Purified human platelet-derived growth factor receptor has ligand-stimulated tyrosine kinase activity.
The platelet-derived growth factor receptor (PDGF-R), a 180-kDa single-chain polypeptide, was purified from membranes of the human osteogenic sarcoma cell line MG-63. Purification was achieved by treatment of membranes with PDGF and ATP, followed by solubilization with nonionic detergent and successive chromatography on solid-phase anti-phosphotyrosine monoclonal antibody and DEAE-cellulose. The PDGF-R, which was estimated to be 50-80% pure by NaDodSO4/polyacrylamide gel electrophoresis of 32P-labeled preparations, was free of contaminating epidermal growth factor receptor and had no detectable phosphatase activity. It specifically bound 125I-labeled PDGF, a reaction quantified by binding of the ligand-PDGF-R complex to the anti-phosphotyrosine antibody. The purified receptor displayed PDGF-stimulatable tyrosine kinase activity, assayed by autophosphorylation of PDGF-R at tyrosine residues and by phosphorylation of angiotensin II. The Km for ATP in the autophosphorylation reaction was 7.5 microM. Addition of PDGF did not change the Km but increased the Vmax 1.7-fold.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000596:Amino Acids / Q000032:analysis; D000804:Angiotensin II / Q000378:metabolism; D006801:Humans; D008024:Ligands; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000032:analysis*; D011956:Receptors, Cell Surface / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D017479:Receptors, Platelet-Derived Growth Factor
D000596:Amino Acids; D008024:Ligands; D010982:Platelet-Derived Growth Factor; D011956:Receptors, Cell Surface; D000804:Angiotensin II; D000255:Adenosine Triphosphate; D011505:Protein-Tyrosine Kinases; D017479:Receptors, Platelet-Derived Growth Factor