Binding interface between the Salmonella σS/RpoS subunit of RNA polymerase and Crl: hints from bacterial species lacking crl

In many Gram-negative bacteria, including Salmonella enterica serovar Typhimurium (S. Typhimurium), the sigma factor RpoS/sigma(S) accumulates during stationary phase of growth, and associates with the core RNA polymerase enzyme (E) to promote transcription initiation of genes involved in general stress resistance and starvation survival. Whereas sigma factors are usually inactivated upon interaction with anti-sigma proteins, sigma(S) binding to the Crl protein increases sigma(S) activity by favouring its association to E. Taking advantage of evolution of the sigma(S) sequence in bacterial species that do not contain a crl gene, like Pseudomonas aeruginosa, we identified and assigned a critical arginine residue in sigma(S) to the S. Typhimurium sigma(S)-Crl binding interface. We solved the solution structure of S. Typhimurium Crl by NMR and used it for NMR binding assays with sigma(S) and to generate in silico models of the sigma(S)-Crl complex constrained by mutational analysis. The sigma(S)-Crl models suggest that the identified arginine in sigma(S) interacts with an aspartate of Crl that is required for sigma(S) binding and is located inside a cavity enclosed by flexible loops, which also contribute to the interface. This study provides the basis for further structural investigation of the sigma(S)-Crl complex.