Multiple molecular forms of phosphoprotein phosphatase from rabbit skeletal muscle

Abstract

1. 1. Phosphohistone phosphatase activity of rabbit skeletal muscle was separated into two fractions (phosphatase I and phosphatase II) by Sephadex G-150 chromatography. The molecular weights estimated using a Sepharose 6B column for phosphatase I and phosphatase II were about 300 000 and 150 000, respectively. 2. 2. Phosphatase I was active in the absence of divalent metal ions and was not inhibited by EDTA or EGTA. However, the activity of phosphatase II was almost totally dependent on Mn 2+ or Co 2+ with K m of 0.8 and 0.5 mM, respectively. Freezing of phosphatase I in the presence of 200 mM mercaptoethanol resulted in a change of the molecular size to that of phosphatase II with the disappearance of substrate ( P-histone)-inhibition property, which was observed in phosphatase I at a concentration higher than 10 μM of phosphohistone (based on the alkali-labile 32P content). 3. 3. Glycogen synthetase-D phosphatase activity, which converts the D form of glycogen synthetase (UDPglucose:glycogen 4-α-glucosyltransferase, EC 2.4.1.11) to the I form, was also found in the two fractions. Synthetase-D phosphatase activity of phosphatase I was less dependent on Mn 2+ than that of phosphatase II, which was essentially inactive without the metal ion. 4. 4. Existence of the two types of phosphohistone phosphatase was shown in crude extract of rat liver, kidney, brain, heart muscle and skeletal muscle.