Characterization of the mycobacteriophage L5 attachment site, attP.
Lysogenization of mycobacteriophage L5 involves integration of the phage genome into the Mycobacterium smegmatis chromosome. Integration occurs by a site-specific recombination event between a phage attachment site, attP, and a bacterial attachment site, attB, which is catalyzed by the phage-encoded integrase protein. DNase I footprinting reveals that L5 integrase binds to two types of sites within attP which span an unexpectedly large region of 413 bp: seven arm-type sites (P1 to P7) each of which correspond to a consensus sequence 5'-TGCaaCtcYy, and core-type sites at the points of strand exchange. Mutational analyses indicate that not all of the arm-type sites are required for integration, and that the P3 site and the rightmost pair of sites (P6 and P7) are dispensable for integration. We show that a 252 bp segment of attP DNA is sufficient for efficient integrative recombination and that int can be provided in trans for simple and efficient transformation of the mycobacteria.
Peña
C E
CE
Department of Biological Sciences, University of Pittsburgh, PA 15260, USA.
Lee
M H
MH
Pedulla
M L
ML
Hatfull
G F
GF
D001435:Bacteriophages / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D016384:Consensus Sequence; D018983:DNA Footprinting; D004252:DNA Mutational Analysis; D003850:Deoxyribonuclease I; D004926:Escherichia coli; D016679:Genome, Viral; D019426:Integrases / Q000378:metabolism*; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000235:genetics / Q000821:virology*; D011995:Recombination, Genetic; D017384:Sequence Deletion; D016662:Virus Integration
D019426:Integrases; D003850:Deoxyribonuclease I