Function of IHF in λ DNA Packaging: I. Identification of the Strong Binding Site for Integration Host Factor and the Locus for Intrinsic Bending in cosB

Abstract

Integration host factor (IHF) plays an accessory role in λ DNA packaging. IHF affects the interaction of the λ DNA packaging protein, terminase, with cos, the site on λ. DNA at which terminase binds and introduces staggered nicks to generate cohesive ends of mature λ chromosomes cos includes cosB, the terminase binding site and cosN, the adjacent nicking site cosB includes multiple binding sites for gpNul, the small subunit of terminase, and an IHF binding site, I1. I1 contains two overlapping sequences, called I1A and I1B, that closely match the consensus sequence for IHF binding sites. The I1A sequence was determined to be the site of IHF binding by hydroxyl radical footprinting experiments. Comparison of the pattern of IHF-induced enhancements and diminishments at I1 with published patterns for IHF binding sites at the λ attachment site identifies I1A as the IHF binding site at I1. The conclusion that I1A is the IHF binding site was confirmed by studies with DNA mutant in I1A. The I1A- mutation, consisting of three adjacent base-pair changes in I1A, abolished IHF binding. In contrast to the I1A- mutation, a mutation in I1B, also consisting of three adjacent base-pair changes, caused a reduction in the affinity of IHF for I1A, and caused a reduction in the magnitude of the net intrinsic bending of cosλ.