DNA conformational changes associated with the cooperative binding of cI-repressor of bacteriophage λ to OR

Daniel Strahs, Michael Brenowitz

Research output: Contribution to journalArticle

31 Scopus citations

Abstract

The cI repressor protein (cI) maintains bacteriophage λ in the lysogenic state in infected Escherichia coli cells by binding cooperatively to three tandemly repeated sequences comprising the right operator (OR). Cooperative interactions occur between alternate pairs of cI dimers bound to adjacent sites. Although crystallographic studies have revealed the structure of the DNA in the 92 amino acid residue amino-terminal fragment-OLI complex, the structure of the DNA within the OR-cI complex with intact, cooperatively bound cI has not been described. In this study, the structure of the DNA within OR was quantitatively examined using sequence and structure-dependent nuclease cleavage patterns as a function of cI binding. The cooperative binding of cI to OR1 and OR2 induces a conformational change in the DNA of OR3 that is detectable by both DNase I and 5-phenyl-1,10-phenanthroline. Hydroxyl radical footprinting indicates the presence of an "A-tract" between OR1 and OR2 at the site of a run of four adenine-thymine base-pairs, implying a stable bend between the sites of approximately 18°. 5-Phenyl-1,10-phenanthroline footprinting reports conformational changes within the central base-pairs of all three sites that is dependent upon the sequence-specific binding of cI. The observed conformational changes are more extensive within OR2 and OR3 compared with OR1, consistent with an "induced-fit" model of sequence-specific recognition. A number of changes in nuclease reactivity within the individual binding sites were quantitatively correlated with cI binding at the other sites within OR. These results demonstrate that changes in the DNA structure are propagated among the sites in response to the binding of cI and imply a role for DNA sequence-dependent conformational changes in the mechanisms of both the intrinsic and cooperative binding reactions of cI to OR.

Original languageEnglish (US)
Pages (from-to)494-510
Number of pages17
JournalJournal of Molecular Biology
Volume244
Issue number5
DOIs
StatePublished - Jan 1 1994

Keywords

  • Bacteriophage λ
  • Cooperativity
  • DNA structure
  • Protein-DNA interactions
  • Thermodynamics

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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