The Critical Roles of Conserved Glu 21 and Asp 23 of a Staphylococcal Anti-Anti-Sigma Factor.

Staphylococcus aureus and similar bacteria cope with stressful environments using a set of conserved proteins including σ^B, an alternative sigma factor. The initiation of transcription by σ^B is obstructed by RsbW, an anti-sigma factor. RsbW also associates and phosphorylates RsbV, an anti-anti-sigma factor. A modeling study previously suggested that Glu 21, Asp 23, and Tyr 54 of S. aureus RsbV form non-covalent bonds with Arg 23, an indispensable residue of cognate RsbW. Herein, we have noted that Glu 21, and Asp 23 are conserved residues, whereas Tyr 54 is a semi-conserved residue. Additionally, our MD simulation studies indicate that both Glu 21 and Asp 23 may maintain the structure of RsbV. To verify the computational data, two RsbV mutants, created by replacing Glu 21 and Asp 23 with an Ala residue, were elaborately investigated using some in vitro tools. The results reveal that both the above residues are critical for preserving the structure of RsbV. Interestingly, the RsbV mutant harboring Ala at position 23 was very little phosphorylated by RsbW. This mutant, compared to the RsbV mutant carrying Ala at position 21, also showed a weaker interaction with RsbW. The ways Glu 21 and Asp 23 keep various properties of RsbV intact have been discussed at length.