Utility of immobilon-bound phosphoproteins as substrates for protein phosphatases from neutrophils.
Immobilon-bound phosphoproteins labeled with 32P were utilized as substrates to study the enzymes in neutrophils that are active against the major products of protein kinase C. The labeled proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred electrophoretically to immobilon-P membranes. Both particulate and soluble phosphatases were found to be active against the blotted phosphoproteins. Reactions were followed by autoradiography as the loss of 32P from individual protein bands. The tumor promoter okadaic acid and the hepatoxin microcystin-LR inhibited these reactions in a manner consistent with the enzymes being type 1 and/or 2A protein phosphatases.
Ding
J B
JB
Department of Cell and Molecular Biology, Boston Biomedical Research Institute, MA 02114.
Badwey
J A
JA
D000818:Animals; D002384:Catalysis; D002478:Cells, Cultured; D006168:Guinea Pigs; D009504:Neutrophils / Q000201:enzymology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011145:Polyvinyls; D012995:Solubility; D013379:Substrate Specificity; D005465:Fluorocarbon Polymers
D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins; D010761:Phosphorus Radioisotopes; D011145:Polyvinyls; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers