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A novel photoactive benzoic acid derivative, 4-azido-2-hydroxybenzoic acid (4-AzHBA), is synthesized and employed as a photoprobe to identify the phenol binding site of UDP-glucuronosyltransferases (UGTs). The combination of photoaffinity labeling, enzymatic digestion, MALDI-TOF (matrix-assisted laser desorption/ioninzation time-of-flight) and LC-MS mass spectrometry are used to determine that Phe(super 90) and Phe(super 93) are directly involved in the catalytic activity of UGT1A10 toward 4-MU (4-methylubelliferone) and pNP (p-nitrophenol).

Through a multiplex promoter spanning 218 kb, the phase II UDP-glucuronosyltransferase 1A (UGT1) gene encodes at least eight differently regulated mRNAs whose protein products function as the principal means to eliminate a vast array of steroids, heme metabolites, environmental toxins, and drugs. The orphan nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) were originally identified as sensors able to respond to numerous environmentally derived foreign compounds (xenobiotics) to promote detoxification by phase I cytochrome P450 genes. In this report, we show that both receptors can induce specific UGT1A isoforms including those involved in estrogen, thyroxin, bilirubin, and carcinogen metabolism. Transgenic mice expressing a constitutively active form of human PXR show markedly increased UGT activity toward steroid, heme, and carcinogens, enhanced bilirubin clearance, as well as massively increased steroid clearance. The ability of PXR and constitutive androstane receptor and their ligands to transduce both the phase I and phase II adaptive hepatic response defines a unique transcriptional interface that bridges the ingestion and metabolism of environmental compounds to body physiology.

The binding of the saturated diterpenoid phytanic acid, nuclear receptor RXR activator, docosahexaenoic acid, and bile acid lithocholic acid compete for binding to the 9-cis-RA binding site of the human retinoid X receptor beta protein.

Results point out that Glu83 and E134 amino acid residues are essential for the catalytic activity of human phenol sulfotransferase. These carboxyl residues are localized to active site of the enzyme as shown by the use of carboxyl-specific amino acid modification reagent, Woodwards reagent K.

Research has been conducted on the cellular retinoic acid-binding proteins I and II (CRABP-I and -II), the transport proteins for all-trans-retinoic acid. The identification of the amino acids that comprise the CRABP-I's ligand binding site has been carried out via direct photoaffinity labeling.

Hepatic hydroxylation is an essential step in the metabolism and excretion of bile acids and is necessary to avoid pathologic conditions such as cholestasis and liver damage. In this report, we demonstrate that the human xenobiotic receptor SXR (steroid and xenobiotic receptor) and its rodent homolog PXR (pregnane X receptor) serve as functional bile acid receptors in both cultured cells and animals. In particular, the secondary bile acid derivative lithocholic acid (LCA) is highly hepatotoxic and, as we show here, a metabolic substrate for CYP3A hydroxylation. By using combinations of knockout and transgenic animals, we show that activation of SXR/PXR is necessary and sufficient to both induce CYP3A enzymes and confer resistance to toxicity by LCA, as well as other xenotoxicants such as tribromoethanol and zoxazolamine. Therefore, we establish SXR and PXR as bile acid receptors and a role for the xenobiotic response in the detoxification of bile acids.