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 (K F?150??m), folding rate constant (k on?1.70×10(3)?s(-1)?m(-1)), unfolding rate constant (k off?0.25?s(-1)), half-life time of the G-quadruplex (t 1/2?2.8?s), and relaxation time (??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.

SUBMITTER: Mironov GG 

PROVIDER: S-EPMC4000168 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature