Identification of a novel small molecule that targets the extracellular regulated kinase‐2 (ERK2) FXFP motif docking site and inhibits substrate phosphorylation and cell proliferation

Abstract

Activation of the ERK/ MAPK signaling pathway is involved in a broad range of cellular processes such as growth, differentiation, death, metabolism, and stress response et al. ERK1/2 proteins play the central role in this signaling pathway by integrating a variety of extracellular signals and distributing the integrated signals to different downstream tiers. ERK1/2 proteins exert their biological activities by phosphorylating and regulating dozens of substrates. The efficiency and specificity of substrate phosphorylation by ERK1/2 are largely dependent on docking domains that facilitate interactions with substrates containing unique binding motifs. To screen for small molecule compounds that selectively manipulate ERK interactions with substrates containing distinct binding motifs, computer aided drug design (CADD) was used to predict small molecules that bind to ERK2 on regions involved in substrate interactions. One compound (2.3.2) was identified and showed high nanomolar binding affinity to ERK2. Compound 2.3.2 selectively inhibited phosphorylation of the DEF motif ERK substrate ELK‐1 as compared to the DEJL motif substrate RSK‐1 and reduced HeLa cell proliferation (IC50~ 1μM). We further co‐crystalized 2.3.2 with ERK2 and solved the structure at 2.2 Å by x‐ray crystallography. The crystal structure confirmed that 2.3.2 binds to ERK2 in the region involved in DEF motif substrate interactions. These findings demonstrate the identification and use of novel small molecules to selectively disrupt ERK activation of substrates involved in cell proliferation.