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OMP peptides activate the DegS stress-sensor protease by a relief of inhibition mechanism.

ABSTRACT: In the E. coli periplasm, C-terminal peptides of misfolded outer-membrane porins (OMPs) bind to the PDZ domains of the trimeric DegS protease, triggering cleavage of a transmembrane regulator and transcriptional activation of stress genes. We show that an active-site DegS mutation partially bypasses the requirement for peptide activation and acts synergistically with mutations that disrupt contacts between the protease and PDZ domains. Biochemical results support an allosteric model, in which these mutations, active-site modification, and peptide/substrate binding act in concert to stabilize proteolytically active DegS. Cocrystal structures of DegS in complex with different OMP peptides reveal activation of the protease domain with varied conformations of the PDZ domain and without specific contacts from the bound OMP peptide. Taken together, these results indicate that the binding of OMP peptides activates proteolysis principally by relieving inhibitory contacts between the PDZ domain and the protease domain of DegS.

SUBMITTER: Sohn J

PROVIDER: S-EPMC2764547 | biostudies-literature | 2009 Oct

REPOSITORIES: biostudies-literature

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OMP peptides activate the DegS stress-sensor protease by a relief of inhibition mechanism.

Sohn Jungsan J Grant Robert A RA Sauer Robert T RT

Structure (London, England : 1993) 20091001 10

In the E. coli periplasm, C-terminal peptides of misfolded outer-membrane porins (OMPs) bind to the PDZ domains of the trimeric DegS protease, triggering cleavage of a transmembrane regulator and transcriptional activation of stress genes. We show that an active-site DegS mutation partially bypasses the requirement for peptide activation and acts synergistically with mutations that disrupt contacts between the protease and PDZ domains. Biochemical results support an allosteric model, in which th ...[more]

PMID: 19836340

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Project description:In bacteria, DegS protease functions as an activating factor of the σE envelope stress response system, which ultimately activates the transcription of stress response genes in the cytoplasm. On the basis of high-throughput RNA sequencing, we have previously found that degS knockout inhibits the expression of flagellum synthesis- and chemotaxis-related genes, thereby indicating that DegS may be involved in the regulation of V. cholerae motility. In this study, we examined the relationships between DegS and motility in V. cholerae. Swimming motility and chemotaxis assays revealed that degS or rpoE deletion promotes a substantial reduction in the motility and chemotaxis of V. cholerae, whereas these activities were restored in ΔdegS::degS and ΔdegSΔrseA strains, indicating that DegS is partially dependent on σE to positively regulate V. cholerae activity. Gene-act network analysis revealed that the cAMP-CRP-RpoS signaling pathway, which plays an important role in flagellar synthesis, is significantly inhibited in ΔdegS mutants, whereas in response to the overexpression of cyaA/crp and rpoS in the ΔdegS strain, the motility and chemotaxis of the ΔdegS + cyaA/crp and ΔdegS + rpoS strains were partially restored compared with the ΔdegS strain. We further demonstrated that transcription levels of the flagellar regulatory gene flhF are regulated by DegS via the cAMP-CRP-RpoS signaling pathway. Overexpression of the flhF gene in the ΔdegS strain partially restored motility and chemotaxis. In addition, suckling mouse intestinal colonization experiments indicated that the ΔdegS and ΔrpoE strains were characterized by the poor colonization of mouse intestines, whereas colonization efficacy was restored in the ΔdegSΔrseA, ΔdegS + cyaA/crp, ΔdegS + rpoS, and ΔdegS + flhF strains. Collectively, our findings indicate that DegS regulates the motility and chemotaxis of V. cholerae via the cAMP-CRP-RpoS-FlhF pathway, thereby influencing the colonization of suckling mouse intestines.

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OMP peptides activate the DegS stress-sensor protease by a relief of inhibition mechanism.

Publications

OMP peptides activate the DegS stress-sensor protease by a relief of inhibition mechanism.

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