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

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 ?M) 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.

SUBMITTER: Hayes RP

PROVIDER: S-EPMC4264193 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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

Hayes Robert P RP Moural Timothy W TW Lewis Kevin M KM Onofrei David D Xun Luying L Kang ChulHee C

International journal of molecular sciences 20141112 11

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 ass ...[more]

PMID: 25397598

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Project description:Pentachlorophenol (PCP) is a highly toxic pesticide that was first introduced in the 1930s. The alphaproteobacterium Sphingobium chlorophenolicum, which was isolated from PCP-contaminated sediment, has assembled a metabolic pathway capable of completely degrading PCP. This pathway produces four toxic intermediates, including a chlorinated benzoquinone that is a potent alkylating agent and three chlorinated hydroquinones that react with O2 to produce reactive oxygen species (ROS). RNA-seq analysis revealed that PCP causes a global stress response that resembles responses to proton motive force uncoupling and membrane disruption, while surprisingly, little of the response resembles the responses expected to be produced by the PCP degradation intermediates. Tn-seq was used to identify genes important for fitness in the presence of PCP. By comparing the genes that are important for fitness in wild-type S. chlorophenolicum and a non-PCP-degrading mutant, we identified genes that are important only when the PCP degradation intermediates are produced. These include genes encoding two enzymes that are likely to be involved in protection against ROS. In addition to these enzymes, the endogenous levels of other enzymes that protect cells from oxidative stress appear to mitigate the toxic effects of the chlorinated benzoquinone and hydroquinone metabolites of PCP. The combination of RNA-seq and Tn-seq results identify important mechanisms for defense against the toxicity of PCP. IMPORTANCE Phenolic compounds such as pentachlorophenol (PCP), triclosan, and 2,4-dichlorophenoxyacetic acid (2,4-D) represent a common class of anthropogenic biocides. Despite the novelty of these compounds, many can be degraded by microbes isolated from contaminated sites. However, degradation of this class of chemicals often generates toxic intermediates, which may contribute to their recalcitrance to biodegradation. We have addressed the stresses associated with degradation of PCP by Sphingobium chlorophenolicum by examining the transcriptional response after PCP exposure and identifying genes necessary for growth during both exposure to and degradation of PCP. This work identifies some of the mechanisms that protect cells from this toxic compound and facilitate its degradation. This information could be used to engineer strains capable of improved biodegradation of PCP or similar phenolic pollutants.

Dataset Information

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

Publications

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

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