Computational protocol: Lead identification for the K-Ras protein: virtual screening and combinatorial fragment-based approaches

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Protocol publication

[…] Compounds were selected from six different databases (Asinex, BindingDB, DrugBank, National Cancer Institute [NCI], PubChem, and Chembridge database) of varying size (numbers of molecules) and type (divers scaffolds) as a starting point for VS. In this study, 1 13,962 diverse small molecules from Asinex platinum collection, 1,602 FDA-approved small molecules drugs from DrugBank,– 408 Ras protein inhibitors from BindingDB,– 4,000 lead-like small molecules from KinaCore library and 10,000 drug-like compounds from KinaSet library of Chembridge database, 1,990 compounds of NCI diversity set from NCI database, 155 kinase inhibitors from PubChem compound databases, and 267 active molecules from PubChem bioassay (AID-759) entitled HTS were taken to identify specific small molecule inhibitors of Ras and Ras-related GTPases, specifically Ras wild type. The entire set of molecules was subjected to ligand preparation. Ligands were submitted to the LigPrep module of Schrödinger (Schrödinger Release 2015-1; Schrödinger, LLC, NY, USA) to generate ionization states populated at crystallized condition of K-Ras protein pH 8.5 by using epik, enumerating isomers and tautomers and generating three-dimensional conformations. [...] The crystal structure of K-Ras protein complex with GDP (Protein Data Bank [PDB] ID: 4OBE) and GTP (PDB ID: 4DSO) was downloaded from PDB. 4OBE is wild-type GDP-bound structure of K-Ras having a resolution 1.24 Å, and 4DSO is a G12D GTP-bound structure of K-Ras having a resolution 1.85 Å. 4DSO mutant is converted into wild-type structure by mutated Asp12 to Gly12. The mutation was incorporated into the structure by homology modeling of K-Ras sequence, for which 4DSO was used as a template. Both proteins were prepared by using protein preparation wizard panel of Schrödinger Suites 2015.1 (Schrödinger Release 2015-1). Hydrogens were added to its corresponding carbon atom and crystallized water molecules were removed from the structures. Protonation states of the titratable residues were assigned at 8.5 pH. The structures were minimized by using Optimized Potentials for Liquid Simulations 2005 force field. The grid was generated by considering cocrystallized GDP/GTP molecule as a grid center using glide for docking calculations. [...] Combinatorial library was generated by using CombiGlide workflow of Schrödinger. These workflows can be used either as a prelude to combinatorial libraries’ design that may be screened for leads and identifying novel scaffolds, or generate focused libraries in support of lead discovery and optimization process. To design lead molecule for the K-Ras GN-binding pocket, combinatorial library design strategy was utilized. The fragments were substituted into a common template core (guanine) at a defined attachment point to graft structural features that are present in different structures (). All possible combinations of fragments and linker with the core were explored. The core, linker, and fragments were assembled into about 260 molecules. […]

Pipeline specifications

Software tools LigPrep, Epik, Protein Preparation Wizard, Glide, CombiGlide
Databases DrugBank BindingDB PubChem BioAssay PubChem Compound
Applications Drug design, Protein interaction analysis
Organisms Rattus norvegicus
Chemicals Guanosine Diphosphate, Guanosine Triphosphate, Nucleotides