Berenil Binds Tightly to Parallel and Mixed Parallel/AntiparallelG?Quadruplex Motifs with Varied Thermodynamic Signatures

摘要

Diminazene, DMZ, (or berenil) has beenreported as a tight binder of G-quadruplexes. G-Quadruplexstructures are often located in the promotor regions ofoncogenes and may play a regulatory role in gene expressionbased on the stability of the folding topology. In this study,attempts have been made to characterize the specificity ofDMZ binding toward multiple G-quadruplex topologies orfoldamers. Mutant sequences of the G-quadruplex formingpromotor regions of several oncogenes were designed toexhibit restricted loop lengths and folding topologies. Circulardichroism was used to confirm the quadruplex topology ofmutant BCL2, KRAS, and c-MYC sequences, human telomere(Na+ and K+) G-quadruplexes and their complexes with DMZand analogs thereof. Isothermal titration calorimetry was used to generate a complete thermodynamic profile (ΔG, ΔH, ?TΔS)for the formation of DMZ and analog complexes with the target G-quadruplexes. DMZ binds to parallel and/or mixed parallel/antiparallel quadruplex DNA motifs with stoichiometries up to 8:1 and via three binding modes with varying affinities. In thecase of the parallel G-quadruplexes, with the exception of the long-looped c-MYC mutant, the highest affinity binding event(mode 1) is driven by enthalpy. DMZ binding to the long-looped c-MYC mutant exhibits a very favorable entropy change inaddition to a moderately favorable enthalpy change. Mode 1 binding to the antiparallel and mixed parallel/antiparallel hTelquadruplexes is also driven by favorable enthalpy changes. In all cases, the intermediate DMZ affinity binding (mode 2) isdriven almost entirely by entropy, with small or unfavorable enthalpic contributions. The weakest binding event (mode 3) isalso entropically driven with small or moderate enthalpic contributions.