ROLE OF ARCHITECTURAL ELEMENTS IN COMBINATORIAL REGULATION OF INITIATION OF DNA-REPLICATION IN ESCHERICHIA-COLI

Bending of DNA is a prerequisite of site-specific recombination and ge ne expression in many regulatory systems involving the assembly of spe cific nucleoprotein complexes. We have investigated how the uniquely c lustered Dam methylase sites, GATCs, in the origin of Escherichia coli replication (oriC) and their methylation status modulate the geometry of oriC and its interaction with architectural proteins, such as inte gration host factor (IHF), factor for inversion stimulation (Fis) and DnaA initiator protein. We note that 3 of the 11 GATC sites at oriC ar e strategically positioned within the IHF protected region. Methylatio n of the GATCs enhances IHF binding and alters the IHF-induced bend at oriC. GATC motifs also contribute to intrinsic DNA curvature at oriC and the degree of bending is modulated by methylation. The IHF-induced bend at oriC is further modified by Fis protein and IHF affinity for its binding site may be impaired by protein(s) binding to GATCs within the IHF site. Thus, GATC sites at oriC affect the DNA conformation an d GATCs, in conjunction with the protein-induced bends, are critical c is-acting elements in specifying proper juxtapositioning of initiation factors in the early steps of DNA replication.