Comparative characterization of membrane-associated and cytosolic Tyr-protein kinases in human erythrocytes.
In recent years, two protein-tyrosine kinase activities, phosphorylating tyrosine residues on the transmembrane band-3 protein, have been isolated from human erythrocyte membranes and partially characterized by different laboratories, i.e. one extracted by non-ionic detergent (Triton X-100 or Nonidet P-40), the other solubilized by 0.25 M NaCl from the detergent-insoluble residue. The present paper shows that these two membrane-associated Tyr-protein kinases purified, in the presence of bovine serum albumin, by phosphocellulose chromatography followed by heparin-Sepharose chromatography, have the same apparent molecular mass (36 kDa) determined by Ultrogel Ac44 filtration. Moreover, both Tyr-protein kinases exhibit several identical properties, including Km values for band 3, the random acidic copolymer poly(Glu,Tyr)4:1 and angiotensin II, pH dependence, response to Mn2+ and Mg2+, response to NaCl and 2,3-bisphosphoglycerate. All these properties are identical or very similar to those exhibited by the Tyr-protein kinase previously isolated by us from human erythrocyte cytosol. These results suggest that the two membrane-associated and the cytosolic Tyr-protein kinase activities are mediated by the same enzyme, distributed between the cytosol and the membrane structures.
Clari
G
G
Istituto di Chimica, Biologica dell'Università di Verona, Italy.
Moret
V
V
D002845:Chromatography / Q000379:methods; D003600:Cytosol / Q000201:enzymology*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D004910:Erythrocyte Membrane / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000032:analysis* / Q000037:antagonists & inhibitors
D008345:Manganese; D011505:Protein-Tyrosine Kinases; D008274:Magnesium