Identification and purification of a cytosolic phosphotyrosyl protein phosphatase from bovine spleen.
A protein tyrosine kinase with an apparent Mr of 60,000 was highly purified from bovine spleen and used to phosphorylate poly(Glu, Tyr) (4:1) on tyrosine residues for the study of phosphotyrosyl protein phosphatases from this tissue. About 70% of the phosphotyrosyl protein phosphatase activity in extracts of bovine spleen was adsorbed on DEAE-Sepharose. Chromatography of the eluted phosphotyrosyl protein phosphatases on phosphocellulose indicated the presence of at least two species, one that did not bind to the phosphocellulose and a second species that did bind and was eluted at about 0.5 M NaCl. The phosphatase that did not bind to phosphocellulose was further purified by successive chromatography on poly(L-lysine)-Sepharose, L-tyrosine-agarose, poly(Glu,Tyr)-Sepharose, and Sephacryl S-200. The enzyme had an apparent Mr of 50,000 as estimated by gel filtration and 52,000 as estimated by NaDodSO4- polyacrylamide gel electrophoresis. The phosphatase exhibited a pH optimum of 6.5-7.0, was inhibited by Zn2+ and vanadate ions, and was stimulated by EDTA. Sodium fluoride and sodium pyrophosphate, inhibitors of phosphoseryl protein phosphatases, had no effect on the enzyme. Protein inhibitors of type 1 phosphoseryl/threonyl phosphatase were also ineffective.
D000818:Animals; D002417:Cattle; D003600:Cytosol / Q000201:enzymology; D008970:Molecular Weight; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D013154:Spleen / Q000201:enzymology*
D010749:Phosphoprotein Phosphatases; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases