Human aryl-hydrocarbon receptor and its interaction with dioxin and physiological ligands investigated by molecular modelling and docking simulations Maria Salzano a , Anna Marabotti b , Luciano Milanesi b , Angelo Facchiano a,⇑ a Institute of Food Science (ISA), CNR, Via Roma 64, 83100 Avellino, Italy b Institute of Biomedical Technologies (ITB), CNR, Via F.lli Cervi 93, 20090 Segrate (MI), Italy article info Article history: Received 22 July 2011 Available online 17 August 2011 Keywords: Aryl-hydrocarbon receptor Dioxin Xenobiotics Docking simulations abstract Molecular structure of the ligand binding domain of hAhR has been modelled by homology modelling techniques and used for docking simulations with dioxin and nine more xenobiotics and endogenous ligands. The study evidences that different sites may bind these ligands, whereas only one binding site has been previously indicated by past studies on the mouse homologous receptor. The differences in the sequence of mouse and human AhR ligand binding domain may explain this observation, being most of them in the additional sites observed. Preferences of the evaluated ligands for the different sites are reported and discussed in view of their functional role. Ó 2011 Elsevier Inc. All rights reserved. 1. Introduction Dioxins are a group of chlorinated heteroaromatic polyciclic organic chemicals, and the term usually includes polychlorinated dibenzodioxins and polychlorinated dibenzofurans. Some of them have harmful characteristics depending on the number and struc- tural position of chlorine atoms [1]. 2,3,7,8-Tetrachlorodibenzo-p- dioxin (TCDD or dioxin), the most toxic member of the group, with four chlorine atoms, is considered one of the most toxic com- pounds ever released into the environment [2,3]. Immune system toxicity and dysfunction are some of the most consistent features observed in all animal species following exposure to dioxins and related chemicals. In humans, TCDD produces a broad spectrum of effects at very low concentrations, leading to indicate TCDD as an ‘‘environmental hormone’’. At non-lethal doses, reproductive and developmental effects, hepato-carcinogenesis, tumour promo- tion, and immune suppression are observed [3,4]. The link between chemicals-induced toxicity and clinical effects on human health is the activation of aryl-hydrocarbon receptor (AhR), a cytosolic transcription factor that, in its latent unliganded state, forms complexes with HSP90, p23 and XAP2. Upon ligand binding, AhR translocates to the nucleus, where it complexes with its hetero-dimerization partner, the AhR Nuclear Translocator (ARNT), to modulate expression of AhR target genes containing specific DNA enhancer sequences, known as AhR responsive ele- ments (AhREs) [5,6]. AhR is a member of the bHLH-PAS (basic he- lix-loop-helix Per ARNT Sim) protein family found in organisms as diverse as Caenorhabditis elegans, Drosophila melanogaster and mammals. bHLH-PAS proteins are biological sensors for a variety of stimuli, controlling neurogenesis, vascularization, circadian rhythms, metabolism and stress responses to hypoxia, among oth- ers [7,8]. AhR is highly conserved in evolution and is present in many cell types, albeit at different abundance [9,10]. The selective forces that led to the high degree of conservation of the AhR amino acid sequence are unknown and its physiological function(s) are still being elucidated. The search for bona fide endogenous ligands that regulate AhR transcriptional activity under physiological con- ditions has had limited success, too. Several efforts were made to model the ligand binding domain (LBD) of AhR in mouse [11,14]. In all these studies, the attention was focused on PAS B domain, one of the two structural repeats (PAS A and PAS B) within the PAS domain, that contains the pri- mary HSP90 and also the ligands binding site [10,13,15]. Studies examining the ability of low- and high-affinity ligands to bind to AhRs with mutations in the ligand-binding pocket, have provided some evidences for differential interactions of ligands with the AhR LBD and for amino acids involved in the binding of ligands, thus identifying a specific binding site. 0006-291X/$ - see front matter Ó 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2011.08.039 Abbreviations: AhR, aryl-hydrocarbon receptor; ARNT, AhR Nuclear Transloca- tor; AhREs, AhR responsive elements; bHLH-PAS, basic helix-loop-helix Per ARNT Sim; hAhR, human AhR; LBD, ligand binding domain; mAhR, mouse AhR; TCDD, 2,3,7,8-tetrachlorodibenzo-p-dioxin; dFICZ, 6,12-diformylindolo(3,2-b)carbazole; FICZ, 6-formylindolo(3,2-b)carbazole; PentaCB, 3,3 0 ,4,4 0 ,5 0 -Pentachlorobiphenyl. ⇑ Corresponding author. Fax: +39 0825 781585. E-mail addresses: maria.salzano@isa.cnr.it (M. Salzano), anna.marabotti@itb. cnr.it (A. Marabotti), luciano.milanesi@itb.cnr.it (L. Milanesi), angelo.facchiano@ isa.cnr.it (A. Facchiano). Biochemical and Biophysical Research Communications 413 (2011) 176–181 Contents lists available at SciVerse ScienceDirect Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc