Role of the transmembrane domain 4/extracellular loop 2 junction of the human gonadotropin-releasing hormone receptor in ligand binding and receptor conformational selection

Recent crystal structures of G protein-coupled receptors (GPCRs) show the remarkable structural diversity of extracellular loop 2 (ECL2), implying its potential role in ligand binding and ligand-induced receptor conformational selectivity. Here we have applied molecular modeling and mutagenesis studies to the TM4/ECL2 junction (residues Pro(174(4.59))-Met(180(4.66))) of the human gonadotropin-releasing hormone (GnRH) receptor, which uniquely has one functional type of receptor but two endogenous ligands in humans. We suggest that the above residues assume an α-helical extension of TM4 in which the side chains of Gln(174(4.60)) and Phe(178(4.64)) face toward the central ligand binding pocket to make H-bond and aromatic contacts with pGlu(1) and Trp(3) of both GnRH I and GnRH II, respectively. The interaction between the side chains of Phe(178(4.64)) of the receptor and Trp(3) of the GnRHs was supported by reciprocal mutations of the interacting residues. Interestingly, alanine mutations of Leu(175(4.61)), Ile(177(4.63)), and Met(180(4.66)) decreased mutant receptor affinity for GnRH I but, in contrast, increased affinity for GnRH II. This suggests that these residues make intramolecular or intermolecular contacts with residues of transmembrane (TM) domain 3, TM5, or the phospholipid bilayer, which couple the ligand structure to specific receptor conformational switches. The marked decrease in signaling efficacy of I177A and F178A also indicates that IIe(177(4.63)) and Phe(178(4.64)) are important in stabilizing receptor-active conformations. These findings suggest that the TM4/ECL2 junction is crucial for peptide ligand binding and, consequently, for ligand-induced receptor conformational selection.     

Molecular modeling of the second extracellular loop of G-protein coupled receptors and its implication on structure-based virtual screening

The current study describes the validation of high-throughput modeling procedures for the construction of the second extracellular loop (ecl2) of all nonolfactory human G Protein-coupled receptors. Our modeling flowchart is based on the alignment of essential residues determining the particular ecl2 fold observed in the bovine rhodopsin (bRho) crystal structure. For a set of GPCR targets, the dopamine D2 receptor (DRD2), adenosine A3 receptor (AA3R), and the thromboxane A2 receptor (TA2R), the implications of including ecl2 atomic coordinates is evaluated in terms of structure-based virtual screening accuracy: the suitability of the 3D models to distinguish between known antagonists and randomly chosen decoys using automated docking approaches. The virtual screening results of different models describing increasingly exhaustive receptor representations (seven helices only, seven helices and ecl2 loop, full model) have been compared. Explicit modeling of the ecl2 loop was found to be important in only one of three test cases whereas a loopless model was shown to be accurate enough in the two other receptors. An exhaustive comparison of ecl2 loops of 365 receptors to that of bRho suggests that explicit ecl2 loop modeling should be reserved to receptors where loop building can be guided by experimental restraints.     

Discovery of Entamoeba histolytica hexokinase 1 inhibitors through homology modeling and virtual screening

Abstract

Entamoeba histolytica, the parasite which causes amebiasis is responsible for 110?000 deaths a year. Entamoeba histolytica depends on glycolysis to obtain ATP for cellular work. According to metabolic flux studies, hexokinase exerts the highest flux control of this metabolic pathway; therefore, it is an excellent target in the search of new antiamebic drugs. To this end, a tridimensional model of E. histolytica hexokinase 1 (EhHK1) was constructed and validated by homology modeling. After virtual screening of 14?400 small molecules, the 100 with the best docking scores were selected, purchased and assessed in their inhibitory capacity. The results showed that three molecules (compounds 2921, 11275 and 2755) inhibited EhHK1 with an I₅₀ of 48, 91 and 96?µM, respectively. Thus, we found the first inhibitors of EhHK1 that can be used in the search of new chemotherapeutic agents against amebiasis.     

