A combination of 19F NMR and surface plasmon resonance for site-specific hit selection and validation of fragment molecules that bind to the ATP-binding site of a kinase

¹⁹F NMR has recently emerged as an efficient, sensitive tool for analyzing protein binding to small molecules, and surface plasmon resonance (SPR) is also a popular tool for this purpose. Herein a combination of ¹⁹F NMR and SPR was used to find novel binders to the ATP-binding pocket of MAP kinase extracellular regulated kinase 2 (ERK2) by fragment screening with an original fluorinated-fragment library. The ¹⁹F NMR screening yielded a high primary hit rate of binders to the ERK2 ATP-binding pocket compared with the rate for the SPR screening. Hit compounds were evaluated and categorized according to their ability to bind to different binding sites in the ATP-binding pocket. The binding manner was characterized by using isothermal titration calorimetry and docking simulation. Combining ¹⁹F NMR with other biophysical methods allows the identification of multiple types of hit compounds, thereby increasing opportunities for drug design using preferred fragments.