Studies on the synthesis of plasmid-coded proteins and their control in Bacillus subtilis minicells.

The minicell system of Bacillus subtilis has been used to study the expression of plasmid genes using several R plasmids derived from Staphylococcus aureus. pE194, pC194, and pUB110 as well as several mutant and in vitro recombinant derivatives of these plasmids segregate into minicells. A copy control mutant of pE194 was used to show that the extent of segregation is proportional to the copy number. The polypeptides specified by these plasmids were examined by SDS-polyacrylamide gel electrophoresis. Six proteins specified by pE194, an erythromycin resistance plasmid, were identified using cop mutants. These comprise about 90% of the potential coding capacity of the 2.4-Mdal pE194 plasmid. One of these proteins (29,000 daltons) is inducible by erythromycin in the wild type pE194 but is synthesized constitutively in a mutant derivative which also expresses antibiotic resistance constitutively. Several other proteins are detected only in copy control mutants. pUB110, a kanamycin resistance plasmid, expresses three major proteins which comprise 50% of the coding capacity of this 3.0-Mdal plasmid. Two additional minor proteins are occasionally observed. pC194 (2.0 Mdal), which confers chloramphenicol resistance, expresses two polypeptides comprising about 25% of its coding capacity. One of these polypeptides (22,000 daltons) is inducible by chloramphenicol. pBD9, an in vitro composite of pUB110 and pE194, probably expresses all of the major parental plasmid proteins with the exception of one from pUB110 and one from pE194.