Expression of the Bacillus subtilis polC gene in Escherichia coli

Abstract

We have cloned a 14 kb DNA segment containing the coding sequence (polC) for DNA polymerase III (PolIII) from the Bacillus subtilis chromosome. The plasmid carrying the sequence, pRO10, directs the synthesis of the 160 kDa PolIII protein and three additional polypeptides in Escherichia coli maxicells from strain CSR603. A set of deletion derivatives of pRO10 was constructed in vitro. The location of the PolIII coding sequence and the direction of transcription through the polC gene were determined by analysis of the truncated polypeptides observed in extracts of CSR603 transformants. Two HindIII segments subcloned from pRO10 were found to contain promoter sequences which function in E. coli and in vegetative phase B. subtilis cells. The location of the promoter sequence was determined with respect to the polC gene. The B. subtilis polC gene did not complement the temperature-sensitive defect of an E. coli PolIII mutant (dnaE486). The presence of the complete B. subtilis polC gene on a multicopy plasmid inhibited the growth of E. coli cells.