Evidence that the thyroid-stimulating hormone (TSH) receptor transmembrane domain influences kinetics of TSH binding to the receptor ectodomain

Thyroid-stimulating hormone (TSH)-induced reduction in ligand binding affinity (negative cooperativity) requires TSH receptor (TSHR) homodimerization, the latter involving primarily the transmembrane domain (TMD) but with the extracellular domain (ECD) also contributing to this association. To test the role of the TMD in negative cooperativity, we studied the TSHR ECD tethered to the cell surface by a glycosylphosphatidylinositol (GPI) anchor that multimerizes despite the absence of the TMD. Using the infinite ligand dilution approach, we confirmed that TSH increased the rate of dissociation (k(off)) of prebound (125)I-TSH from CHO cells expressing the TSH holoreceptor. Such negative cooperativity did not occur with TSHR ECD-GPI-expressing cells. However, even in the absence of added TSH, (125)I-TSH dissociated much more rapidly from the TSHR ECD-GPI than from the TSH holoreceptor. This phenomenon, suggesting a lower TSH affinity for the former, was surprising because both the TSHR ECD and TSH holoreceptor contain the entire TSH-binding site, and the TSH binding affinities for both receptor forms should, theoretically, be identical. In ligand competition studies, we observed that the TSH binding affinity for the TSHR ECD-GPI was significantly lower than that for the TSH holoreceptor. Further evidence for a difference in ligand binding kinetics for the TSH holoreceptor and TSHR ECD-GPI was obtained upon comparison of the TSH K(d) values for these two receptor forms at 4 °C versus room temperature. Our data provide the first evidence that the wild-type TSHR TMD influences ligand binding affinity for the ECD, possibly by altering the conformation of the closely associated hinge region that contributes to the TSH-binding site.     

Discovery of potential novel microsomal triglyceride transfer protein inhibitors via virtual screening of pharmacophore modelling and molecular docking

In order to identify potential natural inhibitors against the microsomal triglyceride transfer protein (MTP), HipHop models were generated using 20 known inhibitors from the Binding Database. Using evaluation indicators, the best hypothesis model, Hypo1, was selected and utilised to screen the Traditional Chinese Medicine Database, which resulted in a hit list of 58 drug-like compounds. A homology model of MTP was built by MODELLER and was minimised by CHARMm force field. It was then validated by Ramachandran plot and Verify-3D so as to obtain a stable structure, which was further used to refine the 58 hits using molecular docking studies. Then, five compounds with higher docking scores which satisfied the docking requirements were discovered. Among them, Ginkgetin and Dauricine were most likely to be candidates that exhibition inhibiting effect on MTP. The screening strategy in this study is relatively new to the discovery of MTP inhibitors in medicinal chemistry. Moreover, it is important to note that, lomitapide, an approved MTP inhibitor, fits well with Hypo1 as well as our homology model of MTP, which confirmed the rationality of our studies. The results indicated the applicability of molecular modeling for the discovery of potential natural MTP inhibitors from traditional Chinese herbs.     

Monte Carlo sampling can be used to determine the size and shape of the steady-state flux space

Constraint-based modeling results in a convex polytope that defines a solution space containing all possible steady-state flux distributions. The properties of this polytope have been studied extensively using linear programming to find the optimal flux distribution under various optimality conditions and convex analysis to define its extreme pathways (edges) and elementary modes. The work presented herein further studies the steady-state flux space by defining its hyper-volume. In low dimensions (i.e. for small sample networks), exact volume calculation algorithms were used. However, due to the #P-hard nature of the vertex enumeration and volume calculation problem in high dimensions, random Monte Carlo sampling was used to characterize the relative size of the solution space of the human red blood cell metabolic network. Distributions of the steady-state flux levels for each reaction in the metabolic network were generated to show the range of flux values for each reaction in the polytope. These results give insight into the shape of the high-dimensional solution space. The value of measuring uptake and secretion rates in shrinking the steady-state flux solution space is illustrated through singular value decomposition of the randomly sampled points. The V(max) of various reactions in the network are varied to determine the sensitivity of the solution space to the maximum capacity constraints. The methods developed in this study are suitable for testing the implication of additional constraints on a metabolic network system and can be used to explore the effects of single nucleotide polymorphisms (SNPs) on network capabilities.     

Monte Carlo docking simulations of cyclomaltoheptaose and dimethyl cyclomaltoheptaose with paclitaxel

The molecular basis for the remarkable enhancement of the solubility of paclitaxel by O-dimethylcyclomaltoheptaose (DM-beta-CD) over cyclomaltoheptaose (beta-cyclodextrin, beta-CD) was investigated with Monte Carlo docking-minimization simulation. As possible guests of inclusion complexation for the host cyclic oligosaccharides, two functional moieties of the suggested solution structure of paclitaxel were used where one is the C-3'N benzoyl moiety (B-ring) and the other is a hydrophobic (HP) cluster site among the C-3' phenyl (C-ring), C-2 benzoate (A-ring), and C-4 acetoxy moieties. The energetic preference of inclusion complexation of DM-beta-CD over beta-CD was analyzed on the basis of more efficient partitioning process of DM-beta-CD into the hydrophobic cluster site of the paclitaxel.     

