Characterization of the attP site of the integrative element pSAM2 from Streptomyces ambofaciens

摘要

pSAM2 is integrated into the Streptomyces ambofaciens chromosome through site-specific recombination between the element (attP) and the chromosomal (attB) site. The 43?kDa integrase protein encoded by pSAM2 catalyses this recombination event. Tools have been developed to study site-specific recombination in Escherichia coli. In vivo studies showed that a 360?bp fragment of attP is required for efficient site-specific recombination and that int can be provided in trans. pSAM2 integrase was purified and overexpressed in E. coli and Int binding at the attP site was studied. DNaseI footprinting revealed two sites that bind integrase strongly and appear to be symmetrical with regard to the core site. These two P1/P2 arm-type sites both contain a 17?bp motif that is identical except at one position, GTCACGCAG(A/T)TAGACAC. P1 and P2 are essential for site-specific recombination.