Identification of a Gene Encoding the Replication Initiator Protein of the Streptomyces Integrating Element, pSAM2

Abstract pSAM2 is an 11-kilobase integrating element from Streptomyces ambofaciens which was previously shown to generate single-stranded DNA during replication, indicating that it probably replicates by a rolling-circle replication (RCR) mechanism. Two separate regions are involved in its replication, one of which was shown to contain the plus origin of replication ( ds origin ). We report here the study of the second regions. Its nucleotide sequence was determined and analysed for open reading frames (ORFs). Three putative ORFs were identified: orf183 (183 amino acids (aa)), orf50 (50 aa), and repSA (459 aa). orf183 is not necessary for replication. The function of orf50 is unknown. repSA is essential for pSAM2 replication; it could encode a protein. RepSA, presenting similarities to the replication initiator proteins (Rep) of elements that replicate by an RCR mechanism. A derivative consisting of repSA , the region containing ds origin , a Streptomyces antibiotic resistance marker, and pBR322, could replicate in Streptomyces , further demonstrating that this ORF encodes the major replication protein of pSAM2. repSA might be co-transcribed with the genes involved in integration and excision of pSAM2.