Ss-LrpB, a novel Lrp-like regulator of Sulfolobus solfataricus P2, binds cooperatively to three conserved targets in its own control region.
Ss-LrpB, a novel Lrp-like DNA-binding protein from the hyperthermophilic crenarchaeon Sulfolobus solfataricus, was shown to bind cooperatively to three regularly spaced targets in its own control region, with as consensus the 15 bp palindrome 5'-TTGYAW WWWWTRCAA-3'. Binding to the border sites occurred with high affinity; the target in the middle proved to be a low affinity site which is stably bound only when both flanking sites are occupied. Ss-LrpB contacts two major groove segments and the intervening minor groove of each site, all aligned on one face of the helix. The operator shows intrinsic bending and is increasingly deformed upon binding of Ss-LrpB to one, two and three targets. Complex formation relies therefore on DNA conformability, protein-DNA and protein-protein contacts. Mobility-shift assays and in gel footprinting indicate that Ss-LrpB and the transcription factors TATA-box binding protein (TBP) and transcription factor B (TFB) can bind simultaneously to the control region. Based on these findings we present a model for the construction of the higher order nucleoprotein complexes and a hypothesis for the autoregulatory process. The latter is based on the concentration-dependent formation of distinct complexes exhibiting different stoichiometries and conformations, which could positively and negatively affect promoter activity.
Peeters
Eveline
E
Erfelijkheidsleer en Microbiologie, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
Thia-Toong
Thia-Lin
TL
Gigot
Daniel
D
Maes
Dominique
D
Charlier
Daniel
D
D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D018983:DNA Footprinting; D019848:Gene Expression Regulation, Archaeal; D019901:Locus Control Region; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D048229:Sulfolobus solfataricus / Q000235:genetics* / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism
D019843:Archaeal Proteins; D046911:Macromolecular Substances; D014157:Transcription Factors