Binding between the par region of plasmids R1 and pSC101 and the outer membrane fraction of the host bacteria

Abstract

The binding between par+ and par plasmid DNA to different membrane fractions of Escherichia coli was investigated. Membrane material from cells carrying different Par+ and Par- derivatives of plasmids R1 and pSC101 was isolated and fractionated into an outer and a cytoplasmic membrane fraction. The presence of plasmid DNA in the two membrane fractions was measured either by nick-translation of the membrane-bound DNA, followed by filter-hybridization to homologous DNA, or by filter-hybridization of the membrane-bound DNA to nick-translated homologous purified plasmid DNA. The DNA of par derivatives of plasmids R1 and pSC101 could be detected only in the cytoplasmic membrane fraction, whereas the corresponding par+ plasmid DNA also appeared in the outer membrane material, indicating a specific binding between the R1 and pSC101 partition loci and the bacterial outer membrane. The experiment was then modified by fractionation of the membrane material from cells carrying hybrids between the vector pSF2124 and the par region or the basic replicon region of plasmid R1. The DNA of the membrane fractions were filter-hybridized to nick-translated probes. Again, the par+ region caused hybridization to the outer membrane material. Therefore, we may conclude that controlled partitioning involves binding of DNA to membrane material that has the same density as the outer membrane of the host bacteria. This finding offers a biochemical 'assay' for studies of the molecular biology of plasmid partitioning.