Binding interface between the Salmonella σ(S)/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/σ(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 σ factors are usually inactivated upon interaction with anti-σ proteins, σ(S) binding to the Crl protein increases σ(S) activity by favouring its association to E. Taking advantage of evolution of the σ(S) sequence in bacterial species that do not contain a crl gene, like Pseudomonas aeruginosa, we identified and assigned a critical arginine residue in σ(S) to the S. Typhimurium σ(S)-Crl binding interface. We solved the solution structure of S. Typhimurium Crl by NMR and used it for NMR binding assays with σ(S) and to generate in silico models of the σ(S)-Crl complex constrained by mutational analysis. The σ(S)-Crl models suggest that the identified arginine in σ(S) interacts with an aspartate of Crl that is required for σ(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 σ(S)-Crl complex.
Institut Pasteur, Laboratoire Systèmes Macromoléculaires et Signalisation, Département de Microbiologie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS ERL3526, rue du Docteur Roux, 75015 Paris, France.
Paola
P
Cavaliere
Institut de Chimie des Substances Naturelles, CNRS UPR2301, 91190 Gif-sur-Yvette, France.
Christina
C
Sizun
Institut Pasteur, Plate-forme de Protéines Recombinantes, Département de Biologie Structurale et Chimie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS UMR 3528, rue du Dr. Roux, 75015 Paris, France.
Fabienne
F
Levi-Acobas
Institut Pasteur, Plate-forme de Protéines Recombinantes, Département de Biologie Structurale et Chimie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS UMR 3528, rue du Dr. Roux, 75015 Paris, France.
Mireille
M
Nowakowski
Institut Pasteur, Laboratoire Systèmes Macromoléculaires et Signalisation, Département de Microbiologie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS ERL3526, rue du Docteur Roux, 75015 Paris, France.
Véronique
V
Monteil
Institut de Chimie des Substances Naturelles, CNRS UPR2301, 91190 Gif-sur-Yvette, France.
François
F
Bontems
Institut Pasteur, Plate-forme de Protéines Recombinantes, Département de Biologie Structurale et Chimie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS UMR 3528, rue du Dr. Roux, 75015 Paris, France.
Jacques
J
Bellalou
CNRS UMR 3528, rue du Dr. Roux, 75015 Paris, France.|Institut Pasteur, Unité de Microbiologie Structurale, Département de Biologie Structurale et Chimie, 25 rue du Docteur Roux, 75015 Paris, France.|Université Paris Diderot, Sorbonne Paris Cité, Paris, France.
Claudine
C
Mayer
Institut Pasteur, Laboratoire Systèmes Macromoléculaires et Signalisation, Département de Microbiologie, 25 rue du Docteur Roux, 75015 Paris, France.|CNRS ERL3526, rue du Docteur Roux, 75015 Paris, France.
Françoise
F
Norel
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D008956:Models, Chemical; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D021122:Protein Subunits; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D012475:Salmonella / Q000378:metabolism*; D012808:Sigma Factor / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D013045:Species Specificity; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D021122:Protein Subunits; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria; C077481:Crl protein, Bacteria; D012321:DNA-Directed RNA Polymerases