GATC MOTIFS MAY ALTER THE CONFORMATION OF DNA DEPENDING ON SEQUENCE CONTEXT AND N-6-ADENINE METHYLATION STATUS - POSSIBLE IMPLICATIONS FOR DNA-PROTEIN RECOGNITION

As part of our analysis of the role of a uniquely clustered set of dam methylation sites (the motif GATC) within the origin of DNA replicati on in Escherichia coli, we have studied the effect of GATCs in various methylation states on the intrinsic curvature of DNA. We have designe d a set of DNA linkers and used commercially available linkers contain ing GATC motifs. The linkers were ligated and the electrophoretic mobi lity of the resulting multimers in different states of methylation was tested relative to reference fragments. We report that properly phase d GATCs in certain sequence environments modulate DNA curvature and th at these effects may be enhanced by N-6-adenine methylation of the GAT Cs. These structural alterations may in turn affect DNA-protein intera ctions, especially those involving proteins that rely on both primary sequence and structure for recognition. We present an example, where i ntroduction of a GATC within an integration host factor (IHF) binding site, which does not alter the consensus sequence, reduces the binding affinity of the protein for the modified site.