Identification of an HP1 phage protein required for site-specific excision

Transposon insertion mutagenesis and transformation were used to locate genes responsible for excision in the temperature phage HP1 of Haemophilus influenzae. A 6.5 kb segment of DNA near the left end of the phage genome was sequenced, and 11 new open reading frames were identified. Two face-to-face overlapping promoter sequences organized these open reading frames into two operons transcribed in opposite directions. Interruption of the first open reading frame in the rightward operon created lysogens unable to produce phages. Provision of the uninterrupted open reading frame in trans restored phage production. The gene identified by this procedure, cox, was cloned and the protein product was expressed at high levels in Escherichia coli. The Cox protein is a 79-residue basic protein with a predicted strong helix-turn-helix DNA-binding motif. Extracts induced to express high levels of Cox contained a 9 kDa protein. These extracts inhibited integrative recombination and were required for excisive recombination mediated by HP1 integrase. The HP1 cox gene location is similar to that of the homologous excisive and regulatory genes from coliphages P2 and 186. These phages appear to share a distinctive organization of recombination proteins and transcriptional domains differing markedly from phage lambda and its relatives.