Conformational dynamics of DNA G-quadruplex in solution studied by kinetic capillary electrophoresis coupled on-line with mass spectrometry

Abstract

G-quadruplex-forming DNA/RNA sequences play an important role in the regulation of biological functions and development of new anticancer and anti-aging drugs. In this work, we couple on-line kinetic capillary electrophoresis with mass spectrometry (KCE-MS) to study conformational dynamics of DNA G-quadruplexes in solution. We show that peaks shift and its widening in KCE can be used for measuring rate and equilibrium constants for DNA-metal affinity interactions and G-quadruplex formation; and ion mobility mass spectrometry (IM-MS) provides information about relative sizes, absolute molecular masses and stoichiometry of DNA complexes. KCE-MS separates a thrombin-binding aptamer d[GGTTGGTGTGGTTGG] from mutated sequences based on affinity to potassium, and reveals the apparent equilibrium folding constant (KF≈150 μm), folding rate constant (kon≈1. 70×103 s-1 m-1), unfolding rate constant (koff≈0.25 s-1), half-life time of the G-quadruplex (t1/2≈2.8 s), and relaxation time (τ≈3.9 ms at physiological 150 mm [K+]). In addition, KCE-MS screens for a GQ-stabilizing/-destabilizing effect of DNA binding dyes and an anticancer drug, cisplatin. Combining forces! Kinetic capillary electrophoresis is coupled on-line with mass spectrometry (KCE-MS) to study conformational dynamics of DNA G-quadruplexes in solution. Peak's shift and its widening in KCE help measure rate and equilibrium constants; and ion mobility spectrometry (IMS) provides relative sizes, absolute molecular masses and stoichiometry. © 2014 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.