Evidence of Intradomain and Interdomain Flexibility in an OmpR/PhoB Homolog from Thermotoga maritima

Buckler, David R.; Zhou, Yuchen; Stock, Ann

doi:10.1016/s0969-2126(01)00706-7

Cited by 102 publications

(117 citation statements)

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“…These insertions presumably activated WspR via disruption of the interface between the two domains, a mechanism common to many responseregulator activation mechanisms (Eldridge et al, 2002;Park et al, 2002;Muchova et al, 2004). A comparison of the domain interaction surfaces of four full-length, two-domain response-regulator crystal structures (Robinson et al, 2003) suggests that the activation mechanisms of the different proteins, NarL (Baikalov et al, 1996), CheB (Djordjevic et al, 1998), DrrD (Buckler et al, 2002) and DrrB (Robinson et al, 2003), vary with the extent of their interdomain interfaces. For example, comparison of the DrrB and DrrD crystal structures suggests that different levels of intramolecular communication between the N-and C-terminal domains lead to significant differences in effector-domain regulation in each case (Robinson et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

et al. 2007

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The GGDEF response regulator WspR couples the chemosensory Wsp pathway to the overproduction of acetylated cellulose and cell attachment in the Pseudomonas fluorescens SBW25 wrinkly spreader (WS) genotype. Here, it is shown that WspR is a diguanylate cyclase (DGC), and that DGC activity is elevated in the WS genotype compared to that in the ancestral smooth (SM) genotype. A structure-function analysis of 120 wspR mutant alleles was employed to gain insight into the regulation and activity of WspR. Firstly, 44 random and defined pentapeptide insertions were produced in WspR, and the effects determined using assays based on colony morphology, attachment to surfaces and cellulose production. The effects of mutations within WspR were interpreted using a homology model, based on the crystal structure of Caulobacter crescentus PleD. Mutational analyses indicated that WspR activation occurs as a result of disruption of the interdomain interface, leading to the release of effector-domain repression by the N-terminal receiver domain. Quantification of attachment and cellulose production raised significant questions concerning the mechanisms of WspR function. The conserved RYGGEEF motif of WspR was also subjected to mutational analysis, and 76 single amino acid residue substitutions were tested for their effects on WspR function. The RYGGEEF motif of WspR is functionally conserved, with almost every mutation abolishing function.

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Section: Discussionmentioning

confidence: 99%

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

et al. 2007

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“…This has been supported by subsequent biochemical and biophysical studies. [29][30][31][32] In contrast, the crystal structures of two full-length RRs of the OmpR/PhoB subfamily from Thermotoga maritima, DrrD 33 and DrrB, 34 showed their recognition helices to be completely exposed, making them readily available for DNA binding. The structural data, together with several biochemical studies that point to the significance of dimerization for members of the OmpR/PhoB subfamily, [34][35][36][37][38] are suggestive of a different mechanism for transcriptional regulation.…”

Section: Introductionmentioning

confidence: 95%

“…A multiple sequence alignment of the regulatory domains of all members of this subfamily from E. coli, 20 in addition to subfamily members from other organisms for which structures of the regulatory domain are available, 33,34,49,50 revealed that the residues involved in key dimer interface interactions are highly conserved within this subfamily (Fig. 5).…”

Section: Dimer Interfacementioning

confidence: 99%

“…Interestingly, in the NtrC/ DctD subfamily, which lacks conservation in the α4-β5-α5 face, different members have been shown to use distinctly different surfaces of the regulatory domain for multimerization. 61 Another notable feature of the OmpR/PhoB subfamily is that in the inactive state the DNA recognition helix of the effector domain is freely exposed to the solvent 33,34 making it readily available for DNA binding in contrast to other RRs in which the functional regions of the effector domains are occluded by the unphosphorylated regulatory domains. 27,28 Taking into account these marked differences, it is hypothesized that members of the OmpR/PhoB subfamily use a common mechanism of regulation by dimerization.…”

Section: A Common Mechanism Of Regulation By Dimerizationmentioning

confidence: 99%

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Structural Analysis and Solution Studies of the Activated Regulatory Domain of the Response Regulator ArcA: A Symmetric Dimer Mediated by the α4-β5-α5 Face

Toro-Roman

Mack²,

Stock

2005

Journal of Molecular Biology

116

137

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SUMMARYEscherichia coli react to changes from aerobic to anaerobic conditions of growth using the ArcAArcB two-component signal transduction system. This system, in conjunction with other proteins, regulates the respiratory metabolic pathways in the organism. ArcA is a member of the OmpR/ PhoB subfamily of response regulator transcription factors that are known to regulate transcription by binding in tandem to target DNA direct repeats. It is still unclear in this subfamily how activation by phosphorylation of the regulatory domain of response regulators stimulates DNA binding by the effector domain and how dimerization and domain orientation, as well as intra-and intermolecular interactions, affect this process. In order to address these questions we have solved the crystal structure of the regulatory domain of ArcA in the presence and absence of the phosphoryl analog, BeF 3 − . In the crystal structures, the regulatory domain of ArcA forms a symmetric dimer mediated by the α4-β5-α5 face of the protein and involving a number of residues that are highly conserved in the OmpR/PhoB subfamily. It is hypothesized that members of this subfamily use a common mechanism of regulation by dimerization. Additional biophysical studies were employed to probe the oligomerization state of ArcA, as well as its individual domains, in solution. The solution studies show the propensity of the individual domains to associate into oligomers larger than the dimer observed for the intact protein, and suggest that the C-terminal DNA-binding domain also plays a role in oligomerization.

