gel electrophoresis (PAGE), circular dichroism (CD) and UV-vis absorption spectroscopy. In the presence of NaCl, the binding number of TMPyP4 per Tel22, determined by AUC-SV, increases with the increasing concentration of TMPyP4 (C_TMPyP4). However, it decreases with the increasing concentration of NaCl (C_NaCl). Moreover, both AUC-SV and PAGE reveal that TMPyP4 can induce the formation of the dimeric G-quadruplex–TMPyP4 complex through a medium affinity binding mode. High affinity binding modes, which may include the inner intercalation mode and end stacking, have no contribution to the formation of dimers. The weak electrostatic binding of TMPyP4 to Tel22 has a negative effect on the formation of dimers, presumably due to the instability of G-quadruplex induced by this binding mode.