Sigma 54 levels and physiological control of the Pseudomonas putida Pu promoter

The cellular levels of the alternative sigma factor σ of Pseudomonas putida have been examined in a variety of growth stages and culture conditions with a single-chain Fv antibody tailored for detection of scarce proteins. The levels of σ were also monitored in P. putida strains with knockout mutations inptsO orptsN, known to be required for the C-source control of the σ-dependent Pu promoter of the TOL plasmid. Our results show that ∼80 ± 26 molecules of σ exist per cell. Unlike that in relatives of Pseudomonas (e.g., Caulobacter), where fluctuations of σ determine adaptation and differentiation when cells face starvation, σ in P. putida remains unexpectedly constant at different growth stages, in nitrogen starvation and C-source repression conditions, and in the ptsO and ptsN mutant strains analyzed. The number of σ molecules per cell in P. putida is barely above the predicted number of σ-dependent promoters. These figures impose a framework on the mechanism by which Pu (and other σ-dependent systems) may become amenable to physiological control.