Inactivation and reactivation of phosphoprotein phosphatase.
The catalytic subunit of phosphoprotein phosphatase (Mr = 35,000) is inactivated by phosphate compounds such as trimetaphosphate, PPi, and ATP. The inactivation of phosphoprotein phosphatase by these phosphate compounds is time- and concentration-dependent, is not reversed by dilution or gel filtration and is protected by Pi. A dissociation constant for the enzyme-trimetaphosphate complex and a rate constant for the reaction were calculated to be 4.6 x 10(-4) M and 0.29 min-1, respectively. The inactivation of phosphatase by PPi and ATP shows more complex kinetics than that by trimetaphosphate. The addition of EDTA to PPi and ATP exhibits more potent inactivation, even though EDTA alone does not inactivate phosphatase. This phosphoprotein phosphatase is not labeled by [gamma-32P]ATP. The inactivation of phosphatase by PPi or ATP can only be reversed by Mn2+ or Co2+, among all other metals or cationic compounds tried. The reactivation also requires sulfhydryl compounds. The effectiveness of sulfhydryl compounds follows the order: dithioerythritol greater than mercaptoethanol greater than cysteine. Glutathione was without effect. Metal analysis of the catalytic subunit did not reveal any significant amounts of Ca, Cd, Co, Cu, Fe, Mg, Mn, Ni, Sn, or Zn. Phosphoprotein phosphatase activity from zinc-deficient rat livers also eliminated the possibility of this phosphatase being a zinc metalloenzyme. Inactivation does not seem to be due to a loss of a critical metal ion. Other mechanisms for inactivation are presented.
D000818:Animals; D002413:Cations, Divalent; D004789:Enzyme Activation; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D010710:Phosphates / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D015032:Zinc / Q000172:deficiency
D002413:Cations, Divalent; D046911:Macromolecular Substances; D010710:Phosphates; D013438:Sulfhydryl Compounds; D010749:Phosphoprotein Phosphatases; D015032:Zinc