Direct Evidence for DNA Bending at the Lambda Replication Origin

Abstract

Replication initiation in bacteriophage lambda appears to require wrapping of origin DNA on an approximately 50 angstrom radius in or around the complex with the initiator protein O. Since short lengths of DNA are not that flexible, it may be that runs of coherently spaced deoxyadenylate residues constitute bend sites in the ori sequence that facilitate the process. Earlier data showed that ori DNA has electrophoretic anomalies characteristic of bend sites and that these are augmented by initiator protein binding. Here origin bending is examined by direct measurement of the ability of polymerized ori sequences to form small circles. The smallest circles observed (84 residues) are compatible with the required radius of curvature. Bend sites within the O protein binding sites, bend sites in the spacers between them, plus the inherent flexibility of non-bent DNA in the origin may all contribute to origin bending. The data also show that a bend site is required for O protein binding to DNA.