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Article Cite This: ACS Omega 2018, 3, 18214−18226
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Deciphering G‑Protein-Coupled Receptor 119 Agonists as Promising Strategy against Type 2 Diabetes Using Systems Biology Approach Aman Chandra Kaushik,†,‡ Ajay Kumar,§ Ashfaq Ur Rehman,† Muhammad Junaid,† Abbas Khan,† Shiv Bharadwaj,*,∥ Shakti Sahi,*,‡ and Dong-Qing Wei*,†
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†
State Key Laboratory of Microbial Metabolism and School of life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China ‡ School of Biotechnology, Gautam Buddha University, Greater Noida 201312, U.P., India § Institute of Biomedical Sciences & Department of Mechanical and Electro-Mechanical Engineering, National Sun Yat-sen University, Kaohsiung 80424, Taiwan ∥ Nanotechnology Research and Application Center, Sabanci University, Istanbul 34956, Turkey S Supporting Information *
ABSTRACT: Type 2 diabetes (T2D) has been established as a serious and chronic medical condition with clinical feature of insulin deficiency and the resultant pathologically high glucose level in blood. Among various approaches purposed for T2D treatment, small molecule like agonist for G-protein-coupled receptor 119 (GPR119) was suggested to regulate blood glucose level by stimulating pancreatic βcells function. This study employed homology and threading modeling approach to predict the GPR119 structure and conducted the structure-based virtual screening (SBVS) to discover novel agonist for GPR119 signal activation. Although the SBVS approach concluded a total of 419 compounds, we selected only 10 compounds for validation based on their docking scores (threshold values were fixed between −16.23 and −10.00) with GPR119 active site. Further, biochemical pathway simulation was also conducted for T2D involving GPR119 and 10 screened compounds using system biology approach. However, we observed that only compound 1 (C23H29F3O6), with characteristics such as docking score of −16.227, MMGBSA value of −66.23, exhibiting attraction for GLN65, ARG71, and THR86 residues, and optimum concentration of 0.50 μM, has the potential to activate GPR119 signaling and subsequently regulated the glucose-dependent insulin secretion in blood. Hence, this novel compound 1 has vindicated its capacity for further development as a potential therapeutic agent in the treatment of T2D.
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INTRODUCTION Diabetes mellitus, also known as type 2 diabetes (T2D), is a progressive chronic disease.1,2 An estimation revealed that approximately 422 million people were affected worldwide with T2D by 2014 and predicted a significant T2D incidence by 2030.3 Generally, T2D has been characterized by a relative high blood glucose level (hyperglycemia) linked to metabolic factors such as insulin resistance, lowered glucose tolerance, or insulin deficiency.2 Moreover, T2D progression has been established with detrimental consequences such as cardiovascular diseases, neuropathy retinopathy, and nephropathy.1,4 Currently, several types of antidiabetic drugs such as short-acting insulin secretagogues, α-glucosidase inhibitors, sodium-glucose transporter 2 inhibitors, sulfonylureas, dipeptidyl peptidase-4 inhibitors, thiazolidines, biguanides, and glucagon-like peptide1 (GLP-1) have been employed to control glycemia.5,6 Although these drugs are widely used in diabetic patients as monotherapy or formulation of drugs with diverse modes of action to treat T2D, most of these drugs induced side effects such as hypoglycemia and β-cell dysfunction.5 Currently, metformin drug has been broadly used as an effective front-line therapy © 2018 American Chemical Society
against T2D, but the need for additional therapies was purposed in case of failure of metformin against appropriate regulation of glucose homeostasis.7 G-protein-coupled receptor 119 (GPR119) was characterized as class A member of the rhodopsin subfamily in Gαs-proteincoupled receptor and predominately expressed in fetal liver, gastrointestinal tract (K- and L-cells), and pancreatic β-cells8−13 via GPR119 gene.14 Recently, GPR119 has been established as a potential target for the treatment of T2D, as is evident from various published reports.1,15 It was suggested that agonist mediated the activation of endogenous expressed GPR119 in HEK293 and HIT-T15 cells through adenylyl cyclase to increase the cyclic adenosine monophosphate (cAMP) levels.11,16 Besides, GPR119 simulation in pancreatic β-cells directly influenced the glucose-dependent insulin secretion and indirectly regulated the gut peptides (GLP-1, GIP, and PYY) secretion from enteroendocrine cells into intestinal tissue and Received: August 8, 2018 Accepted: October 22, 2018 Published: December 26, 2018 18214
DOI: 10.1021/acsomega.8b01941 ACS Omega 2018, 3, 18214−18226
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Figure 1. Schematic plan shows five steps employed in deciphering G-protein-coupled receptor 119 (GPR119) agonist as a promising strategy in the treatment of T2D. In step (a), complete gene sequence for GPR119 gene was retrieved from NCBI database. In step (b), collected sequence was used for predicting protein 3D structure using homology and threading (multiple template) modeling method. In step (c), potential agonists were screened at the predicted GPR119 active site by structure-based virtual screening method. In step (d), selected GPR119 agonists were further validated for their stability complex with protein using molecular dynamics (MD) approach. Finally, step (e), GPR119 agonists were evaluated in the designed complete biochemical pathway that showed significant reduction in T2D symptoms by regulating insulin secretion in blood.
