Identification and characterization of mycobacteriophage L5 excisionase

The well-characterized mycobacteriophage L5 forms stable lysogens in Mycobacterium smegmatis. Establishment of lysogeny involves integration of the phage genome into the chromosome of its mycobacterial hosts through an integrase-mediated site-specific recombination event. As L5 lysogens spontaneously generate free phage particles, prophage excision must also occur, although an L5 excisionase gene had not been identified. We show here that L5 gene 36 encodes the phage excisionase and is a small, heat-stable 56-amino-acid protein that strongly stimulates excisive recombination both in vivo and in vitro. The ability to manipulate the highly directional phage integration and excision reactions will provide powerful tools for the introduction, curing and recovery of foreign genes in recombinant mycobacterial strains.