Snf2/Swi2-related ATPase Mot1 drives displacement of TATA-binding protein by gripping DNA.
Mot1 is a conserved Snf2/Swi2-related transcriptional regulator that uses ATP hydrolysis to displace TATA-binding protein (TBP) from DNA. Several models of the enzymatic mechanism have been proposed, including Mot1-catalyzed distortion of TBP structure, competition between Mot1 and DNA for the TBP DNA-binding surface, and ATP-driven translocation of Mot1 along DNA. Here, DNase I footprinting studies provide strong support for a 'DNA-based' mechanism of Mot1, which we propose involves ATP-driven DNA translocation. Mot1 forms an asymmetric complex with the TBP core domain (TBPc)-DNA complex, contacting DNA both upstream and within the major groove of the TATA Box. Contact with upstream DNA is required for Mot1-mediated displacement of TBPc from DNA. Using the SsoRad54-DNA complex as a model, DNA-binding residues in Mot1 were identified that are critical for Mot1-TBPc-DNA complex formation and catalytic activity, thus placing Mot1 mechanistically within the helicase superfamily. We also report a novel ATP-independent TBPc displacement activity for Mot1 and describe conformational heterogeneity in the Mot1 ATPase, which is likely a general feature of other enzymes in this class.
Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, VA 22908-0733, USA.
Rebekka O
RO
Sprouse
Michael
M
Brenowitz
David T
DT
Auble
D000251:Adenosine Triphosphatases / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D008958:Models, Molecular; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics; D035381:TATA-Binding Protein Associated Factors / Q000737:chemistry* / Q000235:genetics; D035181:TATA-Box Binding Protein / Q000737:chemistry* / Q000235:genetics; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics
D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; C073676:MOT1 protein, S cerevisiae; C499333:SNF2 protein, S cerevisiae; D004265:DNA Helicases