A common switch in activation of the response regulators NtrC and PhoB: phosphorylation induces dimerization of the receiver modules.

Fiedler, Ulrike; Weiss, Verena

doi:10.1002/j.1460-2075.1995.tb00039.x

Cited by 118 publications

(118 citation statements)

References 51 publications

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“…In bacterial two-component systems, a sensor kinase phosphorylates an aspartic acid residue of its cognate response regulator, resulting in activation of the regulator (21). Phosphorylation of the monomeric PhoB protein induces dimerization, and the resultant dimer binds to its DNA binding sites (22,23). In addition, the phosphorylated OmpR forms dimers and trimers as evidenced by chemical cross-linking experiments (24).…”

Section: Discussionmentioning

confidence: 99%

Multimerization of Phosphorylated and Non-phosphorylated ArcA Is Necessary for the Response Regulator Function of the Arc Two-component Signal Transduction System

Jeon¹,

Lee

Han³

et al. 2001

Journal of Biological Chemistry

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To adapt to anaerobic conditions, Escherichia coli operates the Arc two-component signal transduction system, consisting of a sensor kinase, ArcB, and a response regulator, ArcA. ArcA is converted to the active form, phosphorylated ArcA (ArcA-P), by ArcB-mediated phosphorylation. The active ArcA-P binds to the promoter regions of target genes, thereby regulating their transcriptional activities. The phosphoryl group of ArcA-P is unstable with a half-life of 30 min. However, we were able to inhibit the dephosphorylation for more than 12 h by the addition of EDTA; this allowed us to characterize ArcA-P. Gel-filtration and glycerol sedimentation experiments demonstrated that ArcA exists as a homo-dimer. ArcA phosphorylated by either ArcB or carbamyl phosphate multimerizes to form a tetramer of dimers; this multimer binds to the ArcA DNA binding site. Isoelectric focusing gel electrophoresis and nitrocellulose-filter binding analyses indicated that the ArcA multimer is composed of both ArcA-P and ArcA in a ratio, 1:1. The ArcA(D54E) mutant protein was unable to be phosphorylated by ArcB. This defect resulted in the inability of ArcA(D54E) to form a multimer or to bind to the ArcA DNA binding site. These results indicate that phosphorylation of ArcA induces multimerization prior to DNA binding, and the multimerization is a prerequisite for binding. Our results suggest a novel model that phosphorylation of ArcA by ArcB regulates multimerization of ArcA, which in turn functions as a response regulator.

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Section: Discussionmentioning

confidence: 99%

Multimerization of Phosphorylated and Non-phosphorylated ArcA Is Necessary for the Response Regulator Function of the Arc Two-component Signal Transduction System

Jeon¹,

Lee

Han³

et al. 2001

Journal of Biological Chemistry

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“…Phosphorylation of response regulators is believed to prevent the activity of an inhibitory domain, allowing the amino terminus to dimerize or oligomerize and the carboxyl terminus to bind to the target DNA (27). However, like the PhoP protein of Bacillus subtilis, the PmrA protein dimerizes and binds to DNA regardless of its phosphorylation state (28).…”

Section: Discussionmentioning

confidence: 99%

Molecular Characterization of the PmrA Regulon

Wösten¹,

Groisman

1999

Journal of Biological Chemistry

101

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The two-component system PmrA/PmrB of Salmonella enterica controls expression of several loci including those mediating modifications in the lipopolysaccharide that result in polymyxin resistance. To gain insight in the regulation of polymyxin resistance, we mapped the transcription start sites of the PmrA-regulated genes pmrC, pmrG, pbgPE, and ugd and identified a conserved sequence in the promoter region of the first three genes. His-tagged PmrA protein could gel shift DNA fragments containing the promoters of the pmrC, pmrG, and pbgPE genes but not the udg promoter. DNase I footprinting analysis of the pmrC, pmrG, and pbgPE promoters indicate that phosphorylated as well as unphosphorylated PmrA bind to a 16-base pair imperfect inverted repeat sequence (5-TTAAKTTCTTA-AKGTT-3), which is found 40, 80, and 38 nucleotides upstream from the transcription start sites of the pmrC, pmrG, and pbgPE genes, respectively. Our data suggest that a PmrA dimer activates transcription of the divergent pmrG and pbgPE promoters by binding to a single site in the pmrG-pbgPE intergenic region and that the ugd gene is regulated by the PmrA/PmrB system only indirectly.

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“…[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: 86%

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

119

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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A common switch in activation of the response regulators NtrC and PhoB: phosphorylation induces dimerization of the receiver modules.

Cited by 118 publications

References 51 publications

Multimerization of Phosphorylated and Non-phosphorylated ArcA Is Necessary for the Response Regulator Function of the Arc Two-component Signal Transduction System

Multimerization of Phosphorylated and Non-phosphorylated ArcA Is Necessary for the Response Regulator Function of the Arc Two-component Signal Transduction System

Molecular Characterization of the PmrA Regulon

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

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