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Structures of the inducer-binding domain of pentachlorophenol-degrading gene regulator PcpR from Sphingobium chlorophenolicum.

Hayes RP, Moural TW, Lewis KM, Onofrei D, Xun L, Kang C - Int J Mol Sci (2014)

Bottom Line: However, the other cavity was unique to PCP with much weaker affinity (Kd = 70 μM) and thus its significance was not clear.Neither phenol nor benzoic acid displayed any significant affinity to PcpR, indicating a role of chlorine substitution in ligand specificity.The finding concurs that PcpR uses various polychlorophenols as long as it includes 2,4,6-trichlorophenol, as inducers; whereas TcpR is only responsive to 2,4,6-trichlorophenol.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Washington State University, Pullman, WA 99164-4630, USA. robert_hayes@wsu.edu.

ABSTRACT
PcpR is a LysR-type transcription factor from Sphingobium chlorophenolicum L-1 that is responsible for the activation of several genes involved in polychlorophenol degradation. PcpR responds to several polychlorophenols in vivo. Here, we report the crystal structures of the inducer-binding domain of PcpR in the apo-form and binary complexes with pentachlorophenol (PCP) and 2,4,6-trichlorophenol (2,4,6-TCP). Both X-ray crystal structures and isothermal titration calorimetry data indicated the association of two PCP molecules per PcpR, but only one 2,4,6-TCP molecule. The hydrophobic nature and hydrogen bonds of one binding cavity allowed the tight association of both PCP (Kd = 110 nM) and 2,4,6-TCP (Kd = 22.8 nM). However, the other cavity was unique to PCP with much weaker affinity (Kd = 70 μM) and thus its significance was not clear. Neither phenol nor benzoic acid displayed any significant affinity to PcpR, indicating a role of chlorine substitution in ligand specificity. When PcpR is compared with TcpR, a LysR-type regulator controlling the expression of 2,4,6-trichlorophenol degradation in Cupriavidus necator JMP134, most of the residues constituting the two inducer-binding cavities of PcpR are different, except for their general hydrophobic nature. The finding concurs that PcpR uses various polychlorophenols as long as it includes 2,4,6-trichlorophenol, as inducers; whereas TcpR is only responsive to 2,4,6-trichlorophenol.

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Related in: MedlinePlus

Global structure of PcpR. (A) Ribbon diagram representing two PcpR dimers in the asymmetric unit; and (B) Overall global structure of PcpR monomer illustrating the two Rossmann-like domains RDI and RDII, the hinge region, and the C-terminal and N-terminal of this construct (labelled DBD←N and C respectively). This figure was generated using the PyMOL Molecular Graphics System, Version 1.3, Schrödinger, LLC (Cambridge, MA, USA).
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ijms-15-20736-f002: Global structure of PcpR. (A) Ribbon diagram representing two PcpR dimers in the asymmetric unit; and (B) Overall global structure of PcpR monomer illustrating the two Rossmann-like domains RDI and RDII, the hinge region, and the C-terminal and N-terminal of this construct (labelled DBD←N and C respectively). This figure was generated using the PyMOL Molecular Graphics System, Version 1.3, Schrödinger, LLC (Cambridge, MA, USA).

Mentions: Purified recombinant PcpR protein, which covered the substrate-binding domain (residues from 85 to the C-terminus of PcpR), was crystallized in the R3 space group with 4 molecules per asymmetric unit (Figure 2A) and solved at 2.7, 2.2 and 1.9 Å resolution for the apo-form, PCP and 2,4,6-TCP binary complexes, respectively (Table 1). The monomeric structure of the PcpR inducer-binding domain contains two nucleotide-binding (or Rossmann fold) motifs connected by a hinge region (Figure 2B). Although 4 molecules were seen in the asymmetric unit, PcpR appears to exist as dimers in solution as illustrated by multi-angle light scattering (MALS) (Figure 3) consistent with weak interaction between the two dimers in the crystal lattice. The N-, C-terminus and two loop regions (173–177 and 183–186) of the four molecules in the asymmetric unit and among three structures showed somewhat different conformations indicating their flexible nature.


