Activation of phosphorylase phosphatase by a novel procedure: Evidence for a regulatory mechanism involving the release of a catalytic subunit from enzyme-inhibitor complex(es) of higher molecular weight

Regulation of type-1 protein phosphatase in a model of metabolic arrest

“…Furthermore, ethanol treatment increased activity from ACT and EST snails to the same level. Taken together, we interpret that ethanol treatment liberated the catalytic subunit (PP1c) from inhibitory complexes of higher molecular masses, as has been determined in vertebrates (20,21) and appears conserved in invertebrates (4,22). During initial heparin chromatography of snail extracts, a type-2A phosphatase also bound to the column.…”

“…Crude extracts were subjected to precipitation with 95% ethanol (5：1 v：v) for 1 h at 22 o C to remove potential regulatory subunits (20,21). After ethanol treatment, samples were centrifuged for 5 min at 10,000 g, ethanol was removed and pellets were resuspended in the original volume of buffer A. PP1 activity was then purified using modifications of established chromatography methods (10,24).…”

Phosphorylase phosphatase: new horizons for an old enzyme

“…We discovered that ethanol or trypsin treatment of tissue extracts led to large increases in phosphorylase phosphatase activity, concomitant with the reduction in apparent size to the free catalytic subunit (17). These findings could be rationalized by the removal of an inhibitor protein(s ) which were complexed with the catalytic subunit, and led to the discovery of the heat-stable PP1 inhibitor proteins of (19).…”

“…These findings could be rationalized by the removal of an inhibitor protein(s ) which were complexed with the catalytic subunit, and led to the discovery of the heat-stable PP1 inhibitor proteins of (19). The findings for the existence of heat-stable, trypsin-sensitive protein inhibitors of PP1 (17,19) played an important role in unraveling the enzymology of PP1 from two perspectives, the role of inhibitor proteins in regulating phosphatase activity, and the existence of holoenzyme forms. Subsequent studies (20) identified two such inhibitors: inhibitor-1 and inhibitor-2, which have been extensively studied (for reviews, see [8][9][10].…”

“…Underlying this success was discovery that treatment of tissue extracts with denaturants (alcohol or urea) led to the release of a 35 kDa catalytic subunit which could be readily isolated (17). (For the remainder of this review, we will refer to the catalytic subunit simply as PP1.)…”