Brenke, J.K.J.K.BrenkeSalmina, E.S.E.S.SalminaRingelstetter, L.L.RingelstetterDornauer, S.S.DornauerKuzikov, M.M.KuzikovRothenaigner, I.I.RothenaignerSchorpp, K.K.SchorppGiehler, F.F.GiehlerGopalakrishnan, J.J.GopalakrishnanKieser, A.A.KieserGul, S.S.GulTetko, I.V.I.V.TetkoHadian, K.K.Hadian2022-03-052022-03-052016https://publica.fraunhofer.de/handle/publica/24607510.1177/1087057116639992In high-throughput screening (HTS) campaigns, the binding of glutathione S-transferase (GST) to glutathione (GSH) is used for detection of GST-tagged proteins in protein-protein interactions or enzyme assays. However, many false-positives, so-called frequent hitters (FH), arise that either prevent GST/GSH interaction or interfere with assay signal generation or detection. To identify GST-FH compounds, we analyzed the data of five independent AlphaScreen-based screening campaigns to classify compounds that inhibit the GST/GSH interaction. We identified 53 compounds affecting GST/GSH binding but not influencing His-tag/Ni2+-NTA interaction and general AlphaScreen signals. The structures of these 53 experimentally identified GST-FHs were analyzed in chemoinformatic studies to categorize substructural features that promote interference with GST/GSH binding.en572journal article