The identification and properties of phosphatases in skeletal muscle with activity towards the inhibitory subunit of troponin, and their relationship to other phosphoprotein phosphatases.
1. Phosphoprotein phosphatases with activity towards the inhibitory subunit of troponin (troponin I), phosphorylase a and lysine-rich histone (fraction F1) have been fractionated from rat skeletal muscle by chromatography on Sephadex G-200 and polylysine-Sepharose. Six separate fractions were identified on the basis of substrate specificity and behaviour during chromatography. 2. All fractions showed similar Km values for any given protein substrate. The Km for troponin I (5 muM) was significantly lower than that previously reported. 3. Phosphatase activities towards troponin I and hosphorylase a did not show a requirement for bivalent-metal ions. Two of the fractions with only minor activity towards histone were activated by Mn2+. 4. Discontinuous polyacrylamide-gel-electrophoresis studies indicated that several of the fractions contained more than one phosphatase activity, and additionally showed that several of the activities could exist in different aggregation states. On the basis of these studies at least two phosphatases with activity only towards troponin I were identified. In addition, phosphorylase phosphatase (which has considerable activity towards troponin I) and a general phosphatase with activity towards all three substrates were found. 5. A fraction with mol.wt. of 150000 could be activated by freezing with 2-mercaptoethanol or by heating to 55 degrees C. This activation was accompanied by a decrease in mol.wt. to 25000. 6. The total amount of phosphatase with activity towards troponin I which was extracted would be sufficient to dephosphorylate all the troponin I present in skeletal muscle in approximately 10s.
D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D006657:Histones / Q000378:metabolism; D006358:Hot Temperature; D007700:Kinetics; D008345:Manganese; D008623:Mercaptoethanol / Q000494:pharmacology; D009124:Muscle Proteins / Q000378:metabolism*; D009132:Muscles / Q000201:enzymology*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010765:Phosphorylase Phosphatase / Q000032:analysis; D006005:Phosphorylases / Q000378:metabolism; D051381:Rats; D014336:Troponin / Q000378:metabolism*
D006657:Histones; D009124:Muscle Proteins; D014336:Troponin; D008345:Manganese; D008623:Mercaptoethanol; D006005:Phosphorylases; D010765:Phosphorylase Phosphatase; D010744:Phosphoric Monoester Hydrolases