Partial purification from rat liver microsomes of three native protein phosphatases with activity towards HMG-CoA reductase.
A procedure for the isolation and partial purification of three hydroxymethylglutaryl coenzyme A reductase phosphatases in their native high molecular weight form from rat liver microsomes is described for the first time. Reductase phosphatase Ex (Mr 90,000), IM (Mr 75,000), and IIM (Mr 180,000) were purified 132-, 55-, and 98-fold, respectively. Treatment with 80% ethanol irreversibly inactivated the three enzymes contrary to what is found for cytosolic reductase phosphatases. The three microsomal reductase phosphatases differ among themselves and with respect to the cytosolic reductase phosphatases in molecular weight, response to inhibitors, thermal stability, and optimum pH. Indirect evidence that these three proteins are phosphatases includes their inhibition by inhibitors of phosphatase activity, such as KF, Pi, and PPi. Direct evidence includes their ability to release 32P from highly radioactive homogeneous 32P-labeled HMG-CoA reductase, this dephosphorylation being concomitant with activation of HMG-CoA reductase. The three phosphatases dephosphorylate 32P-labeled phosphorylase a, but only reductase phosphatase IIM shows glycogen synthase phosphatase activity.
D000818:Animals; D002845:Chromatography; D011756:Diphosphates / Q000494:pharmacology; D004355:Drug Stability; D000431:Ethanol / Q000494:pharmacology; D005459:Fluorides; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology*; D008970:Molecular Weight; D010710:Phosphates / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D011188:Potassium / Q000494:pharmacology; D017680:Potassium Compounds; D051381:Rats; D013379:Substrate Specificity
D011756:Diphosphates; D010710:Phosphates; D017680:Potassium Compounds; D000431:Ethanol; C033320:potassium fluoride; C024938:HMG-CoA reductase phosphatase; D010749:Phosphoprotein Phosphatases; D005459:Fluorides; D011188:Potassium