In vitro interactions of Pseudomonas RNA polymerases with tac and RNA I promoters

Abstract

The activities of RNA polymerases from Pseudomonas putida and Pseudomonas aeruginosa were compared with that of Escherichia coli RNA polymerase in an in vitro transcription system. All three enzymes initiated transcription at the tac promoter and the RNA I promoter of E. coli. We measured the rate of open complex formation between the RNA polymerases and the promoters, and the saturation level of open complex formation at equilibrium in single-round transcription. The relative rates of open complex formation were P. putida > E. coli > P. aeruginosa and the relative saturation levels of open complex formation at equilibrium were E. coli > P. putida > P. aeruginosa for the tac and RNA I promoters. The interaction of the RNA polymerases with the promoters was also studied by DNase I footprinting. The patterns of protection of the Pseudomonas RNA polymerases on the tac promoter were similar to that of E. coli RNA polymerase. However, the protection patterns of the Pseudomonas RNA polymerases on the RNA I promoter were slightly different from that of E. coli RNA polymerase.