IDENTIFICATION OF A GENE ENCODING THE REPLICATION INITIATOR PROTEIN OF THE STREPTOMYCES INTEGRATING ELEMENT, PSAM2

pSAM2 is an 11-kilobase integrating element from Streptomyces ambofaci ens which was previously shown to generate single-stranded DNA during replication, indicating that it probably replicates by a rolling-circl e replication (RCR) mechanism. Two separate regions are involved in it s replication, one of which was shown to contain the plus origin of re plication (ds origin). We report here the study of the second region. Its nucleotide sequence was determined and analysed for open reading f rames (ORFs). Three putative ORFs were identified: orf183 (183 amino a cids (aa)), orf50 (50 aa), and repSA (459 aa). orf183 is not necessary for replication. The function of or150 is unknown. repSA is essential for pSAM2 replication; it could encode a protein, RepSA, presenting s imilarities to the replication initiator proteins (Rep) of elements th at replicate by an RCR mechanism. A derivative consisting of repSA, th e region containing ds origin, a Streptomyces antibiotic resistance ma rker, and pBR322, could replicate in Streptomyces, further demonstrati ng that this ORF encodes the major replication protein of pSAM2. repSA might be co-transcribed with the genes involved in integration and ex cision of pSAM2. (C) 1994 Academic Press, Inc.