Biosynthesis and NMR-studies of a double transmembrane domain from the Y4 receptor,a human GPCR

The human Y4 receptor, a class A G-protein coupled receptor (GPCR) primarily targeted by the pancreatic polypeptide (PP), is involved in a large number of physiologically important functions. This paper investigates a Y4 receptor fragment (N-TM1-TM2) comprising the N-terminal domain, the first two transmembrane (TM) helices and the first extracellular loop followed by a (His)₆ tag, and addresses synthetic problems encountered when recombinantly producing such fragments from GPCRs in Escherichia coli. Rigorous purification and usage of the optimized detergent mixture 28 mM dodecylphosphocholine (DPC)/118 mM% 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG) resulted in high quality TROSY spectra indicating protein conformational homogeneity. Almost complete assignment of the backbone, including all TM residue resonances was obtained. Data on internal backbone dynamics revealed a high secondary structure content for N-TM1-TM2. Secondary chemical shifts and sequential amide proton nuclear Overhauser effects defined the TM helices. Interestingly, the properties of the N-terminal domain of this large fragment are highly similar to those determined on the isolated N-terminal domain in the presence of DPC micelles.     

Understanding the structural and functional differences between mouse thyrotropin-releasing hormone receptors 1 and 2

Multiple computational methods have been employed in a comparative study of thyrotropin-releasing hormone receptors 1 and 2 (TRH-R1 and TRH-R2) to explore the structural bases for the different functional properties of these G protein-coupled receptors. Three-dimensional models of both murine TRH receptors have been built and optimized by means of homology modeling based on the crystal structure of bovine rhodopsin, molecular dynamics simulations, and energy minimizations in a membrane-aqueous environment. The comparison between the two models showed a correlation between the higher flexibility and higher basal activity of TRH-R2 versus the lesser flexibility and lower basal activity of TRH-R1 and supported the involvement of the highly conserved W6.48 in the signaling process. A correlation between the level of basal activity and conformational changes of TM5 was detected also. Comparison between models of the wild type receptors and their W6.48A mutants, which have reversed basal activities compared with their respective wild types, further supported these correlations. A flexible molecular docking procedure revealed that TRH establishes a direct interaction with W6.48 in TRH-R2 but not in TRH-R1. We designed and performed new mutagenesis experiments that strongly supported these observations.     

Dosimetric and Monte Carlo verification of jaws-only IMRT plans calculated by the Collapsed Cone Convolution algorithm for head and neck cancers

AimThe aim of this study is to verify the Prowess Panther jaws-only intensity modulated radiation therapy (JO-IMRT) treatment planning (TP) by comparing the TP dose distributions for head-and-neck (H&N) cancer with the ones simulated by Monte Carlo (MC).BackgroundTo date, dose distributions planned using JO-IMRT for H&N patients were found superior to the corresponding three-dimensional conformal radiotherapy (3D-CRT) plans. Dosimetry of the JO-IMRT plans were also experimentally verified using an ionization chamber, MapCHECK 2, and Octavius 4D and good agreements were shown.Materials and methodsDose distributions of 15 JO-IMRT plans of nasopharyngeal patients were recalculated using the EGSnrc Monte Carlo code. The clinical photon beams were simulated using the BEAMnrc. The absorbed dose to patients treated by fixed-field IMRT was computed using the DOSXYZnrc. The simulated dose distributions were then compared with the ones calculated by the Collapsed Cone Convolution (CCC) algorithm on the TPS, using the relative dose error comparison and the gamma index using global methods implemented in PTW-VeriSoft with 3%/3 mm, 2%/2 mm, 1%/1 mm criteria.ResultsThere is a good agreement between the MC and TPS dose. The average gamma passing rates were 93.3 ± 3.1%, 92.8 ± 3.2%, 92.4 ± 3.4% based on the 3%/3 mm, 2%/2 mm, 1%/1 mm criteria, respectively.ConclusionsAccording to the results, it is concluded that the CCC algorithm was adequate for most of the IMRT H&N cases where the target was not immediately adjacent to the critical structures.     

