title: A ruthenium dimer complex with a flexible linker slowly threads between DNA bases in two distinct steps.
abstract: Several multi-component DNA intercalating small molecules have been designed around ruthenium-based intercalating monomers to optimize DNA binding properties for therapeutic use. Here we probe the DNA binding ligand [μ-C4(cpdppz)2(phen)4Ru2](4+), which consists of two Ru(phen)2dppz(2+) moieties joined by a flexible linker. To quantify ligand binding, double-stranded DNA is stretched with optical tweezers and exposed to ligand under constant applied force. In contrast to other bis-intercalators, we find that ligand association is described by a two-step process, which consists of fast bimolecular intercalation of the first dppz moiety followed by ∼10-fold slower intercalation of the second dppz moiety. The second step is rate-limited by the requirement for a DNA-ligand conformational change that allows the flexible linker to pass through the DNA duplex. Based on our measured force-dependent binding rates and ligand-induced DNA elongation measurements, we are able to map out the energy landscape and structural dynamics for both ligand binding steps. In addition, we find that at zero force the overall binding process involves fast association (∼10 s), slow dissociation (∼300 s), and very high affinity (Kd ∼10 nM). The methodology developed in this work will be useful for studying the mechanism of DNA binding by other multi-step intercalating ligands and proteins.
authors.affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA.
authors.forename: Meriem
authors.initials: M
authors.lastname: Bahira
authors.affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA.
authors.forename: Micah J
authors.initials: MJ
authors.lastname: McCauley
authors.affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA.
authors.forename: Ali A
authors.initials: AA
authors.lastname: Almaqwashi
authors.affiliation: Department of Chemistry and Chemical Engineering, Chalmers University of Technology, Gothenburg, Sweden.
authors.forename: Per
authors.initials: P
authors.lastname: Lincoln
authors.affiliation: Department of Biology and Biological Engineering, Chalmers University of Technology, Gothenburg, Sweden.
authors.forename: Fredrik
authors.initials: F
authors.lastname: Westerlund
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Ioulia
authors.initials: I
authors.lastname: Rouzina
authors.affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA mark@neu.edu.
authors.forename: Mark C
authors.initials: MC
authors.lastname: Williams
mesh_terms: D004247:DNA / Q000737:chemistry*; D007364:Intercalating Agents / Q000737:chemistry*; D007700:Kinetics; D009942:Organometallic Compounds / Q000737:chemistry*; D010618:Phenanthrolines / Q000737:chemistry*
chemical_list: D007364:Intercalating Agents; D009942:Organometallic Compounds; D010618:Phenanthrolines; C470943:mu-C4-bis(12-cyano-12,13-dihydro-11H-cyclopenta(b)dipyrido(3,2-h-2',3'-j)phenazine-12-carbonyl)tetrakis(1,10-phenathroline)diruthenium(II); D004247:DNA