MK2 directly phosphorylates K18 and K20 in vitro. A, comparison of phosphorylation sites in K18/20 with that of MK2 substrates mHSPB1/hHSPB1 and consensus MK2/3 phosphorylation site. Matching residues are in bold, and the phosphorylated S/T is denoted. B, in vitro kinase assay with GST-p38/His-MK2 followed by Western blotting for K8-Ser73 phosphorylation and for total K8 protein. C, in vitro kinase assay for K20-Ser13 phosphorylation. mHSPB1-Ser86 and MK2-Thr334 phosphorylations are shown as controls. D, in vitro kinase assay for K18-Ser52 phosphorylation.