Structural Basis of Response Regulator Dephosphorylation by Rap Phosphatases

Parashar, Vijay; Mirouze, Nicolas; Dubnau, David; Neiditch, Matthew B.

doi:10.1371/journal.pbio.1000589

Cited by 92 publications

(186 citation statements)

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“…In previous studies, the concave surface of the repeat domain of peptide pheromone receptors was identified as the pheromone-binding site (Fig. S5 A-D) (13,14,(16)(17)(18). We hypothesized that the Streptococcus Rgg receptors similarly used the concave surface of their repeat domain as the SHP-binding site.…”

Section: Resultsmentioning

confidence: 92%

“…A search (30) of the PDB database for proteins structurally similar to Rgg2 Sd identified PrgX (PDB ID code 2AXZ, Z score = 19.2) (18), PlcR (PDB ID code 3U3W, Z score = 13.1) (15), NprR (PDB ID code 4GPK, Z score = 11.7) (19), and RapI (PDB ID code 4I1A, Z score = 10.7) (17). In previous studies, the concave surface of the repeat domain of peptide pheromone receptors was identified as the pheromone-binding site (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The Rap proteins are phosphatases and transcriptional antiactivators, whereas NprR, PlcR, and PrgX are DNA-binding transcription factors. Structure-function studies revealed that Rap, NprR, PlcR, and PrgX (the RNPP family proteins) use a structurally similar C-terminal tetratricopeptide (TPR)-like repeat domain to bind their cognate peptide pheromones (12)(13)(14)(15)(16)(17)(18)(19).…”

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confidence: 99%

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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: 92%

Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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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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“…In the structure, the side chain of RapH Gln-47 inserts into the Spo0F active site. RapH Gln-47 may orient water for direct in-line hydrophilic attack on the Spo0F phosphoaspartate 54 phosphorous atom and thus cause hydrolysis of the phosphoryl group (54). In these cases, the interactions are primarily responsible for the specific histidine kinase-response regulator or connector-response regulator binding.…”

Section: Discussionmentioning

confidence: 99%

Structural Basis of a Physical Blockage Mechanism for the Interaction of Response Regulator PmrA with Connector Protein PmrD from Klebsiella pneumoniae

Luo

Lou

Rajasekaran

et al. 2013

Journal of Biological Chemistry

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Background: PmrD binds to phospho-PmrA and sustains its phosphorylation state. Results: Phospho-PmrA interacts with PmrD via several specific intermolecular interactions. Conclusion: A steric inhibition mechanism was proposed for protecting phospho-PmrA against dephosphorylation. Significance: This work provides novel data revealing how a connector protein protects an activated response regulator.

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“…1) (8,9). In addition to RapA and RapB, many other Rap proteins, including RapE, RapH, and RapJ, function similarly to dephosphorylate Spo0F (10)(11)(12). The ultimate effect of dephosphorylating Spo0F is a decrease in the cellular concentration of phosphorylated Spo0A (Spo0AϳP).…”

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confidence: 99%

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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Structural Basis of Response Regulator Dephosphorylation by Rap Phosphatases

Abstract: Crystallographic, biochemical, and genetic studies reveal the mechanism of Rap protein phosphatase activity within the phosphorelay pathway leading to sporulation in Bacillus species.

Cited by 92 publications

References 50 publications

Rgg protein structure–function and inhibition by cyclic peptide compounds

Rgg protein structure–function and inhibition by cyclic peptide compounds

Structural Basis of a Physical Blockage Mechanism for the Interaction of Response Regulator PmrA with Connector Protein PmrD from Klebsiella pneumoniae

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

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