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Crystal structure of a functional dimer of the PhoQ sensor domain.

Cheung J, Bingman CA, Reyngold M, Hendrickson WA, Waldburger CD - J. Biol. Chem. (2008)

Bottom Line: Analysis of the wild-type structure reveals a dimer that allows for the formation of a salt bridge across the dimer interface between Arg-50' and Asp-179 and with nickel ions bound to aspartate residues in the acidic cluster.The physiological importance of the salt bridge to in vivo PhoQ function has been confirmed by mutagenesis.The mutant structure has an alternative, non-physiological dimeric association.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biophysics, Howard Hughes Medical Institute, Columbia University, New York, New York 10032, USA.

ABSTRACT
The PhoP-PhoQ two-component system is a well studied bacterial signaling system that regulates virulence and stress response. Catalytic activity of the histidine kinase sensor protein PhoQ is activated by low extracellular concentrations of divalent cations such as Mg2+, and subsequently the response regulator PhoP is activated in turn through a classic phosphotransfer pathway that is typical in such systems. The PhoQ sensor domains of enteric bacteria contain an acidic cluster of residues (EDDDDAE) that has been implicated in direct binding to divalent cations. We have determined crystal structures of the wild-type Escherichia coli PhoQ periplasmic sensor domain and of a mutant variant in which the acidic cluster was neutralized to conservative uncharged residues (QNNNNAQ). The PhoQ domain structure is similar to that of DcuS and CitA sensor domains, and this PhoQ-DcuS-CitA (PDC) sensor fold is seen to be distinct from the superficially similar PAS domain fold. Analysis of the wild-type structure reveals a dimer that allows for the formation of a salt bridge across the dimer interface between Arg-50' and Asp-179 and with nickel ions bound to aspartate residues in the acidic cluster. The physiological importance of the salt bridge to in vivo PhoQ function has been confirmed by mutagenesis. The mutant structure has an alternative, non-physiological dimeric association.

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Structure of the wild-type PhoQ 43–190 dimer in complex with nickel. A, the overall structure in complex with nickel is shown as a ribbon diagram. Nickel ions are shown as green spheres. The Arg-50′ → Asp-179 salt bridge is shown with residue side chains depicted in a stick representation (carbon colored yellow, nitrogen colored blue, and oxygen colored red). A line of black dots marks interacting atoms of the salt bridge. The Cα atoms of Gly-54 residues are shown as yellow spheres. Residues are identified by one-letter-code labels. Colored dots are drawn to represent a hypothetical path of the protein backbone through disordered regions corresponding to residues 135 and 136 of molecule A (light purple), and residues 136 and 137 of molecule B (light red). B, the structure of one subunit (molecule A) of the wild-type PhoQ 43–190 dimer is shown from a 90° rotation about the vertical axis of the dimer as shown in A. Secondary structure elements are labeled in black. The diagram was created using MolScript (43).
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fig1: Structure of the wild-type PhoQ 43–190 dimer in complex with nickel. A, the overall structure in complex with nickel is shown as a ribbon diagram. Nickel ions are shown as green spheres. The Arg-50′ → Asp-179 salt bridge is shown with residue side chains depicted in a stick representation (carbon colored yellow, nitrogen colored blue, and oxygen colored red). A line of black dots marks interacting atoms of the salt bridge. The Cα atoms of Gly-54 residues are shown as yellow spheres. Residues are identified by one-letter-code labels. Colored dots are drawn to represent a hypothetical path of the protein backbone through disordered regions corresponding to residues 135 and 136 of molecule A (light purple), and residues 136 and 137 of molecule B (light red). B, the structure of one subunit (molecule A) of the wild-type PhoQ 43–190 dimer is shown from a 90° rotation about the vertical axis of the dimer as shown in A. Secondary structure elements are labeled in black. The diagram was created using MolScript (43).

Mentions: Overall Structure of PhoQ 43–190—The structure of wild-type PhoQ 43–190 (Fig. 1) was determined by multiwavelength anomalous dispersion from a single SeMet crystal at the selenium K-edge. Two molecules were found forming an apparent dimer in the asymmetric unit in space group P21. Clear electron density allowed residues to be traced in for residues 45–134 and 137–188 of molecule A and for residues 45–75, 83–135, and 138–186 of molecule B. Missing residues correspond to N- and C-terminal segments and to loops. A total of 275 ordered residues, 184 water molecules, 2 nickel ions, and 1 acetate ion was refined against the low remote data to a resolution of 2.5 Å with R and Rfree values of 20.3% and 30.1%, respectively (Tables 1 and 2).