Homology modelling,virtual screening and molecular dynamics of the MARK3 KA1 domain for cancer drug design

Structural homology modelling was done with the software AMPS, MODELLER, PROCHECK, WHATIF AND VERIFY-3D to generate a quality model of human MARK3. Macromolecular docking simulations seem to confirm recent data in the literature and in MARK3 there does not occur intramolecular interactions between the associated kinase domain KA1 and the catalytic domain. Using virtual screening, we were able to identify and suggest the principal residues of MARK3 which interact with the ligands in addition to those reported in the literature. The pharmacophoric model obtained from Discovery Studio coincides with those obtained by molecular interaction fields, indicating the principal ligand residues of the MARK3 KA1 domain. Using virtual screening with pharmacophoric constraints as well as molecular dynamics, the most stable compounds in the ligand site as well as their potential toxicities were used to select potential inhibitors for further in vitro and in vivo investigations of human MARK3 KA1 domain, which could eventually pass to the market to be used for the treatment of head and neck cancer.     

Improved model building and assessment of the Calcium-sensing receptor transmembrane domain

A three-dimensional model of the human Calcium-sensing receptor (CaSR) seven transmembrane domain was built via a novel sequence alignment method based on the conserved contacts in proteins using the crystal structure of bovine rhodopsin as the template. This model was tested by docking NPS 2143, the first identified allosteric antagonist of CaSR. In our model, Glu837 plays a critical role in anchoring the protonated nitrogen atom and hydroxy oxygen atom of NPS 2143. The phenyl moiety of the ligand contacts residues Phe668, Pro672, and Ile841. The naphthalene moiety is surrounded by several hydrophobic residues, including Phe684, Phe688, and Phe821. Our model appears to be consistent with all six residues that have been demonstrated to be critical for NPS 2143 binding, in contrast with existing homology models based on traditional sequence alignment of CaSR to rhodopsin. This provides validation of our sequence alignment method and the use of the rhodopsin backbone as the initial structure in homology modeling of other G protein-coupled receptors that are not members of the rhodopsin family.     

A novel and efficient ligand-based virtual screening approach using the HWZ scoring function and an enhanced shape-density model

In this work, we extend our previous ligand shape-based virtual screening approach by using the scoring function Hamza–Wei–Zhan (HWZ) score and an enhanced molecular shape-density model for the ligands. The performance of the method has been tested against the 40 targets in the Database of Useful Decoys and compared with the performance of our previous HWZ score method. The virtual screening results using the novel ligand shape-based approach demonstrated a favorable improvement (area under the receiver operator characteristics curve AUC?=?.89?±?.02) and effectiveness (hit rate HR^1%?=?53.0%?±?6.3 and HR^10%?=?71.1%?±?4.9). The comparison of the overall performance of our ligand shape-based method with the highest ligand shape-based virtual screening approach using the data fusion of multi queries showed that our strategy takes into account deeper the chemical information of the set of active ligands. Furthermore, the results indicated that our method are suitable for virtual screening and yields superior prediction accuracy than the other study derived from the data fusion using five queries. Therefore, our novel ligand shape-based screening method constitutes a robust and efficient approach to the 3D similarity screening of small compounds and open the door to a whole new approach to drug design by implementing the method in the structure-based virtual screening.     

Identifying the novel inhibitors of lysine-specific demethylase 1 (LSD1) combining pharmacophore-based and structure-based virtual screening

Abstract

Lysine-specific demethylase 1 (LSD1) has been reported to connect with a range of solid tumors. Thus, the exploration of LSD1 inhibitors has emerged as an effective strategy for cancer treatment. In this study, we constructed a pharmacophore model based on a series of flavin adenine dinucleotide (FAD)-competing inhibitors bearing triazole???dithiocarbamate scaffold combining docking, structure–activity relationship (SAR) study, and molecular dynamic (MD) simulation. Meanwhile, another pharmacophore model was also constructed manually, relying on several speculated substrate-competing inhibitors and reported putative vital interactions with LSD1. On the basis of the two pharmacophore models, multi-step virtual screenings (VSs) were performed against substrate-binding pocket and FAD-binding pocket, respectively, combining pharmacophore-based and structure-based strategy to exploit novel LSD1 inhibitors. After bioassay evaluation, four compounds among 21 hits with diverse and novel scaffolds exhibited inhibition activity at the range of 3.63–101.43?μM. Furthermore, substructure-based enrichment was performed, and four compounds with a more potent activity were identified. After that, the time-dependent assay proved that the most potent compound with IC₅₀ 2.21?μM inhibits LSD1 activity in a manner of time-independent. In addition, the compound exhibited a cellular inhibitory effect against LSD1 in MGC-803 cells and may inhibit cell migration and invasion by reversing EMT in cultured gastric cancer cells. Considering the binding mode and SAR of the series of compounds, we could roughly deem that these compounds containing 3-methylxanthine scaffold act through occupying substrate-binding pocket competitively. This study presented a new starting point to develop novel LSD1 inhibitors.     

