Structural Basis of Response Regulator Inhibition by a Bacterial Anti-Activator Protein

Baker, Melinda D.; Neiditch, Matthew B.

doi:10.1371/journal.pbio.1001226

Cited by 52 publications

(107 citation statements)

References 68 publications

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“…This structure resembles a TPR domain superhelix, but the primary amino acid sequence does not (11) and ComA (5). The RRNPP proteins are depicted here as dimers, but Rap proteins were shown to be monomeric in solution (16,44), and PrgX likely forms tetramers (18). In contrast to the RRNPP systems, extracellular pheromone detection occurs by two-component signal transduction (TCST) pathways that control transcription through phosphorylation of a response regulator.…”

Section: Resultsmentioning

confidence: 99%

Rgg protein structure–function and inhibition by cyclic peptide compounds

Parashar

Aggarwal

Federle

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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100

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Peptide pheromone cell-cell signaling (quorum sensing) regulates the expression of diverse developmental phenotypes (including virulence) in Firmicutes, which includes common human pathogens, e.g., Streptococcus pyogenes and Streptococcus pneumoniae. Cytoplasmic transcription factors known as "Rgg proteins" are peptide pheromone receptors ubiquitous in Firmicutes. Here we present X-ray crystal structures of a Streptococcus Rgg protein alone and in complex with a tight-binding signaling antagonist, the cyclic undecapeptide cyclosporin A. To our knowledge, these represent the first Rgg protein X-ray crystal structures. Based on the results of extensive structure-function analysis, we reveal the peptide pheromone-binding site and the mechanism by which cyclosporin A inhibits activation of the peptide pheromone receptor. Guided by the Rgg-cyclosporin A complex structure, we predicted that the nonimmunosuppressive cyclosporin A analog valspodar would inhibit Rgg activation. Indeed, we found that, like cyclosporin A, valspodar inhibits peptide pheromone activation of conserved Rgg proteins in medically relevant Streptococcus species. Finally, the crystal structures presented here revealed that the Rgg protein DNA-binding domains are covalently linked across their dimerization interface by a disulfide bond formed by a highly conserved cysteine. The DNA-binding domain dimerization interface observed in our structures is essentially identical to the interfaces previously described for other members of the XRE DNA-binding domain family, but the presence of an intermolecular disulfide bond buried in this interface appears to be unique. We hypothesize that this disulfide bond may, under the right conditions, affect Rgg monomer-dimer equilibrium, stabilize Rgg conformation, or serve as a redox-sensitive switch.

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Section: Resultsmentioning

confidence: 99%

Rgg protein structure–function and inhibition by cyclic peptide compounds

Parashar

Aggarwal

Federle

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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100

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“…One Rap protein, RapH, both dephosphorylates Spo0F and binds to the ComA DNA binding domain, directly inhibiting its interaction with DNA and allosterically inhibiting ComA dimerization (11,21). Therefore, we explored the possibility that RapP might function to both dephosphorylate Spo0F and bind ComA, inhibiting ComA-driven srfA expression.…”

Section: Resultsmentioning

confidence: 99%

“…Despite their sequence and structural similarity to the Rap proteins that dephosphorylate Spo0F, some B. subtilis Rap proteins, such as RapC, RapF, and RapG, are transcriptional antiactivator proteins that inhibit the function of their response regulator tar-gets without dephosphorylating their receiver domains (20)(21)(22)(23). For example, RapC and RapF bind to the DNA binding domain of the response regulator ComA (20,23) and RapG binds to DegU (22).…”

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confidence: 99%

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A Plasmid-Encoded Phosphatase Regulates Bacillus subtilis Biofilm Architecture, Sporulation, and Genetic Competence

et al. 2013

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bBacillus subtilis biofilm formation is tightly regulated by elaborate signaling pathways. In contrast to domesticated lab strains of B. subtilis which form smooth, essentially featureless colonies, undomesticated strains such as NCIB 3610 form architecturally complex biofilms. NCIB 3610 also contains an 80-kb plasmid absent from laboratory strains, and mutations in a plasmid-encoded homolog of a Rap protein, RapP, caused a hyperrugose biofilm phenotype. Here we explored the role of rapP phrP in biofilm formation. We found that RapP is a phosphatase that dephosphorylates the intermediate response regulator Spo0F. RapP appears to employ a catalytic glutamate to dephosphorylate the Spo0F aspartyl phosphate, and the implications of the RapP catalytic glutamate are discussed. In addition to regulating B. subtilis biofilm formation, we found that RapP regulates sporulation and genetic competence as a result of its ability to dephosphorylate Spo0F. Interestingly, while rap phr gene cassettes routinely form regulatory pairs; i.e., the mature phr gene product inhibits the activity of the rap gene product, the phrP gene product did not inhibit RapP activity in our assays. RapP activity was, however, inhibited by PhrH in vivo but not in vitro. Additional genetic analysis suggests that RapP is directly inhibited by peptide binding. We speculate that PhrH could be subject to posttranslational modification in vivo and directly inhibit RapP activity or, more likely, PhrH upregulates the expression of a peptide that, in turn, directly binds to RapP and inhibits its Spo0F phosphatase activity.

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“…Regarding the other members of the RNPP family, both structures of RapH (24) and RapF (25) have been obtained in complex with their respective protein targets, Spo0F and ComAc, in the absence of their cognate inhibitory Phr peptides. The regulatory mechanism of the Rap proteins by Phr peptides is therefore unknown.…”

Section: Discussionmentioning

confidence: 99%

Structural basis for the activation mechanism of the PlcR virulence regulator by the quorum-sensing signal peptide PapR

Grenha

Slamti

Nicaise

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The quorum-sensing regulator PlcR is the master regulator of most known virulence factors in Bacillus cereus. It is a helix-turn-helix (HTH)-type transcription factor activated upon binding of its cognate signaling peptide PapR on a tetratricopeptide repeat-type regulatory domain. The structural and functional properties of PlcR have defined a new family of sensor regulators, called the RNPP family (for Rap, NprR, PrgX, and PlcR), in Gram-positive bacteria. To fully understand the activation mechanism of PlcR, we took a closer look at the conformation changes induced upon binding of PapR and of its target DNA, known as PlcR-box. For that purpose we have determined the structures of the apoform of PlcR (Apo PlcR) and of the ternary complex of PlcR with PapR and the PlcR-box from the plcA promoter. Comparison of the apoform of PlcR with the previously published structure of the PlcR-PapR binary complex shows how a small conformational change induced in the C-terminal region of the tetratricopeptide repeat (TPR) domain upon peptide binding propagates via the linker helix to the N-terminal HTH DNA-binding domain. Further comparison with the PlcR-PapR-DNA ternary complex shows how the activation of the PlcR dimer allows the linker helix to undergo a drastic conformational change and subsequent proper positioning of the HTH domains in the major groove of the two half sites of the pseudopalindromic PlcR-box. Together with random mutagenesis experiments and interaction measurements using peptides from distinct pherogroups, this structural analysis allows us to propose a molecular mechanism for this functional switch

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Structural Basis of Response Regulator Inhibition by a Bacterial Anti-Activator Protein

Cited by 52 publications

References 68 publications

Rgg protein structure–function and inhibition by cyclic peptide compounds

Rgg protein structure–function and inhibition by cyclic peptide compounds

A Plasmid-Encoded Phosphatase Regulates Bacillus subtilis Biofilm Architecture, Sporulation, and Genetic Competence

Structural basis for the activation mechanism of the PlcR virulence regulator by the quorum-sensing signal peptide PapR

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