Structural Disorder in the Complex of Human Pregnane X Receptor and the Macrolide Antibiotic Rifampicin

Abstract

The human nuclear xenobiotic receptor, pregnane X receptor (PXR), detects a variety of structurally distinct endogenous and xenobiotic compounds and controls expression of genes central to drug and cholesterol metabolism. The macrolide antibiotic rifampicin, a front-line treatment for tuberculosis, is an established PXR agonist and, at 823 Da, is one of the largest known ligands for the receptor. We present the 2.8 A crystal structure of the ligand-binding domain of human PXR in complex with rifampicin. We also use structural and mutagenesis data to examine the origins of the directed promiscuity exhibited by the PXRs across species. Three structurally flexible loops adjacent to the ligand-binding pocket of PXR are disordered in this crystal structure, including the 200-210 region that is part of a sequence insert novel to the promiscuous PXRs relative to other members of the nuclear receptor superfamily. The 4-methyl-1-piperazinyl ring of rifampicin, which would lie adjacent to the disordered protein regions, is also disordered and not observed in the structure. Taken together, our results indicate that one wall of the PXR ligand-binding cavity can remain flexible even when the receptor is in complex with an activating ligand. These observations highlight the key role that structural flexibility plays in PXR's promiscuous response to xenobiotics.

The statin class of HMG-CoA reductase inhibitors demonstrate differential activation of the nuclear receptors PXR, CAR and FXR, as well as their downstream target genes

“…This product was cloned into the pTriEx1.1 expression plasmid (Clontech, UK) and sequenced to ensure 100% identity to the published sequence, In silico analysis: Ligand docking was undertaken using Autodock 4 suite and Autodock Tools for Windows (Morris et al 2009). Molecular coordinates for the crystal structure of PXR with rifampicin bound (Chrencik et al 2005) were obtained from the RCSB protein databank (Accession: 1SKX; http://www.rcsb.org/pdb/home/home.do), whereas three dimensional structures of statins were designed using Corina (http://www.molecular-networks.com/products/corina). Docking was undertaken using a 0.375Å grid centred upon the ligand-binding pocket of PXR, using a genetic algortim with the following parameters: 150 iterations; 100 population size; 1E+6 energy evaluations; 1E+5 maximum generations.…”

Milk Thistle’s Active Components Silybin and Isosilybin: Novel Inhibitors of PXR-Mediated CYP3A4 Induction

“…Docking of hyperforin was started from its X-ray position in 1M13. Subsequently, docking of rifampicin was started after superimposing the hPXR-rifampicin complex 1SKX (Chrencik et al, 2005) onto 1M13. The PXR antagonist A-792611 was built from the similar HIV-protease inhibitor in Protein Data Bank ID 3GGV (Degoey et al, 2009), guided by the structure drawing from Healan-Greenberg et al (2008).…”

Identification and Validation of Novel Human Pregnane X Receptor Activators among Prescribed Drugs via Ligand-Based Virtual Screening

“…For example, coadministration of rifampicin, a hPXR activator used for treatment of tuberculosis (Chrencik et al, 2005) with a variety of drugs [including oral contraceptives (Ma et al, 2008), the anesthetic midazolam (Backman et al, 1996), and HIV protease inhibitors (Ivanovic et al, 2008)], resulted in decreased drug efficacy mainly due to hPXR-mediated increased expression of CYP3A4 (Ivanovic et al, 2008;Ma et al, 2008;di Masi et al, 2009). Thus, identification of novel hPXR activators among commercial drugs is important in predicting hPXR-mediated drug-drug interactions.…”

“…Crystal structures of the hPXR ligand-binding domain (LBD) indicate that its binding cavity is much larger than that of other NR family members (Xu et al, 2004;Chrencik et al, 2005;di Masi et al, 2009). Several key amino acid residues are responsible for the high flexibility of its binding site that is critical for recognizing promiscuous ligands of various dimensions and chemical properties (Ekins et al, 2009).…”