Figure 2. Ramachandran plot for the predicted GPR119 model using RAMPAGE server. Both (a) original model and (b) minimized model for GPR119 structure showed no significant changes in the favored region (92.6%), indicating the stability of the predicted GPR119 model.
agonist to improve β-cells function and lower the blood glucose levels in T2D.19 Indeed, significant research has been conducted to discover synthetic GPR119 agonists such as MBX 2982, PSN821, LEZ 763, DS 8500, and GSK-1292263 for therapeutic treatment against T2D.1,7 Despite tremendous exertions, these agonists were discontinued in the Phase II clinical trials17 and
bloodstream.1,17 Moreover, animal model studies also exhibited that GPR119 mediated glucagon-like peptide 1 (GLP-1) secretion, induced glucose-dependent insulinotropic effects, and improved glucose tolerance.11,16 Besides, upregulation in insulin secretion and essential genes for pancreatic β-cells was also documented.18 These results suggested the use of GPR119 18215
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theoretical assistance of GPR119 agonist for glucose-dependent insulin regulation in T2D is still a challenge to demonstrate clinically.20,21 In the past decade, remarkable progress in structural biology of G-protein-coupled receptors (GPRs) resulted in the publication of 140 GPR crystal structures.22 Also, GPR binding pockets were suggested to be well suited for docking studies and opened the probability to apply structure-based virtual screening (SBVS) approaches to GPR drug discovery efforts.23,24 However, the absence of the GPR119 crystal structure created hurdles in the discovery of novel agonist using in silico approach as treatment against T2D. Hence, we attempted to generate three-dimensional (3D) structure of GPR119 followed by screening of potential agonists from various chemical databases using the SBVS approach. Furthermore, dynamic behavior of selected agonists was also evaluated in the entire biochemical pathway involving GPR119 using systems biology approach and concluded as promising treatment for T2D as shown in Figure 1.
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RESULTS AND DISCUSSION Three-Dimensional Structure Prediction, Minimization, and Validation of GPR119 Model. Using advance python scripts of Modeler, we generated 50 homology models for GPR119 using multitemplates (himology and threadug method) of protein. However, only one homology model was selected for further validation studies based on discrete optimized protein energy (DOPE) score. Following, predicted GPR119 model was optimized and then validated by Ramachandran plot. It was observed that GPR119 model showed no significant changes in the favored region for minimized model (92.6%) against original model (92.6%), establishing the stability of the designed model (Figure 2). Moreover, minimized GPR119 model exhibited significant changes in the allowed (5.5%) and outlier (1.9%) regions against the original model (allowed region, 5.2%; outlier region, 2.3%). Additionally, the minimized model displayed lower energy levels (−11980.84 KJ/mol) in comparison to the original model (−8259.089 KJ/mol), further demonstrating the stable state of the minimized model (Figure 2 and Table S1). Protein Preparation and Seven Transmembrane Domain Identifications. The predicted GPR119 model was prepared for docking study using Protein Prep tool of Schrodinger suite with default predefined and standard parameters. The protein was assigned with bond order at pH 7.0, labeled with disulfide bonds, and marked for zero-order bonds to metals (Na+). Moreover, hydrogen atoms and water molecules were added and deleted beyond 3 Å, respectively, from hetero groups in the GPR119 model using OPLS2005 force fields. The intracellular region in the predicted GPR119 structure was found to possess seven α-helix (α-helix) structural transmembrane domains. These α-helix domains were marked as TM1 TM2, TM3, TM4, TM5, TM6, and TM7 from 10−32, 39−61, 79−101, 121−143, 163−185, 226−248, and 263−282 amino acid positions, respectively, in the predicted structure for GPR119 (Figure 3a). Significantly, TM2 and TM4 domains were predicted to cover CYS42, LEU45, ASN46, VAL49, GLY123, ILE126, and TRP130 residues, respectively, in the GPR119 structure and established as potential targets for T2D inhibition. Also, it was observed that GPR119 can transmit the signals within and outside the cell, whereas extracellular molecules were discovered to react with the archetypal receptor that induced changes in the cell metabolism. The processed signal transduction was also documented to play an important
Figure 3. Three-dimensional structure and active sites−agonist complex of GPR119 model. (a) GPR119 structure exhibited seven αhelix transmembrane domain positions in the intracellular region that are differentiated through various colors, namely, orange, dark yellow, light yellow, light green, dark green, blue, and purple color reflected the domain 1, domain 2, domain 3, domain 4, domain 5, domain 6, and domain 7, respectively. (b) Extracellular and intracellular regions along with short listed 10 potential compounds complexed at the predicted active site of the GPR119 model.