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“…For example, if the N-terminal β-sheet of the DNA-binding domain prefers to interact with the N-terminal regulatory domain, the tandem association in the hexamer would not be formed, and the recognition helix would be exposed for productive DNA -binding in the presence of the N-terminal domain. In the structures of the DrrD (25) and DrrB (24) of T. maritima, the N-terminal β-sheet of the DNA-binding domain is found to interact with the N-terminal domain. Ribbon diagram of the structure of the C-terminal domain of MTB PhoP.…”

Section: Possible Dna-binding Mechanism and Residues Involvedmentioning

confidence: 99%

Structure of the DNA-Binding Domain of the Response Regulator PhoP from Mycobacterium tuberculosis^,

Wang¹,

Engohang‐Ndong²,

Smith³

2007

Biochemistry

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The PhoP-PhoR two-component signaling system from Mycobacterium tuberculosis is essential for the virulence of the tubercle bacillus. The response regulator, PhoP, regulates expression of over 110 genes. In order to elucidate the regulatory mechanism of PhoP, we determined the crystal structure of its DNA-binding domain (PhoPC). PhoPC exhibits a typical fold of the winged helix-turn-helix subfamily of response regulators. The structure starts with a four-stranded anti-parallel β-sheet, followed by a three-helical bundle of α-helices, and then a C-terminal β-hairpin, which together with a short β-strand between the first and second helices forms a three-stranded anti-parallel β-sheet. Structural elements are packed through a hydrophobic core, with the first helix providing a scaffold for the rest of the domain to pack. The second and third helices and the long, flexible loop between them form the helix-turn-helix motif, with the third helix being the recognition helix. The C-terminal β-hairpin turn forms the wing motif. The molecular surfaces around the recognition helix and the wing residues show strong positive electrostatic potential, consistent with their roles in DNA binding and nucleotide sequence recognition. The crystal packing of PhoPC gives a hexamer ring, with neighboring molecules interacting in a head-to-tail fashion. This packing interface suggests that PhoPC could bind DNA in a tandem association. However, this mode of DNA binding is likely to be non-specific because the recognition helix is partially blocked and would be prevented from inserting into the major groove of DNA. Detailed structural analysis and implications with respect to DNA binding are discussed.Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis, is one of the most successful human pathogens. MTB infects nearly one-third of the world's population and kills over 2 million people annually worldwide. Multi-drug resistant (MDR) and extreme drug resistant (XDR) strains of MTB are on the rise in the clinic (1), stressing the urgent need to develop novel anti-tuberculosis drugs. Two-component systems (TCS) are major signaling systems in bacteria that mediate a variety of processes such as sporulation, transformation competence, membrane transport, inorganic nutrient uptake, chemotaxis, stress response, and virulence. In general, a TCS consists of a histidine kinase, which senses environmental signals, and a response regulator, which is † This work was supported by NIH Grants GM079185 to S.W. and AI65987 to I.S. J.E.-N. was supported by a Heiser Program Fellowship from the New York Community Trust. Coordinates and observed structure factor amplitudes have been deposited in the Protein Data Bank (pdb code 2PMU). phosphorylated by the cognate histidine kinase and in most cases functions as a transcription regulator to regulate expression of certain genes. Because they are absent in mammals, TCSs are potential targets for developing novel drugs. The MTB genome encodes 30 TCS proteins, comprising 11 systems in which ...

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Evidence of Intradomain and Interdomain Flexibility in an OmpR/PhoB Homolog from Thermotoga maritima

Cited by 102 publications

References 66 publications

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

Structural Analysis and Solution Studies of the Activated Regulatory Domain of the Response Regulator ArcA: A Symmetric Dimer Mediated by the α4-β5-α5 Face

Structure of the DNA-Binding Domain of the Response Regulator PhoP from Mycobacterium tuberculosis^,

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Evidence of Intradomain and Interdomain Flexibility in an OmpR/PhoB Homolog from Thermotoga maritima

Cited by 102 publications

References 66 publications

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

The structure–function relationship of WspR, a Pseudomonas fluorescens response regulator with a GGDEF output domain

Structural Analysis and Solution Studies of the Activated Regulatory Domain of the Response Regulator ArcA: A Symmetric Dimer Mediated by the α4-β5-α5 Face

Structure of the DNA-Binding Domain of the Response Regulator PhoP from Mycobacterium tuberculosis,

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Structure of the DNA-Binding Domain of the Response Regulator PhoP from Mycobacterium tuberculosis^,