The phospholamban phosphatase associated with cardiac sarcoplasmic reticulum is a type 1 enzyme.
Canine cardiac sarcoplasmic reticulum vesicles contain intrinsic protein phosphatase activity, which can dephosphorylate phospholamban and regulate calcium transport. This phosphatase has been suggested to be a mixture of both type 1 and type 2 enzymes (E. G. Kranias and J. Di Salvo, 1986, J. Biol. Chem. 261, 10,029-10,032). In the present study the sarcoplasmic reticulum phosphatase activity was solubilized with n-octyl-beta-D-glucopyranoside and purified by sequential chromatography on DEAE-Sephacel, polylysine-agarose, heparin-agarose, and DEAE-Sephadex. A single peak of phosphatase activity was eluted from each column and it was coincident for both phospholamban and phosphorylase a, used as substrates. The partially purified phosphatase could dephosphorylate the sites on phospholamban phosphorylated by either cAMP-dependent or calcium-calmodulin-dependent protein kinase(s). Enzymatic activity was inhibited by inhibitor-2 and by okadaic acid (I50 = 10-20 nM), using either phosphorylase a or phospholamban as substrates. The sensitivity of the phosphatase to inhibitor-2 or okadaic acid was similar for the two sites on phospholamban, phosphorylated by the cAMP-dependent and the calcium-calmodulin-dependent protein kinases. Phospholamban phosphatase activity was enhanced (40%) by Mg2+ or Mn2+ (3 mM) while Ca2+ (0.1-10 microM) had no effect. These characteristics suggest that the phosphatase associated with cardiac sarcoplasmic reticulum is a type 1 enzyme, and this activity may participate in the regulation of Ca2+ transport through dephosphorylation of phospholamban in cardiac muscle.
Steenaart
N A
NA
Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, Ohio 45267-0575.
Ganim
J R
JR
Di Salvo
J
J
Kranias
E G
EG
D000818:Animals; D002122:Calcium Chloride / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002712:Chlorides; D000242:Cyclic AMP / Q000378:metabolism; D004285:Dogs; D004988:Ethers, Cyclic / Q000494:pharmacology; D006003:Glycogen / Q000378:metabolism; D015636:Magnesium Chloride / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D017895:Manganese Compounds; D009206:Myocardium / Q000201:enzymology*; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000145:classification* / Q000378:metabolism; D011189:Potassium Chloride / Q000494:pharmacology; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D012995:Solubility; D000099211:Phospholamban
D002122:Calcium Chloride; D002135:Calcium-Binding Proteins; D002712:Chlorides; D000242:Cyclic AMP; D004988:Ethers, Cyclic; D006003:Glycogen; D015636:Magnesium Chloride; D008345:Manganese; D017895:Manganese Compounds; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases; D011189:Potassium Chloride; D000099211:Phospholamban; C057399:phospholamban phosphatase; C025340:manganese chloride