title: Facilitated dissociation of transcription factors from single DNA binding sites.
abstract: The binding of transcription factors (TFs) to DNA controls most aspects of cellular function, making the understanding of their binding kinetics imperative. The standard description of bimolecular interactions posits that TF off rates are independent of TF concentration in solution. However, recent observations have revealed that proteins in solution can accelerate the dissociation of DNA-bound proteins. To study the molecular basis of facilitated dissociation (FD), we have used single-molecule imaging to measure dissociation kinetics of Fis, a keyEscherichia coliTF and major bacterial nucleoid protein, from single dsDNA binding sites. We observe a strong FD effect characterized by an exchange rate [Formula: see text], establishing that FD of Fis occurs at the single-binding site level, and we find that the off rate saturates at large Fis concentrations in solution. Although spontaneous (i.e., competitor-free) dissociation shows a strong salt dependence, we find that FD depends only weakly on salt. These results are quantitatively explained by a model in which partially dissociated bound proteins are susceptible to invasion by competitor proteins in solution. We also report FD of NHP6A, a yeast TF with structure that differs significantly from Fis. We further perform molecular dynamics simulations, which indicate that FD can occur for molecules that interact far more weakly than those that we have studied. Taken together, our results indicate that FD is a general mechanism assisting in the local removal of TFs from their binding sites and does not necessarily require cooperativity, clustering, or binding site overlap.
authors.affiliation: Department of Molecular Biosciences, Northwestern University, Evanston, IL 60208.
authors.forename: Ramsey I
authors.initials: RI
authors.lastname: Kamar
authors.affiliation: Department of Physics and Astronomy, Northwestern University, Evanston, IL 60208.
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Banigan
authors.affiliation: Department of Materials Science and Engineering, Northwestern University, Evanston, IL 60208.
authors.forename: Aykut
authors.initials: A
authors.lastname: Erbas
authors.affiliation: Department of Molecular Biosciences, Northwestern University, Evanston, IL 60208.
authors.forename: Rebecca D
authors.initials: RD
authors.lastname: Giuntoli
authors.affiliation: Department of Physics and Astronomy, Northwestern University, Evanston, IL 60208; m-olvera@northwestern.edu john-marko@northwestern.edu.|Department of Materials Science and Engineering, Northwestern University, Evanston, IL 60208.|Department of Chemistry, Northwestern University, Evanston, IL 60208.
authors.forename: Monica
authors.initials: M
authors.lastname: Olvera de la Cruz
authors.affiliation: Department of Biological Chemistry, David Geffen School of Medicine at the University of California, Los Angeles, CA 90095.
authors.forename: Reid C
authors.initials: RC
authors.lastname: Johnson
authors.affiliation: Department of Molecular Biosciences, Northwestern University, Evanston, IL 60208; m-olvera@northwestern.edu john-marko@northwestern.edu.|Department of Physics and Astronomy, Northwestern University, Evanston, IL 60208.
authors.forename: John F
authors.initials: JF
authors.lastname: Marko
mesh_terms: D001665:Binding Sites; D004271:DNA, Fungal / Q000737:chemistry / Q000378:metabolism*; D024001:HMGN Proteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004271:DNA, Fungal; C464859:FIS1 protein, S cerevisiae; D024001:HMGN Proteins; D024101:Mitochondrial Proteins; C086471:NHP6A protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors
keywords: DNA–protein interactions; biomolecule binding; chemical kinetics; facilitated dissociation; transcription factor