A 56 kDa protein kinase was molecularly cloned from human fetal brain. This protein kinase (p56 KKIAMRE) shares homology with p42 KKIALRE (Meyerson et al., 1992) and is related to the proline-directed protein kinase group of signal transducing enzymes. The p56 KKIAMRE and p42 KKIALRE protein kinases exhibit mutually exclusive expression in reproductive tissues; p56 KKIAMRE in testis and p42 KKIALRE in ovary. p56 KKIAMRE and p42 KKIALRE may therefore contribute to signal transduction within these highly differentiated tissues. p56 KKIAMRE and p42 KKIALRE are activated by treatment of cells with epidermal growth factor (EGF). Although p56 KKIAMRE and p42 KKIALRE contain the MAP kinase dual phosphorylation motif Thr-Xaa-Tyr (Thr-Asp-Tyr), phosphorylation on Thr and Tyr within this motif is not required for EGF-stimulated protein kinase activity.