Removal of phosphate from phosphohistidine in proteins

Abstract

Kinetic constants of KM = 0.8 microM, 3 microM and 1.6 microM, and kcat = 9 s-1, 7 s-1 or 9 s-1 were determined for histidine dephosphorylation by protein phosphatases 1, 2A and 2C respectively. IC50 values were determined for the inhibition of protein phosphatase 1 by inhibitor 1 (IC50 = 1 nM), inhibitor-2 (IC50 = 3 nM) and okadaic acid (IC50 = 30 nM) and for the inhibition of protein phosphatase 2A by okadaic acid (IC50 = 0.02 nM) and microcystin-LR (IC50 = 1 nM). Inhibitor-1 (Ki = 0.7 nM) and okadaic acid (Ki = 32 nM) are noncompetitive with protein phosphatase 1. Some of the IC50 values were low enough to violate the assumptions of the usual inhibition equations and a more general approach to the analysis of the data was used. On the basis of these kinetic parameters and the presence of phosphohistidine, the major cellular protein serine/threonine phosphatases are likely to act as protein histidine phosphatases in the cell.