Phosphorylation/dephosphorylation assays

Abstract

The P-type ATPases form an autophosphorylated intermediate with ATP, and its isomeric transition and hydrolysis are obligatory events in the ATP-driven pump and thus for the energy coupling. The analyses of these reactions are therefore crucial for understanding the mechanism of the pump function and diseases caused by its defects. Here we describe the methods to analyze these processes in the transport cycle with a representative member of P-type ATPase family, SERCA1a, sarco(endo)plasmic reticulum Ca2+-ATPase.