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Title: Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride

Abstract

The response regulator PhoP is part of the PhoQ/PhoP two-component system involved in responses to depletion of extracellular Mg{sup 2+}. Here, we report the crystal structures of the receiver domain of Escherichia coli PhoP determined in the absence and presence of the phosphoryl analog beryllofluoride. In the presence of beryllofluoride, the active receiver domain forms a twofold symmetric dimer similar to that seen in structures of other regulatory domains from the OmpR/PhoB family, providing further evidence that members of this family utilize a common mode of dimerization in the active state. In the absence of activating agents, the PhoP receiver domain crystallizes with a similar structure, consistent with the previous observation that high concentrations can promote an active state of PhoP independent of phosphorylation.

Authors:
;
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL) National Synchrotron Light Source
Sponsoring Org.:
Doe - Office Of Science
OSTI Identifier:
929980
Report Number(s):
BNL-80587-2008-JA
Journal ID: ISSN 0021-9193; JOBAAY; TRN: US200822%%1137
DOE Contract Number:
DE-AC02-98CH10886
Resource Type:
Journal Article
Resource Relation:
Journal Name: Journal of Bacteriology; Journal Volume: 189; Journal Issue: 16
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; CRYSTAL STRUCTURE; DIMERIZATION; DIMERS; ESCHERICHIA COLI; PHOSPHORYLATION; national synchrotron light source

Citation Formats

Bachhawat,P., and Stock, A. Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride. United States: N. p., 2007. Web. doi:10.1128/JB.00049-07.
Bachhawat,P., & Stock, A. Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride. United States. doi:10.1128/JB.00049-07.
Bachhawat,P., and Stock, A. Mon . "Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride". United States. doi:10.1128/JB.00049-07.
@article{osti_929980,
title = {Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride},
author = {Bachhawat,P. and Stock, A.},
abstractNote = {The response regulator PhoP is part of the PhoQ/PhoP two-component system involved in responses to depletion of extracellular Mg{sup 2+}. Here, we report the crystal structures of the receiver domain of Escherichia coli PhoP determined in the absence and presence of the phosphoryl analog beryllofluoride. In the presence of beryllofluoride, the active receiver domain forms a twofold symmetric dimer similar to that seen in structures of other regulatory domains from the OmpR/PhoB family, providing further evidence that members of this family utilize a common mode of dimerization in the active state. In the absence of activating agents, the PhoP receiver domain crystallizes with a similar structure, consistent with the previous observation that high concentrations can promote an active state of PhoP independent of phosphorylation.},
doi = {10.1128/JB.00049-07},
journal = {Journal of Bacteriology},
number = 16,
volume = 189,
place = {United States},
year = {Mon Jan 01 00:00:00 EST 2007},
month = {Mon Jan 01 00:00:00 EST 2007}
}
  • The crystal structure of the yeast SLN1 response regulator (RR) domain bound to both a phosphoryl analog [beryllium fluoride (BeF3 -)] and Mg2 +, in complex with its downstream phosphorelay signaling partner YPD1, has been determined at a resolution of 1.70 Angstroms. Comparisons between the BeF3 --activated complex and the unliganded (or apo) complex determined previously reveal modest but important differences. The SLN1-R1{center_dot}Mg2 +{center_dot}BeF3 - structure from the complex provides evidence for the first time that the mechanism of phosphorylation-induced activation is highly conserved between bacterial RR domains and this example from a eukaryotic organism. Residues in and around themore » active site undergo slight rearrangements in order to form bonds with the essential divalent cation and fluorine atoms of BeF3 -. Two conserved switch-like residues (Thr1173 and Phe1192) occupy distinctly different positions in the apo versus BeF3 --bound structures, consistent with the 'Y-T' coupling mechanism proposed for the activation of CheY and other bacterial RRs. Several loop regions and the a4-{beta}5-a5 surface of the SLN1-R1 domain undergo subtle conformational changes ({approx} 1-3 Angstroms displacements relative to the apo structure) that lead to significant changes in terms of contacts that are formed with YPD1. Detailed structural comparisons of protein-protein interactions in the apo and BeF3 --bound complexes suggest at least a two-state equilibrium model for the formation of a transient encounter complex, in which phosphorylation of the RR promotes the formation of a phosphotransfer-competent complex. In the BeF3 --activated complex, the position of His64 from YPD1 needs to be within ideal distance of and in near-linear geometry with Asp1144 from the SLN1-R1 domain for phosphotransfer to occur. The ground-state structure presented here suggests that phosphoryl transfer will likely proceed through an associative mechanism involving the formation of a pentacoordinate phosphorus intermediate.« less
  • The Escherichia coli Rsd protein binds tightly and specifically to the RNA polymerase (RNAP) {sigma}{sup 70} factor. Rsd plays a role in alternative {sigma} factor-dependent transcription by biasing the competition between {sigma}{sup 70} and alternative {sigma} factors for the available core RNAP. Here, we determined the 2.6 {angstrom}-resolution X-ray crystal structure of Rsd bound to {sigma}{sup 70} domain 4 ({sigma}{sup 70}{sub 4}), the primary determinant for Rsd binding within {sigma}{sup 70}. The structure reveals that Rsd binding interferes with the two primary functions of {sigma}{sup 70}{sub 4}, core RNAP binding and promoter -35 element binding. Interestingly, the most highly conservedmore » Rsd residues form a network of interactions through the middle of the Rsd structure that connect the {sigma}{sup 70}{sub 4}-binding surface with three cavities exposed on distant surfaces of Rsd, suggesting functional coupling between {sigma}{sup 70}{sub 4} binding and other binding surfaces of Rsd, either for other proteins or for as yet unknown small molecule effectors. These results provide a structural basis for understanding the role of Rsd, as well as its ortholog, AlgQ, a positive regulator of Pseudomonas aeruginosa virulence, in transcription regulation.« less
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