Background: Selenoprotein K is important for calcium-dependent activation of immune cells. Results: Selenoprotein K is cleaved by m-calpain in resting macrophages, but Toll-like receptor activation induces calpastatin generating full-length, functional selenoprotein K. Conclusion: Proteolytic modulation of selenoprotein K is important for macrophage activation. Significance: New roles are defined for the calpain/calpastatin system and selenoprotein K during macrophage activation and inflammation.