Lambda+ is able to grow in Escherichia coli cells lacking integration host factor (IHF), producing a burst of approximately 25% that produced in IHF+ cells. In vitro, however, we find that the lambda DNA packaging enzyme terminase is strongly dependent on IHF in both cos cleavage reactions and DNA packaging reactions. The cos59 mutation renders lambda dependent on IHF in vivo. The cos59 mutation is a deletion of 3 base pairs at the XmnI site in the cohesive end site (cos) of lambda. Variants of lambda cos59 that were able to grow in the absence of IHF were isolated and found to carry a mutation, called ms1, in the Nu1 gene, which codes for the small subunit of terminase. The Nu1ms1 mutation results in a change of the 40th amino acid of the Nu1 gene product from leucine to phenylalanine. The Nu1ms1 terminase was independent of IHF in packaging reactions in vitro. The results indicate that the mutation either renders terminase: (1) able to utilize some host protein other than IHF, or (2) totally independent of host factors.