title: Protein-protein interactions dominate the assembly thermodynamics of a transcription repression complex.
abstract: Assembly of the transcription repression complex at the Escherichia coli biotin biosynthetic operon occurs via coupled protein-protein and protein-DNA interactions in which the holoBirA dimer binds to the forty base pair biotin operator sequence. The thermodynamic driving forces for the assembly process have been dissected using sedimentation equilibrium measurements and DNaseI footprint titrations. Measurements of the temperature dependence of dimerization indicate that this process is strongly enthalpically opposed and is driven by a very favorable entropy. By contrast, the DNA binding step is enthalpically driven and opposed by a modest entropy. Neither step is accompanied by a heat capacity change. The convoluted protein-protein and protein-DNA binding reaction is dominated by the thermodynamic signature of the dimerization step. This observed dominance of the dimerization step illustrates the importance of dissecting complex DNA binding reactions into their constituent steps in elucidation of the thermodynamic driving forces for these processes. Measurements of the salt dependence of dimerization and DNA binding indicate modest contributions of electrostatic interactions to each contributing step as well as the total assembly of the repression complex. In light of the known structural features of this system, this modest dependence of the DNA binding equilibrium on salt concentration was unanticipated.
authors.affiliation: Department of Chemistry & Biochemistry, College of Chemical & Life Sciences, Center for Biological Structure & Organization, University of Maryland, College Park, Maryland 20742, USA.
authors.forename: Huaying
authors.initials: H
authors.lastname: Zhao
authors.forename: Emily
authors.initials: E
authors.lastname: Streaker
authors.forename: Weilan
authors.initials: W
authors.lastname: Pan
authors.forename: Dorothy
authors.initials: D
authors.lastname: Beckett
mesh_terms: D001483:Base Sequence; D001710:Biotin / Q000378:metabolism; D019731:Carbon-Nitrogen Ligases / Q000737:chemistry* / Q000378:metabolism; D018983:DNA Footprinting; D019281:Dimerization; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009876:Operon; D012097:Repressor Proteins / Q000737:chemistry* / Q000378:metabolism; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D001710:Biotin; D019731:Carbon-Nitrogen Ligases; C058125:birA protein, E coli