Function of IHF in λ DNA Packaging: II. Effects of Mutations Altering the IHF Binding Site and the Intrinsic Bend in cosB on λ Development

Abstract

cosB is the binding site on λ DNA for terminase, the phage DNA packaging protein. cosB contains three binding sites for gpNu1, the small subunit of terminase, and a site for integration host factor (IHF). IHF plays an accessory role in λ DNA packaging, and IHF stimulates the burst size of λ several-fold, presumably by assisting the interaction of terminase with cosB. The present work includes a study of the effect on λ development of a mutation, called I1A-, which consists of three adjacent base-pair changes in the IHF binding site. The I1A- mutation was found to abolish IHF stimulation of the λ burst size, indicating that IHF is unable to bind to the mutant I1A site. A second mutation, called I1B- and also consisting of three adjacent base-pair changes, is a mutation that reduces an intrinsic bend found in cosB. λ I1B- was more dependent on IHF than λ+, raising the possibility that the intrinsic bend in cosB plays a role in cos function for λ+ under the IHF- conditions. In vitro DNA packaging experiments established that the I1 mutations affect DNA packaging per se. A series of Nu1 mutations that create terminuses able to suppress a variety of cosB defects were found to suppress the defects of the I1A- and I1B- mutations under IHF- conditions.