Modulation of calcineurin phosphotyrosyl protein phosphatase activity by calmodulin and protease treatment.

Calcineurin (CN) dephosphorylated [32P] phosphotyrosyl glutamine synthetase, a model phosphoprotein substrate containing approximately 1 mol of phosphotyrosine per mol subunit. Phosphatase activity with and without calmodulin (CaM) was greatly stimulated by Mn2+; with Ca2+, even in the presence of CaM, activity was very low. CaM-stimulated phosphatase activity exhibited deactivation with time; initial rates declined markedly after 2-3 min. The Michaelis constant for substrate (3 microM) was identical whether 2 or 12 min assays (with CaM) were used suggesting that the decreased rate of hydrolysis did not result from a decrease in affinity for the phosphoprotein substrate. Limited proteolysis of CN by chymotrypsin increased phosphatase activity 2-3 times that of CaM-supported activity; however, addition of CaM to assays with protease-activated CN reduced activity to that observed for non-proteolyzed enzyme. These data suggest that, in addition to stimulation, CaM can inhibit certain activated conformations of the phosphatase.