Comparative Modeling and Evaluation of Leukotriene B4 Receptors for Selective Drug Discovery Towards the Treatment of Inflammatory Diseases

摘要

Leukotriene B4 (LTB4) exerts its biological effects through stimulation of specific G protein-coupled receptors (GPCRs)—namely BLT1 and BLT2. Due to the absence of human BLT1 and BLT2 crystal structures, the current study was set to predict the 3D structures of these two receptors for structure-based anti-inflammatory drug discovery. Homology modeling of the BLT1 receptor was first constructed, based on various X-ray and NMR GPCR templates, followed by molecular dynamics (MD) refinement. Using a single-template approach, nine well-established alignment methods and ten secondary structure prediction methods during the backbone generation were implemented and assessed. The binding sites of the BLT1 receptor were then mapped using fifteen chemical probes with the help of FTMAP and AutoDock Vina 4.2 software. Model validation was performed through the docking of eight specific antagonists that have experimental inhibition constants ( k ~( i )) towards BLT1. The antagonists-BLT1 docked structures were then subjected to AMBER-based molecular mechanical minimization and the corresponding binding energies were calculated using molecular mechanics–generalized Born surface area (MM/GBSA) approach. According to the results, the most energetically stable models were constructed using SAlign method for the alignment process and PSIPRED for secondary structure prediction. In comparison, the refined BLT1 model built on 2KS9 as an NMR template has the lowest DOPE energy compared to those built on 4EA3 and 4XT1 as X-ray templates. According to the mapping results, two main binding sites were identified: one was among TMs II, III and VII and the other was among TMs III, IV and V. For the antagonists, correlation between binding energies and experimental data was in a good agreement, with a correlation coefficient ( R _(2)value) of 0.91. Due to the great amino acid sequence similarity between BLT1 and BLT2 receptors (calculated as 45.2%), BLT2 model was constructed based on the predicted BLT1 model.