Crystal structure of a functional dimer of the PhoQ sensor domain.

Cheung J, Bingman CA, Reyngold M, Hendrickson WA, Waldburger CD - J. Biol. Chem. (2008)

Structure of the wild-type PhoQ 43–190 dimer in complex with nickel. A, the overall structure in complex with nickel is shown as a ribbon diagram. Nickel ions are shown as green spheres. The Arg-50′ → Asp-179 salt bridge is shown with residue side chains depicted in a stick representation (carbon colored yellow, nitrogen colored blue, and oxygen colored red). A line of black dots marks interacting atoms of the salt bridge. The Cα atoms of Gly-54 residues are shown as yellow spheres. Residues are identified by one-letter-code labels. Colored dots are drawn to represent a hypothetical path of the protein backbone through disordered regions corresponding to residues 135 and 136 of molecule A (light purple), and residues 136 and 137 of molecule B (light red). B, the structure of one subunit (molecule A) of the wild-type PhoQ 43–190 dimer is shown from a 90° rotation about the vertical axis of the dimer as shown in A. Secondary structure elements are labeled in black. The diagram was created using MolScript (43).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2376233&req=5

fig1: Structure of the wild-type PhoQ 43–190 dimer in complex with nickel. A, the overall structure in complex with nickel is shown as a ribbon diagram. Nickel ions are shown as green spheres. The Arg-50′ → Asp-179 salt bridge is shown with residue side chains depicted in a stick representation (carbon colored yellow, nitrogen colored blue, and oxygen colored red). A line of black dots marks interacting atoms of the salt bridge. The Cα atoms of Gly-54 residues are shown as yellow spheres. Residues are identified by one-letter-code labels. Colored dots are drawn to represent a hypothetical path of the protein backbone through disordered regions corresponding to residues 135 and 136 of molecule A (light purple), and residues 136 and 137 of molecule B (light red). B, the structure of one subunit (molecule A) of the wild-type PhoQ 43–190 dimer is shown from a 90° rotation about the vertical axis of the dimer as shown in A. Secondary structure elements are labeled in black. The diagram was created using MolScript (43).
Mentions: Overall Structure of PhoQ 43–190—The structure of wild-type PhoQ 43–190 (Fig. 1) was determined by multiwavelength anomalous dispersion from a single SeMet crystal at the selenium K-edge. Two molecules were found forming an apparent dimer in the asymmetric unit in space group P21. Clear electron density allowed residues to be traced in for residues 45–134 and 137–188 of molecule A and for residues 45–75, 83–135, and 138–186 of molecule B. Missing residues correspond to N- and C-terminal segments and to loops. A total of 275 ordered residues, 184 water molecules, 2 nickel ions, and 1 acetate ion was refined against the low remote data to a resolution of 2.5 Å with R and Rfree values of 20.3% and 30.1%, respectively (Tables 1 and 2).

Bottom Line: Analysis of the wild-type structure reveals a dimer that allows for the formation of a salt bridge across the dimer interface between Arg-50' and Asp-179 and with nickel ions bound to aspartate residues in the acidic cluster.The physiological importance of the salt bridge to in vivo PhoQ function has been confirmed by mutagenesis.The mutant structure has an alternative, non-physiological dimeric association.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biophysics, Howard Hughes Medical Institute, Columbia University, New York, New York 10032, USA.

ABSTRACT
The PhoP-PhoQ two-component system is a well studied bacterial signaling system that regulates virulence and stress response. Catalytic activity of the histidine kinase sensor protein PhoQ is activated by low extracellular concentrations of divalent cations such as Mg2+, and subsequently the response regulator PhoP is activated in turn through a classic phosphotransfer pathway that is typical in such systems. The PhoQ sensor domains of enteric bacteria contain an acidic cluster of residues (EDDDDAE) that has been implicated in direct binding to divalent cations. We have determined crystal structures of the wild-type Escherichia coli PhoQ periplasmic sensor domain and of a mutant variant in which the acidic cluster was neutralized to conservative uncharged residues (QNNNNAQ). The PhoQ domain structure is similar to that of DcuS and CitA sensor domains, and this PhoQ-DcuS-CitA (PDC) sensor fold is seen to be distinct from the superficially similar PAS domain fold. Analysis of the wild-type structure reveals a dimer that allows for the formation of a salt bridge across the dimer interface between Arg-50' and Asp-179 and with nickel ions bound to aspartate residues in the acidic cluster. The physiological importance of the salt bridge to in vivo PhoQ function has been confirmed by mutagenesis. The mutant structure has an alternative, non-physiological dimeric association.

Show MeSH
Related in: MedlinePlus