FTY720 (Gilenya) phosphate selectivity of sphingosine 1-phosphate receptor subtype 1 (S1P1) G protein-coupled receptor requires motifs in intracellular loop 1 and transmembrane domain 2

FTY720 phosphate (FTY720P) is a high potency agonist for all the endothelial differentiation gene family sphingosine 1-phosphate (S1P) receptors except S1P receptor subtype 2 (S1P(2)). To map the distinguishing features of S1P(2) ligand recognition, we applied a computational modeling-guided mutagenesis strategy that was based on the high degree of sequence homology between S1P(1) and S1P(2). S1P(2) point mutants of the ligand-binding pocket were characterized. The head group-interacting residues Arg3.28, Glu3.29, and Lys7.34 were essential for activation. Mutation of residues Ala3.32, Leu3.36, Val5.41, Phe6.44, Trp6.48, Ser7.42, and Ser7.46, predicted to interact with the S1P hydrophobic tail, impaired activation by S1P. Replacing individual or multiple residues in the ligand-binding pocket of S1P(2) with S1P(1) sequence did not impart activation by FTY720P. Chimeric S1P(1)/S1P(2) receptors were generated and characterized for activation by S1P or FTY720P. The S1P(2) chimera with S1P(1) sequence from the N terminus to transmembrane domain 2 (TM2) was activated by FTY720P, and the S1P(2)(IC1-TM2)(S1P1) domain insertion chimera showed S1P(1)-like activation. Twelve residues in this domain, distributed in four motifs a-d, differ between S1P(1) and S1P(2). Insertion of (78)RPMYY in motif b alone or simultaneous swapping of five other residues in motifs c and d from S1P(1) into S1P(2) introduced FTY720P responsiveness. Molecular dynamics calculations indicate that FTY720P binding selectivity is a function of the entropic contribution to the binding free energy rather than enthalpic contributions and that preferred agonists retain substantial flexibility when bound. After exposure to FTY720P, the S1P(2)(IC1-TM2)(S1P1) receptor recycled to the plasma membrane, indicating that additional structural elements are required for the selective degradative trafficking of S1P(1).     

3D structural model of the G-protein-coupled cannabinoid CB2 receptor

The potential for therapeutic specificity in regulating diseases and for reduced side effects has made cannabinoid (CB) receptors one of the most important G-protein-coupled receptor (GPCR) targets for drug discovery. The cannabinoid (CB) receptor subtype CB2 is of particular interest due to its involvement in signal transduction in the immune system and its increased characterization by mutational and other studies. However, our understanding of their mode of action has been limited by the absence of an experimental receptor structure. In this study, we have developed a 3D model of the CB2 receptor based on the recent crystal structure of a related GPCR, bovine rhodopsin. The model was developed using multiple sequence alignment of homologous receptor sub-types in humans and mammals, and compared with other GPCRs. Alignments were analyzed with mutation scores, pairwise hydrophobicity profiles and Kyte-Doolittle plots. The 3D model of the transmembrane segment was generated by mapping the CB2 sequence onto the homologous residues of the rhodopsin structure. The extra- and intracellular loop regions of the CB2 were generated by searching for homologous C(alpha) backbone sequences in published structures in the Brookhaven Protein Databank (PDB). Residue side chains were positioned through a combination of rotamer library searches, simulated annealing and minimization. Intermediate models of the 7TM helix bundles were analyzed in terms of helix tilt angles, hydrogen-bond networks, conserved residues and motifs, possible disulfide bonds. The amphipathic cytoplasmic helix domain was also correlated with biological and site-directed mutagenesis data. Finally, the model receptor-binding cavity was characterized using solvent-accessible surface approach.     

