Fig. 5. 4ax-mediated activation of phosphothreonine lyase OspF in living cells. (A) Schematic representation of the manipulation of OspF's activity with PABK incorporated in place of the catalytic residue K134 (OspF-K134PABK), rendering the enzymatic activity completely masked. The addition of 4ax will effectively decage PABK, resulting in unmasked OspF activity (e.g. irreversible dephosphorylation of p-Erk) (PDB 3I0U). (B) The p-Erk dephosphorylation assay was performed in HEK293T cells. In contrast to wild-type OspF, cells expressing OspF-K134PABK exhibited no dephosphorylation activity on p-Erk. The addition of 4ax rescued OspF's dephosphorylation activity and leads to a decrease in p-Erk levels. 500 µM 4ax was used to treat the cells for 1.5 h.