Structures of the inducer-binding domain of pentachlorophenol-degrading gene regulator PcpR from Sphingobium chlorophenolicum.

Hayes RP, Moural TW, Lewis KM, Onofrei D, Xun L, Kang C - Int J Mol Sci (2014)

Global structure of PcpR. (A) Ribbon diagram representing two PcpR dimers in the asymmetric unit; and (B) Overall global structure of PcpR monomer illustrating the two Rossmann-like domains RDI and RDII, the hinge region, and the C-terminal and N-terminal of this construct (labelled DBD←N and C respectively). This figure was generated using the PyMOL Molecular Graphics System, Version 1.3, Schrödinger, LLC (Cambridge, MA, USA).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4264193&req=5

ijms-15-20736-f002: Global structure of PcpR. (A) Ribbon diagram representing two PcpR dimers in the asymmetric unit; and (B) Overall global structure of PcpR monomer illustrating the two Rossmann-like domains RDI and RDII, the hinge region, and the C-terminal and N-terminal of this construct (labelled DBD←N and C respectively). This figure was generated using the PyMOL Molecular Graphics System, Version 1.3, Schrödinger, LLC (Cambridge, MA, USA).
Mentions: Purified recombinant PcpR protein, which covered the substrate-binding domain (residues from 85 to the C-terminus of PcpR), was crystallized in the R3 space group with 4 molecules per asymmetric unit (Figure 2A) and solved at 2.7, 2.2 and 1.9 Å resolution for the apo-form, PCP and 2,4,6-TCP binary complexes, respectively (Table 1). The monomeric structure of the PcpR inducer-binding domain contains two nucleotide-binding (or Rossmann fold) motifs connected by a hinge region (Figure 2B). Although 4 molecules were seen in the asymmetric unit, PcpR appears to exist as dimers in solution as illustrated by multi-angle light scattering (MALS) (Figure 3) consistent with weak interaction between the two dimers in the crystal lattice. The N-, C-terminus and two loop regions (173–177 and 183–186) of the four molecules in the asymmetric unit and among three structures showed somewhat different conformations indicating their flexible nature.

Bottom Line: However, the other cavity was unique to PCP with much weaker affinity (Kd = 70 μM) and thus its significance was not clear.Neither phenol nor benzoic acid displayed any significant affinity to PcpR, indicating a role of chlorine substitution in ligand specificity.The finding concurs that PcpR uses various polychlorophenols as long as it includes 2,4,6-trichlorophenol, as inducers; whereas TcpR is only responsive to 2,4,6-trichlorophenol.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Washington State University, Pullman, WA 99164-4630, USA. robert_hayes@wsu.edu.

ABSTRACT
PcpR is a LysR-type transcription factor from Sphingobium chlorophenolicum L-1 that is responsible for the activation of several genes involved in polychlorophenol degradation. PcpR responds to several polychlorophenols in vivo. Here, we report the crystal structures of the inducer-binding domain of PcpR in the apo-form and binary complexes with pentachlorophenol (PCP) and 2,4,6-trichlorophenol (2,4,6-TCP). Both X-ray crystal structures and isothermal titration calorimetry data indicated the association of two PCP molecules per PcpR, but only one 2,4,6-TCP molecule. The hydrophobic nature and hydrogen bonds of one binding cavity allowed the tight association of both PCP (Kd = 110 nM) and 2,4,6-TCP (Kd = 22.8 nM). However, the other cavity was unique to PCP with much weaker affinity (Kd = 70 μM) and thus its significance was not clear. Neither phenol nor benzoic acid displayed any significant affinity to PcpR, indicating a role of chlorine substitution in ligand specificity. When PcpR is compared with TcpR, a LysR-type regulator controlling the expression of 2,4,6-trichlorophenol degradation in Cupriavidus necator JMP134, most of the residues constituting the two inducer-binding cavities of PcpR are different, except for their general hydrophobic nature. The finding concurs that PcpR uses various polychlorophenols as long as it includes 2,4,6-trichlorophenol, as inducers; whereas TcpR is only responsive to 2,4,6-trichlorophenol.

Show MeSH
Related in: MedlinePlus