Homology modeling,virtual screening and dynamics study of proteins involved in Pebrine - Serine protease inhibitor 106 and spore wall protein 26

Abstract

Pebrine is a microsporidian disease caused by Nosema bombycis in Bombyx mori (silk worm) which results in brown/black spots. The affected larvae either spin cocoons which are flimsy with low silk content or not spin a cocoon. It has been hypothesised that Serine Protease Inhibitor 106 (SPN106) is responsible for evasion of host immune system by inhibiting the melanization process in silkworms. Also, Spore Wall Protein 26 (SWP26) has been observed to bind with Ig- like protein Bombyx mori turtle-like protein (Bm-TLP) facilitating the attachment of the microsporidian to the host and contributing to infectivity. Till date, there is no crystal structure of the proteins SPN106, SWP26 and Bm-TLP available. In this study, we performed homology modeling of the three structures using Modeller v9.18 and the binding pockets were identified. Virtual screening was conducted using AutoDock Vina on a ligand library consisting of 28,870 lead-like molecules. The protein stability, compactness, fluctuations and protein-ligand interactions were investigated through Molecular Dynamics (MD) simulations studies using Desmond Maestro 11.3 and a potential lead molecule was identified.

Communicated by Ramaswamy H. Sarma     

Identification of some novel pyrazolo[1,5-a]pyrimidine derivatives as InhA inhibitors through pharmacophore-based virtual screening and molecular docking

The InhA inhibitors play key role in mycolic acid synthesis by preventing the fatty acid biosynthesis pathway. In this present article, Pharmacophore modelling and molecular docking study followed by in silico virtual screening could be considered as effective strategy to identify newer enoyl-ACP reductase inhibitors. Pyrrolidine carboxamide derivatives were opted to generate pharmacophore models using HypoGen algorithm in Discovery studio 2.1. Further it was employed to screen Zinc and Minimaybridge databases to identify and design newer potent hit molecules. The retrieved newer hits were further evaluated for their drug likeliness and docked against enoyl acyl carrier protein reductase. Here, novel pyrazolo[1,5-a]pyrimidine analogues were designed and synthesized with good yields. Structural elucidation of synthesized final molecules was perform through IR, MASS, ¹H-NMR, ¹³C-NMR spectroscopy and further tested for its in vitro anti-tubercular activity against H37Rv strain using Microplate Alamar blue assay (MABA) method. Most of the synthesized compounds displayed strong anti-tubercular activities. Further, these potent compounds were gauged for MDR-TB, XDR-TB and cytotoxic study.     

Discovery of selective farnesoid X receptor agonists for the treatment of hyperlipidemia from traditional Chinese medicine based on virtual screening and in vitro validation

Abstract

Farnesoid X receptor (FXR), a bile acid receptor, has important roles in maintaining bile acid and cholesterol homeostasis, which is an attractive target for hyperlipidemia. Present study aimed to discover potential selective FXR agonists over G-protein coupled bile acid receptor 1 (GPBAR1, TGR5) from traditional Chinese medicine (TCM) by using virtual screening, in vitro studies and molecular dynamics simulation (MD). Ligand-based pharmacophore model for FXR was firstly built to screen FXR agonists from the Traditional Chinese Medicine Database (TCMD). Then, 21 FXR crystal structures were clustered in two types and two representative structures (PDB ID: 3OMM and 3P89) were, respectively, used to carry out molecular docking to refine the screened result. Moreover, the pharmacophore model for GPBAR1 was built to screen selective FXR agonists with no activity on GPBAR1. A set of 24 candidate selective FXR agonists which fitvalue of FXR pharmacophore model and docking score of 3OMM and 3P89 were in the top 100 and cannot match the pharmacophore model for GPBAR1 were obtained. By the lipid-lowering activity test in HepG2 cell lines, Arctigenin was identified to be potential selective FXR agonist with the activity of 20?μmol·L^?1. After down-regulating FXR, Arctigenin could increase the mRNA of FXR while exerted no effect on the mRNA of GPBAR1. MD was further used to interpret the mechanism of Arctigenin with the representative structures. This research provided a new screening procedure for finding selective candidate compounds and appropriate docking models of a target by considering the structure diversity of PDB structures, which was applied to discovery novel selective FXR agonists to treat hyperlipidemia.

Communicated by Ramaswamy H. Sarma