Computational protocol: Evaluation of a novel virtual screening strategy using receptor decoy binding sites

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[…] Ligand and decoy sets for fifteen target proteins were downloaded from the Database of Useful Decoys []. The complexes were selected from several different protein categories in the database such as hormone receptors, kinases, proteases and other enzymes to represent a wide range of targets, including 10 targets which had previously been evaluated []. Virtual screening for all fifteen targets was performed using Autodock Vina version 1.1.1 with the default parameters []. The FTMap binding site prediction server [] was used to help define the decoy site for docking. The FTMap server identifies binding hot-spots by computational solvent mapping whereby 16 different molecular probes are docked onto the protein surface to locate favorable binding regions []. The decoy site was chosen based on the following criteria: 1) contains no binding hotspot predicted by FTMap, 2) it appears structurally different to the actual binding site and 3) it does not form an obvious binding cavity but is at a flat region on the exterior surface of the protein. The search space for docking was defined via a grid box manually specified with Autodock Tools [] around the binding or decoy site. A grid spacing of 0.375 Å was used to determine the box dimensions. The box dimensions remained the same for binding site and decoy site docking. Adjusted rank lists were generated from the binding site list by considering molecules that were in the top 10 %, 15 %, 20 %, 30 % and 50 % of the decoy site list, and adjusting the rank of the binding site list using the following formula:Adjustedrank=Bindingsiterank−Decoysiterank+Totalno.ofligandsinlistThe fraction of decoy-site docking results was varied in order to find a cut-off where maximum enrichment is achieved. The numbers of active ligands in the database were then used to calculate the ROC Enrichment (ROCE) factors at 1 % and 2 % of the number of molecules. The ROCEx% was calculated as the fraction of true positives divided by the fraction of false positives at x% of the ligand/decoy database according to the equation:ROCEx%=factives1−Ndecoys−NinactivesNdecoysWhere factives = (number of actives at x%) / (number of all actives),Ndecoys = the total number of inactive decoys,Ninactives = the number of decoys chosen at x% of the ligand/decoy database.Binding site and decoy sites were analysed post-docking with the KVFinder Cavity Detection PyMol Plugin [] to provide a quantitative description of the two sites. The software enables comparison and characterisation of protein binding sites by the number, area and volume of cavities in a specified search space. The default parameters were used for all fifteen targets which included a probe in size of 1.4 Å, probe out size of 4.0 Å and a step size of 0.6 Å. The minimum cavity volume was set at 5.0 Å. The binding site search space was set around the position of the actual ligand molecule obtained from the Protein Data Bank, and the decoy site search space was set using a docked molecule from the decoy site screening. […]

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