role in potential ligand binding and further caused the cascading of chemical events. Hence, extracellular and intracellular compartments of GPR119 were predicted to participate in interaction with lipid molecules and accountable for GPR119 active site, respectively (Figure 3b). Structure-to-Function Relationship and Grid Generation of GPR119. The GPR119 model was compared with similar X-ray-derived crystal structures (PDB ID: 2VT4, 4Y00, and 2R4S) retrieved from PDB databank for elucidating the structure-to-function relationship of GPR119. The comparison study revealed that CYS42, LEU45, ASN46, VAL49, GLY123, ILE126, and TRP130 residues composed the active region in GPR119. Also, cross-validation of the predicted residues at the active region was further supported by the results produced in SiteMap tool of Schrodinger suite. Also, grid generation was done using Glide tool of Schrodinger suite, whereas van der Waals scaling factor and partial charge were set at 1.0 and 0.25, respectively, with fixed coordinates (15 Å on X-axis, 15 Å on Y-axis, and 15 Å on Z-axis) around the predicted active site in the GPR119 model. 18216
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Figure 4. Docking attraction of SBVS compounds (compounds 1−10) with the GPR119 model. Protein structure is mentioned in magenta and screened compound structures are depicted in green. Also, hydrogen bonding, pi−pi stacking, and aromatic hydrogen bonding are reflected in green dotted, cyan, and pink, respectively.
Table 1. Screened 10 Potential Compounds Are Present in Their Descending Docking Score with the GPR119a compound
docking Score
XLogP3
glide emodel
weight (g/mol)
formula
donorHB
accptHB
H bond interactions
MMGBSA
compound 1 compound 2 compound 3 compound 4 compound 5 compound 6 compound 7 compound 8 compound 9 compound10
−16.227 −15.026 −14.880 −14.850 −14.444 −14.412 −14.405 −14.356 −14.325 −14.315
2.9 3.3 2.44 3.11 2.9 2.9 4.3 3.59 1.23 2.0
−83.630 −81.074 −65.327 −77.930 −4.491 −69.792 −85.445 −86.782 −87.028 −86.308
458.470 423.910 370.480 425.871 525.630 444.400 446.459 438.479 416.413 355.364
C23H29F3O6 C22H28ClO6 C20H34O6 C22H19Cl1N3O4 C32H33N2O5 C23H29F3O6 C25H21N2O6− C24H25N2O6 C21H24N2O5P1 C19H19N2O5−
4 3 3 2 4 4 2 1 4 3
9 6 6 6 9 9 7 9 8 5
GLN65, ARG71, THR86 GLN65, ARG71, THR86 GLN65, ARG71, THR86 GLN65, ARG71 ARG71 ARG71, ARG262 GLN65, ARG71 GLN65, ARG71 GLN65, ARG71 GLN65, ARG71, THR86
−66.23 −74.46 −66.01 −73.26 −87.04 −89.55 −83.36 −70.14 −78.96 −67.79
a
These compound were collected through structure-based virtual screening process of various chemical databases.
Structure-Based Virtual Screening (SBVS), Molecular Modeling, and Pre−Post Dynamics Analysis of Screened Compounds. Following, ligands as agonist from various compound databases were docked at the active site of the
GPR119 model using the SBVS approach. A total of 428507 ligands were evaluated at the active site of GPR119 through a stepwise filter process that yielded a total of 419 compounds (Table S2). Briefly, complete SBVS process was executed in 18217
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Figure 5. X-, Y-plots show time-dependent RMSD behavior from initial state to over 30 ns trajectory. (a) Different curves represent MD simulation (RMSD) analysis for screened 10 compounds, where X-axis represents the times in nanosecond (30 ns for each complex) and Y-axis represents the RMSD in Angstroms (Å). (b) Time-dependent RMSF behavior from initial state to over 30 ns trajectory of GPR119 in a complex with top 10 potential compounds, where X-axis represents the times in nanosecond (30 ns for each complex) and Y-axis represents RMSF in Angstroms (Å).
atoms in docked complexes during simulation. The selected compounds showed acceptable RMSD (
