Expression, crystallization and preliminary crystallographic analysis of the PAS domain of RsbP, a stress-response phosphatase from<i>Bacillus subtilis</i>

Makino, Masatomo; Kondo, Shinpei; Kaneko, Tomonori; Baba, Seiki; Hirata, Kunio; Kumasaka, Takashi

doi:10.1107/s1744309109014158

Cited by 3 publications

(3 citation statements)

References 15 publications

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“…Preliminary crystallization of the RsbP-PAS domain has been reported [25] but the structure is not yet available. Therefore, the secondary structure assignments included in Fig.…”

Section: Resultsmentioning

confidence: 99%

An α/β Hydrolase and Associated Per-ARNT-Sim Domain Comprise a Bipartite Sensing Module Coupled with Diverse Output Domains

2011

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The RsbQ α/β hydrolase and RsbP serine phosphatase form a signaling pair required to activate the general stress factor σB of Bacillus subtilis in response to energy limitation. RsbP has a predicted N-terminal Per-ARNT-Sim (PAS) domain, a central coiled-coil, and a C-terminal protein phosphatase M (PPM) domain. Previous studies support a model in which RsbQ provides an activity needed for PAS to regulate the phosphatase domain via the coiled-coil. RsbQ and the PAS domain (RsbP-PAS) therefore appear to form a sensory module. Here we test this hypothesis using bioinformatic and genetic analysis. We found 45 RsbQ and RsbP-PAS homologues encoded by adjacent genes in diverse bacteria, with PAS and a predicted coiled-coil fused to one of three output domains: PPM phosphatase (Gram positive bacteria), histidine protein kinase (Gram negative bacteria), and diguanylate cyclase (both lineages). Multiple alignment of the RsbP-PAS homologues suggested nine residues that distinguish the class. Alanine substitutions at four of these conferred a null phenotype in B. subtilis, indicating their functional importance. The F55A null substitution lay in the Fα helix of an RsbP-PAS model. F55A inhibited interaction of RsbP with RsbQ in yeast two-hybrid and pull-down assays but did not significantly affect interaction of RsbP with itself. We propose that RsbQ directly contacts the PAS domains of an RsbP oligomer to provide the activating signal, which is propagated to the phosphatase domains via the coiled-coil. A similar mechanism would allow the RsbQ-PAS module to convey a common input signal to structurally diverse output domains, controlling a variety of physiological responses.

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“…Preliminary crystallization of the RsbP-PAS domain has been reported [25] but the structure is not yet available. Therefore, the secondary structure assignments included in Fig.…”

Section: Resultsmentioning

confidence: 99%

An α/β Hydrolase and Associated Per-ARNT-Sim Domain Comprise a Bipartite Sensing Module Coupled with Diverse Output Domains

2011

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“…Finally the PAS domain of RsbP from B. subtilis has recently been crystallized (Makino et al, 2009) but no structural data are yet available. It will be very interesting to establish whether that PAS domain also binds FAs.…”

Section: Discussionmentioning

confidence: 99%

Structural Characterization of the Multidomain Regulatory Protein Rv1364c from Mycobacterium tuberculosis

et al. 2011

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The open reading frame rv1364c of Mycobacterium tuberculosis, which regulates the stress-dependent σ factor, σ(F), has been analyzed structurally and functionally. Rv1364c contains domains with sequence similarity to the RsbP/RsbW/RsbV regulatory system of the stress-response σ factor of Bacillus subtilis. Rv1364c contains, sequentially, a PAS domain (which shows sequence similarity to the PAS domain of the B. subtilis RsbP protein), an active phosphatase domain, a kinase (anti-σ(F) like) domain and a C-terminal anti-σ(F) antagonist like domain. The crystal structures of two PAS domain constructs (at 2.3 and 1.6 Å) and a phosphatase/kinase dual domain construct (at 2.6 Å) are described. The PAS domain is shown to bind palmitic acid but to have 100 times greater affinity for palmitoleic acid. The full-length protein can exist in solution as both monomer and dimer. We speculate that a switch between monomer and dimer, possibly resulting from fatty acid binding, affects the accessibility of the serine of the C-terminal, anti-σ(F) antagonist domain for dephosphorylation by the phosphatase domain thus indirectly altering the availability of σ(F).

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“…However, until the hypothesized ligand is identified, this model remains speculative. The crystallization of the PAS domain of RsbP has been reported, but no structure or cofactor has yet been identified . Interestingly, a very similar regulatory network of the GSR exists in Listeria monocytogenes .…”

Section: Rsbp‐mediated Red‐light Activation Of the Energy‐branch Of Tmentioning

confidence: 99%

Activation of the General Stress Response of Bacillus subtilis by Visible Light

Steen

Hellingwerf

2015

Photochem & Photobiology

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A key challenge for microbiology is to understand how evolution has shaped the wiring of regulatory networks. This is amplified by the paucity of information of power-spectra of physicochemical stimuli to which microorganisms are exposed. Future studies of genome evolution, driven by altered stimulus regimes, will therefore require a versatile signal transduction system that allows accurate signal dosing. Here, we review the general stress response of Bacillus subtilis, and its upstream signal transduction network, as a candidate system. It can be activated by red and blue light, and by many additional stimuli. Signal integration therefore is an intricate function of this system. The blue-light response is elicited via the photoreceptor YtvA, which forms an integral part of stressosomes, to activate expression of the stress regulon of B. subtilis. Signal transfer through this network can be assayed with reporter enzymes, while intermediate steps can be studied with live-cell imaging of fluorescently tagged proteins. Different parts of this system have been studied in vitro, such that its computational modeling has made significant progress. One can directly relate the microscopic characteristics of YtvA with activation of the general stress regulon, making this system a very well-suited system for network evolution studies.

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Expression, crystallization and preliminary crystallographic analysis of the PAS domain of RsbP, a stress-response phosphatase fromBacillus subtilis

Cited by 3 publications

References 15 publications

An α/β Hydrolase and Associated Per-ARNT-Sim Domain Comprise a Bipartite Sensing Module Coupled with Diverse Output Domains

An α/β Hydrolase and Associated Per-ARNT-Sim Domain Comprise a Bipartite Sensing Module Coupled with Diverse Output Domains

Structural Characterization of the Multidomain Regulatory Protein Rv1364c from Mycobacterium tuberculosis

Activation of the General Stress Response of Bacillus subtilis by Visible Light

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