title: 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 μm), 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 (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.
authors.affiliation: Department of Chemistry, University of Ottawa 10 Marie Curie, Ottawa K1N 6N5 (Canada).
authors.forename: Gleb G
authors.initials: GG
authors.lastname: Mironov
authors.affiliation: Department of Chemistry, University of Ottawa 10 Marie Curie, Ottawa K1N 6N5 (Canada).
authors.forename: Victor
authors.initials: V
authors.lastname: Okhonin
authors.affiliation: Department of Chemistry, University of Ottawa 10 Marie Curie, Ottawa K1N 6N5 (Canada).
authors.forename: Nasrin
authors.initials: N
authors.lastname: Khan
authors.affiliation: Department of Chemistry, University of Ottawa 10 Marie Curie, Ottawa K1N 6N5 (Canada).
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Clouthier
authors.affiliation: Department of Chemistry, University of Ottawa 10 Marie Curie, Ottawa K1N 6N5 (Canada).
authors.forename: Maxim V
authors.initials: MV
authors.lastname: Berezovski
keywords: DNA folding; G-quadruplexes; capillary electrophoresis; kinetics; mass spectrometry; structure characterization of biomolecules