Xenobiotic-sensing nuclear receptors involved in drug metabolism: a structural perspective

Abstract

Xenobiotic compounds undergo a critical range of biotransformations performed by the phase I, II, and III drug-metabolizing enzymes. The oxidation, conjugation, and transportation of potentially harmful xenobiotic and endobiotic compounds achieved by these catalytic systems are significantly regulated, at the gene expression level, by members of the nuclear receptor (NR) family of ligand-modulated transcription factors. Activation of NRs by a variety of endo- and exogenous chemicals are elemental to induction and repression of drug-metabolism pathways. The master xenobiotic sensing NRs, the promiscuous pregnane X receptor and less-promiscuous constitutive androstane receptor are crucial to initial ligand recognition, jump-starting the metabolic process. Other receptors, including farnesoid X receptor, vitamin D receptor, hepatocyte nuclear factor 4 alpha, peroxisome proliferator activated receptor, glucocorticoid receptor, liver X receptor, and RAR-related orphan receptor, are not directly linked to promiscuous xenobiotic binding, but clearly play important roles in the modulation of metabolic gene expression. Crystallographic studies of the ligand-binding domains of nine NRs involved in drug metabolism provide key insights into ligand-based and constitutive activity, coregulator recruitment, and gene regulation. Structures of other, noncanonical transcription factors also shed light on secondary, but important, pathways of control. Pharmacological targeting of some of these nuclear and atypical receptors has been instituted as a means to treat metabolic and developmental disorders and provides a future avenue to be explored for other members of the xenobiotic-sensing NRs.

Figure 1

(A–I) LBD crystal structures of the nuclear receptors, PXR, CAR, LXRα, GR, VDR, RORα, FXR, HNF4α, and PPARα. Each LBD contains the conserved, three-layered α-helical sandwich, along with structural elements that line the lower left portion of the ligand-binding pocket.

Figure 2

(A) Structural overlay of all nine NR LBD crystal structures. In blue are the conserved α-helical bundles, and colored in tan are the divergent structural features that line the binding pocket. (B) Amino acids that are found to envelop the lower left ligand-binding pocket for each NR. Identities of these regions are disparate in terms of both length and chemical property. (C) Structural alignment and sequence identity performed by Dali (Holm et al., 2008) using known LBD crystal structures reveals both primary and secondary similarities.

Figure 3

(A) Crystal structure of the PXR LBD, with the ligand-binding pocket residues highlighted in magenta. A space-filling model of the binding pocket is shown to better understand the cavity of the pocket. PXR has the unique ability to alter its pocket to allow for the binding of different chemical structures, as exemplified by the pocket volume change from ~1,300 (SR12813 bound) to ~1,600 ų (rifampicin bound). (B) Overlay of the SR12813-bound (gray/yellow) and rifampicin-bound (cyan/green) PXR LBDs. Of note is the conformational change required to accommodate the large, macrocyclic compound, rifampicin, which results in three regions in the LBD becoming disordered (yellow). (C) Chemical properties of the residues that surround the ligand-binding pocket. Amino acids are colored to represent the chemical property: orange (hydrophobic); green (polar); blue (basic); red (acidic); yellow (cysteine); and white (glycine).

Figure 4

(A) Crystal structure of the CAR LBD. Ligand-binding pocket residues are highlighted in magenta. The cavity of the binding pocket is represented as a space-filled model. The CAR pocket has a calculated volume of ~700 ų. (B) Chemical properties of the residues that surround the CAR ligand-binding pocket. Amino acids are colored to represent the chemical property: orange (hydrophobic); green (polar); blue (basic); red (acidic); yellow (cysteine); and white (glycine).

Figure 5

(A) The WT PXR LBD (cyan) with residues that would be removed in the various PXR splice variants highlighted (PXR.2, left; PXR.3, right; yellow). The deletion of these residues would still maintain the canonical LBD structure, but would also result in a smaller binding pocket, which would alter PXR's ability to bind to larger xenobiotics. (B) WT CAR LBD (cyan) with residues predicted to be deleted from CAR splice variants (but still maintaining the classical LBD structure) (yellow/green). Exon 7 is a major deletion in many of CAR's variants (left, yellow) and would affect ligand binding. Shown on the right, exon 5 (predicted, yellow), exon 9 (predicted, green), and various insertion and deletion sites (magenta) are highlighted, describing additional alterations to CAR, resulting in many of the known splice variants.

Figure 6

(A) Overlay of VDR, LXRα, and GR showing secondary structural elements that cap the left portion of the ligand-binding pocket; these NRs contain a beta-hairpin cap to enclose the cavity. (B) RORα ligand-binding pocket is capped by a beta-hairpin, as with other receptors, and an additional alpha-helical cap on top of the beta-hairpin element. (C) Binding cavity of FXR is enclosed only by an alpha-helical cap, but not by a beta-hairpin. (D) HNF4α maintains the beta-hairpin structural element and incorporates half of an alpha-helical cap. (E) The PPARα ligand-binding pocket is enclosed by a beta-hairpin element found in most NRs and a unique helix-beta-helix feature to add unique specificity to this receptor.

Figure 7

(A–G) Details of the ligand-binding pockets for the nine NR LBDs. Residues are color coded based on chemical properties: orange (hydrophobic); green (polar); blue (basic); red (acidic); yellow (cysteine); and white (glycine). For each illustration, approximate measurements of binding cavities are determined to compare their relative size and shape to each other. As further detailed in Table 2, each pocket is heavily hydrophobic in nature, with various elements of polarity and charge to facilitate ligand-binding specificity.

Figure 8

Coregulator recruitment for agonist versus antagonist binding. (A) Crystal structures of the PPARα LBD have been solved in complex with an agonist/coactivator (left) and antagonist/corepressor (right). Comparing the two structures, it is noted that the AF helix changes conformation dramatically upon antagonist binding, producing a unique site for corepressor binding. (B) RXRα LBD crystal structures complexed with agonist/coactivator (left) and antagonist/corepressor (right) elements. As with other NRs, the consequences of antagonist/ corepressor binding are evident in the conformational change of the AF helix.