title: Simultaneous binding and bending of promoter DNA by the TATA binding protein: real time kinetic measurements.
abstract: The binding and bending of tetramethylrhodamine-5'-(GGGCTATAAAAGGG) duplex-3'-fluorescein by native Saccharomyces cerevisiae TATA binding protein (TBP) have been investigated using fluorescence resonance energy transfer. Probability distributions derived from fluorescein emission lifetime measurements show a decrease in the mean 3'-fluorescein-5'- rhodamine distance from 56.5 to 46.8 A upon binding of the oligomer to TBP, consistent with the DNA bend observed by X-ray crystallography. The kinetics, monitored in real time using stopped flow fluorimetry, demonstrate simultaneous binding and bending of a TATA box by TBP with a single second-order rate constant of (2.4 +/- 0.3) x 10(6) M-1 s-1 at 30 degrees C.
authors.affiliation: Department of Chemistry, University of Nebraska, Lincoln 68588-0304, USA.
authors.forename: K M
authors.initials: KM
authors.lastname: Parkhurst
authors.forename: M
authors.initials: M
authors.lastname: Brenowitz
authors.forename: L J
authors.initials: LJ
authors.lastname: Parkhurst
mesh_terms: D001483:Base Sequence; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D015342:DNA Probes; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012235:Rhodamines; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D016385:TATA Box; D035181:TATA-Box Binding Protein; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D015342:DNA Probes; D004268:DNA-Binding Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D009838:Oligodeoxyribonucleotides; D012235:Rhodamines; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; C005358:tetramethylrhodamine