title: Single-molecule FRET analysis of DNA binding and bending by yeast HMGB protein Nhp6A.
abstract: High-mobility group B (HMGB) proteins bind duplex DNA without sequence specificity, facilitating the formation of compact nucleoprotein structures by increasing the apparent flexibility of DNA through the introduction of DNA kinks. It has remained unclear whether HMGB binding and DNA kinking are simultaneous and whether the induced kink is rigid (static) or flexible. The detailed molecular mechanism of HMGB-induced DNA 'softening' is explored here by single-molecule fluorescence resonance energy transfer studies of single yeast Nhp6A (yNhp6A) proteins binding to short DNA duplexes. We show that the local effect of yNhp6A protein binding to DNA is consistent with formation of a single static kink that is short lived (lifetimes of a few seconds) under physiological buffer conditions. Within the time resolution of our experiments, this static kink occurs at the instant the protein binds to the DNA, and the DNA straightens at the instant the protein dissociates from the DNA. Our observations support a model in which HMGB proteins soften DNA through random dynamic binding and dissociation, accompanied by DNA kinking and straightening, respectively.
authors.affiliation: Department of Physics, Emory University, Atlanta, GA 30322, USA.
authors.forename: Julie E
authors.initials: JE
authors.lastname: Coats
authors.forename: Yuyen
authors.initials: Y
authors.lastname: Lin
authors.forename: Emily
authors.initials: E
authors.lastname: Rueter
authors.forename: L James
authors.initials: LJ
authors.lastname: Maher
authors.forename: Ivan
authors.initials: I
authors.lastname: Rasnik
mesh_terms: D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D024001:HMGN Proteins / Q000737:chemistry* / Q000378:metabolism; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D024001:HMGN Proteins; C086471:NHP6A protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA