Characterization of the gene products produced in minicells by pSM1, a derivative of R100

Abstract

At least ten polypeptides larger than 6 kilodaltons (K) are produced in minicells from the miniplasmid pSM1 in vivo. pSM1 (5804 bp) is a small derivative of the drug resistance plasmid R100 (ca. 90 kb) and carries the R100 essential replication region as well as some non-essential functions. Cloned restriction fragments of pSM1 and plasmids with deletions within pSM1 sequences were used to assign eight of the ten observed polypeptides to specific coding regions of pSM1. Two of these polypeptides were identified as RepA1 and RepA2, proteins encoded by the essential replication region of pSM1/R100. The nucleotide sequence consisting of 885 bp outside the essential replication region is presented here. This sequence contains an open reading frame, orf4, for a protein 22.9 K in size, and one of the pSM1-encoded polypeptides was identified as the orf4 gene product. Five additional polypeptides were shown to be the products of other open reading frames mapping outside the essential replication region. Specific functions have been assigned to four of these polypeptides and tentatively to the fifth.