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Homogenous preparations of the catalytic subunit of protein phosphatase-1 and inhibitor-2 can be combined to produce an inactive enzyme that consists of a 1:1 complex between these two proteins. This species is indistinguishable from the Mg-ATP-dependent protein phosphatase in that preincubation with glycogen synthase kinase-3 and Mg-ATP is required to generate activity. Activation results from the phosphorylation of inhibitor-2. The molar concentrations of protein phosphatase-1 and inhibitor-2 in rabbit skeletal muscle (0.25-0.5 microM) are similar. Incubation of the reconstituted Mg-ATP-dependent protein phosphatase with chymotrypsin is accompanied by limited proteolysis of inhibitor-2 and the loss of its phosphorylation site(s). This species can be activated by glycogen synthase kinase-3 and Mg-ATP provided that inhibitor-2 is added. This exogenous inhibitor-2 appears to displace the fragments of inhibitor-2 from the enzyme that were generated by chymotryptic digestion. These experiments may explain the report [Yang, S.D., Vandenheede, J.R. and Merlevede, W. (1981) J. Biol. Chem. 256, 10231-10234] that inhibitor-2 can function as an 'activator' as well as an inhibitor of the Mg-ATP-dependent protein phosphatase. Incubation of the catalytic subunit of protein phosphatase-1 with sodium fluoride or sodium pyrophosphate converted the enzyme to an inactive form that could be partially reactivated by manganese ions, but not by glycogen synthase kinase-3 and Mg-ATP. Conversely, the reconstituted Mg-ATP-dependent protein phosphatase could only be activated by glycogen synthase kinase-3 and Mg-ATP, and not by manganese ions. It is concluded that the conversion of protein phosphatase-1 to a manganese-ion dependent form is a quite separate phenomenon from the formation of the Mg-ATP-dependent protein phosphatase. Inhibitor-2 can inactivate protein phosphatase-1 by a second mechanism that is not reversed by preincubation with glycogen synthase kinase-3 and Mg-ATP. This occurs at higher concentrations of inhibitor-2 than those required to form the Mg-ATP-dependent protein phosphatase, and appears to result from the binding of inhibitor-2 to a distinct site on the enzyme.
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
January 1984,
Current topics in cellular regulation,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
June 1987,
FEBS letters,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
May 1988,
FEBS letters,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
November 1987,
FEBS letters,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
June 1988,
Analytical biochemistry,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
October 1981,
The Journal of biological chemistry,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
February 1980,
FEBS letters,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
January 1982,
Progress in clinical and biological research,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
November 1994,
The Journal of biological chemistry,
T J Resink, and
B A Hemmings, and
H Y Tung, and
P Cohen
October 1984,
Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.),
