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Distinct DNA elements contribute to Rap1p affinity for its binding sites.
- Source :
-
Journal of molecular biology [J Mol Biol] 2004 May 14; Vol. 338 (5), pp. 877-93. - Publication Year :
- 2004
-
Abstract
- The essential Saccharomyces cerevisiae regulatory protein Rap1 contains two tandem Myb-like DNA binding sub-domains that interact with two defined DNA "hemisites", separated by a trinucleotide linker sequence. We have mapped the thermodynamically defined DNA-binding site of Rap1 by a primer extension method coupled with electrophoretic separation of bound and unbound DNAs. Relative to published consensus sequences, we detect binding interactions that extend 3 bp beyond the 5'-end of the putative DNA-binding site. This new site of interaction is located where the DNA minor groove faces the protein, and may account for the major DNA bending induced by Rap1p that previous studies have mapped to a site immediately upstream of the consensus binding site. In addition, we show that a minimal DNA-binding site made of one single consensus hemisite, preceded or followed by a spacer trinucleotide that interacts with the unstructured protein linker between the two Rap1p DNA binding domains, is able to bind the protein, although at lower affinity. These findings may explain the observed in vivo binding properties of Rap1p at many promoters that lack canonical binding sites.
- Subjects :
- Binding Sites
Chromatin metabolism
DNA Footprinting
Promoter Regions, Genetic
Protein Binding
Protein Structure, Tertiary
Saccharomyces cerevisiae metabolism
Shelterin Complex
DNA metabolism
Saccharomyces cerevisiae Proteins metabolism
Telomere-Binding Proteins metabolism
Transcription Factors metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0022-2836
- Volume :
- 338
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- Journal of molecular biology
- Publication Type :
- Academic Journal
- Accession number :
- 15111054
- Full Text :
- https://doi.org/10.1016/j.jmb.2004.03.047