Regulation of protein phosphatase‐1_G from rabbit skeletal muscle: 2. Catalytic subunit translocation is a mechanism for reversible inhibition of activity toward glycogen‐bound substrates

The glycogen‐associated form of protein phosphatase‐1 (PP‐1_G) comprises a 37‐kDa catalytic (C) subunit and a 161‐kDa glycogen‐binding (G) subunit. In the preceding paper in this issue of the journal we showed that the C subunit is released from PP‐1_G in response to phosphorylation of the G subunit by cAMP‐dependent protein kinase. We now show that at 0.15–02 M KCl the phosphorylase phosphatase activity of glycogen‐bound PP‐1_G is 5–8 times higher than that of released C subunit or unbound PP‐1_G, which are strongly inhibited at these ionic strengths. The activity of glycogen‐bound PP‐1_G towards glycogen synthase was about 5‐fold higher than that of released C subunit at 0.15M KCl. Studies with glycogen‐bound substrates and myosin P‐light chain (which does not interact with glycogen) indicated that PP‐1_G activity is only enhanced compared to free C subunit at near physiological ionic strength and when both PP‐1_G and substrate are glycogen‐associated. The inhibition by increasing ionic strength and enhanced activity upon binding to glycogen reflected changes in K′_m, but not V_max. From the determined specificity constant, k′_cat/K′_m∼ 4 × 10⁶ s⁻¹ M⁻¹, it was calculated that at physiological levels of glycogen‐bound PP‐1_G (200 nM) and phosphorylase (70 μM). dephosphorylation of the latter could occur with a half time of 15 s, sufficient to account for inactivation rates in vivo. The much higher catalytic efficiency of glycogen‐bound PP‐1_G toward the glycogen‐metabolising enzymes at physiological ionic strength compared to free C subunit substantiates the role of PP‐1_G in the regulation of these substrates, and establishes a novel mechanism for selectively regulating their phosphorylation states in response to adrenalin and other factors affecting phosphorylation of the G subunit.