SLP1 is a 17.2-kbp genetic element indigenous to the Streptomyces coelicolor chromosome. During conjugation, SLP1 can undergo excision and subsequent site-specific integration into the chromosomes of recipient cells. We report here the localization, nucleotide sequences, and initial characterization of the genes mediating these recombination events. A region of SLP1 adjacent to the previously identified site of integration, attP, was found to be sufficient to promote site-specific integration of an unrelated Streptomyces plasmid. Nucleotide sequence analysis of a 2.2-kb segment of this region reveals two open reading frames that are adjacent to and transcribed toward the attP site. One of these, the 1,365-bp int gene of SLP1, encodes a predicted 50.6-kDa basic protein having substantial amino acid sequence similarity to a family of site-specific recombinases that includes the Escherichia coli bacteriophage A integrase. A linker insertion in the 5' end of the cloned int gene prevents integration, indicating that Int is essential for promoting integration. An open reading frame (orf6M) lying immediately 5' to int encodes a predicted 7.1-kDa basic peptide showing limited sequence similarity to the excisionase (xis) genes of other site-specific recombination systems.Streptomyces are gram-positive filamentous soil bacteria notable for their complex morphology as well as the production of a variety of antibiotics and other secondary metabolites, many of which have medical or agricultural importance. Numerous extrachromosomal elements have been identified for these organisms, including a novel family of chromosomally integrated self-transmissible plasmidogenic genetic elements (reviewed in reference 35). SLP1, indigenous to the chromosome of Streptomyces coelicolor (6), was the first of this group to be identified. A model in which the integrated SLP1 element (i.e., SLPlr') can excise from the S. coelicolor chromosome during conjugation with a strain lacking SLP1, transfer to the recipient strain as a transiently existing circular molecule, and either integrate into the recipient chromosome at a site identical to that occupied in the donor strain or undergo deletion of sequences that suppress autonomous replication and consequently be maintained as an autonomously replicating plasmid (33) has been proposed.The recombination reactions mediated by SLP1 are sitespecific and involve 112-bp regions of homology on SLP1 (attP) and the host chromosome (attB) that have only a single-base-pair difference (34). The integration event generates sites at the left (attL) and right (attR) of the integrated element that retain the homologies between attP and attB. Characterization of these sequences (29) integrants. A derivative of SLP1 containing an insertion at a KpnI site 1.3 kb from attP ( Fig. 1), within a locus designated intB, fails to integrate. In addition, integration of SLP1-derived plasmids deleted for a second locus (i.e., intA, Fig. 1) situated at some distance from attP and intB was not observed. This defect could be c...

SLPlINt is a conjugative Streptomyces coelicolor genetic element that can transfer to Streptomyces lividans and integrate site specifically into the genome of the new bacterial host. Recombination of SLP1 previously has been shown to occur within nearly identical 112-base-pair act sequences on the plasmid and host chromosome. We report here that both integrative recombination and intermolecular transfer of SLPlINt require no more than a 48-base-pair segment of the att sequence and that SLP1 transfer occurs by a conservative rather than a replicative mechanism. The functions responsible for the excision of the element as a discrete DNA segment are induced during the conjugal transfer of SLP1.The SLP1 genetic element, which exists in nature as an integrated sequence (i.e., SLPlint) in the Streptomyces coelicolor chromosome, is capable of transfer into Streptomyces lividans and integration into a specific site on the S. lividans genome (1, 27). SLP1-mediated recombination events associated with transfer and integration occur at the attachment sites attP of SLP1 and attB of the host chromosome and at attL and attR, the recombination loci that flank the integrated element. The att sites are nearly identical 112-basepair (bp) sequences; a 1-bp difference occurring at position 95 yields a BclI site in attB and attR that is absent from attP and attL (Fig. 1). We previously have proposed (27,28) that SLPlint is transferred between species as a transiently existing, physically autonomous 17-kilobase (kb) plasmid that integrates into the attB site of the recipient upon entering the cell. Autonomously replicating SLP1-derived extrachromosomal elements (1, 27, 28) can result from deletion of segments of the SLP1 sequence that are required either for integration or for maintenance of SLP1 in the integrated state.Examination of the DNA sequences of the four SLP1 att sites reveals two regions of dyad symmetry (i.e., inverted repeats) (29) (Fig. 1). In several other systems, such inverted repeat sequences serve as loci for site-specific recombination (3,9,12,13,17,18). Both inverted repeats in the SLP1 att segments lie in the region where SLP1-mediated recombination events are known to occur (29). We report here the results of insertion and deletion analyses that identify regions of the att sites essential for SLP1-mediated recombination and localize more precisely the actual site of recombination. We also provide direct evidence that intermolecular transfer of SLPlint involves conservative excision of the 17-kb SLP1 sequence as a discrete DNA segment and show that the functions responsible for such excisive recombination are regulated rather than constitutively expressed. MATERIALS AND METHODSBacterial strains and plasmids. The plasmids pACYC177 (5), pUC18, and pUC19 (37) were used as vectors for cloning in Escherichia coli, and the pIJlOl-derived replicons pIJ364 (22) and pIJ702 (19) were used in S. lividans. E. coli MC1061 (4), S. lividans TK64 (16), and S. lividans 1326 (15) were * Corresponding author. used as cloning hosts...