Comprehensive Approach of 19F Nuclear Magnetic Resonance, Enzymatic, and In Silico Methods for Site-Specific Hit Selection and Validation of Fragment Molecules that Inhibit Methionine γ-Lyase Activity

Abstract

Fragment-based screening using ¹⁹F NMR (¹⁹F-FS) is an efficient method for exploring seed and lead compounds for drug discovery. Here, we demonstrate the utility and merits of using ¹⁹F-FS for methionine γ-lyase-binding fragments, together with a ¹⁹F NMR-based competition and mutation assay, as well as enzymatic and in silico methods. ¹⁹F NMR-based assays provided useful information on binding between ¹⁹F-FS hit fragments and target proteins. Although the ¹⁹F-FS and enzymatic assay were weakly correlated, they show that the ¹⁹F-FS hit fragments contained compounds with inhibitory activity. Furthermore, we found that in silico calculations partially account for the differences in activity levels between the ¹⁹F-FS hits as per NMR analysis. A comprehensive approach combining the ¹⁹F-FS and other methods not only identified fragment hits but also distinguished structural differences in chemical groups with diverse activity levels.