ADP-Sensitive and -Insensitive Phosphorylated Intermediates of Solubilized Ca2+, Mg2+-Dependent ATPase of the Sarcoplasmic Reticulum from Skeletal Muscle1

Abstract

Solubilized Ca²⁺, Mg²⁺-dependent ATPase [EC 3.6.1.3] with stable activity was prepared from sarcoplasmic reticulum of skeletal muscle by a modification of the method of le Maire et al. (1976) (Biochemistry 15, 2336–2342). The dependence on ATP concentration of the ATPase activity (v₀), the amount of EP in the steady state and the initial rate of EP formation followed the Michaelis-Menten equation. ADP inhibited the ATPase activity non-competitively and the value of v₀/[EP] was constant over a wide range of ADP concentrations. These results indicate the existence of EP which decreased in amount on addition of ADP. The amounts of two kinds of EP, one of which could react with ADP to form ATP and the other of which could not, were measured at alkaline pH where the EP decomposition was very slow. The following results were obtained.