Cytoplasmic RNA polymerase in Escherichia coli

To obtain an estimate for the concentration of free functional RNA polymera se in the bacterial cytoplasm, the content of RNA polymerase beta and beta' subunits in DNA-free minicells from the minicell-producing Escherichia col i strain chi 925 was determined. In bacteria grown in Luria-Bertani medium at 2.5 doublings/h, 1.0% of the total protein was RNA polymerase. The conce ntration of cytoplasmic RNA polymerase beta and beta' subunits in minicells produced by this strain corresponded to about 17% (or 2.5 muM) of the valu e found in whole cells. Literature data suggest that a similar portion of c ytoplasmic RNA polymerase subunits is in RNA polymerase assembly intermedia tes and imply that free functional RNA polymerase can form a small percenta ge of the total functional enzyme in the cell. On infection,vith bacterioph age T7, 20% of the minicells produced progeny phage, whereas infection in 8 0% of the cells was abortive. RNA polymerase subunits in lysozyme-freeze-th aw lysates of minicells were associated with minicell envelopes and were wi thout detectable activity in an in vitro transcription assay. Together, the se results suggest that most functional RNA polymerase is associated with t he DNA and that little if any segregates into DNA-free minicells.