Identification of two distinct regions of p38 MAPK required for substrate binding and phosphorylation

The mechanism by which different mitogen activated protein kinases (MAPKs) distinguish between different substrates is poorly understood. For example, p38 and SAPK4 are two closely related p38 MAPKs that both phosphorylate ATF2 and MBP. However, p38 phosphorylates MAPKAPK-2 and -3, whereas SAPK4 does not. In this study, we have used mutagenesis to determine the regions of p38 required for substrate selection. Alanine scanning mutagenesis identified one region of p38 that was required for its ability to phosphorylate MAPKAPK-2 and -3, but that did not significantly affect its binding to these substrates. Chimeras of p38 and SAPK4 identified a second region of p38 that affected the ability of p38 to both bind and phosphorylate MAPKAPK-2 and -3. Hence, we show for the first time that MAPKs contain two distinct regions for recognizing and phosphorylating protein substrates.