EFFECTS OF RIFAMPICIN ON SYNTHESIS AND FUNCTIONAL ACTIVITY OF DNA-DEPENDENT RNA-POLYMERASE IN ESCHERICHIA-COLI

During the course of kinetic studies on the synthesis of RNA polymerase [EC 2.7.7.6] subunits in E. coli K12, strain Km7 (CP372), certain anomalies were found that seemed to be associated with the system of reversible inhibition of RNA and protein synthesis by rifampicin. The effects of rifampicin on RNA chain elongation and on residual synthesis of polymerase subunits were investigated with several strains including Km7 to explain these anomalies. Examination of mRNA synthesis for the tryptophan operon suggested that RNA chain growth and RNA chain initiation are inhibited at high drug concentration (500 .mu.g/ml); RNA chain initiation is inhibited specifically at low concentration (20 .mu.g/ml). Analysis of rifampicin concentration effect on total RNA synthesis gave results that are also consistent with this conclusion. The need for selecting a proper drug concentration whenever rifampicin or other related antibiotic is used as a specific inhibitor of transcription initiation is emphasized. When rifampicin was added to a culture of these strains, absolute rates of synthesis of all subunits of RNA polymerase increased for several minutes and then decreased. The extent of this transient stimulation varied depending on the strain, drug concentration and other conditions, but was most striking for the .beta.'' and .sigma. subunits with strain Km7 at high drug concentration (500 .mu.g/ml). with a rifampicin-sensitive wild-type strain, the maximum stimulation was at .apprx. 50 .mu.g/ml with a particularly marked effect for .sigma. subunit. Streptolydigin inhibited the synthesis of core subunits much faster than the bulk of the protein, but inhibited synthesis of .sigma. subunit only after a lag. A specific effect of rifampicin but not the inactivation of .beta. subunit per se may be involved in transient stimulation of polymerase synthesis. Implications of these findings on the control of RNA polymerase synthesis are discussed.