title: Insect cell-expressed p180erbB3 possesses an impaired tyrosine kinase activity.
abstract: Protein kinases share a number of highly conserved or invariant amino acid residues in their catalytic domains, suggesting that these residues are necessary for kinase activity. In p180erbB3, a receptor tyrosine kinase belonging to the epidermal growth factor (EGF) receptor subfamily, three of these residues are altered, suggesting that this protein might have an impaired protein tyrosine kinase activity. To test this hypothesis, we have expressed human EGF receptor and bovine p180erbB3 in insect cells via baculovirus infection and have compared their autophosphorylation and substrate phosphorylation activities. We have found that, while the EGF receptor readily undergoes EGF-stimulated autophosphorylation and catalyzes the incorporation of phosphate into the model substrates (E4Y1)n (random 4:1 copolymer of glutamic acid and tyrosine) and GST-p85 (glutathione S-transferase fusion protein with the 85-kDa subunit of phosphatidylinositol 3-kinase), p180erbB3 autophosphorylation and substrate phosphorylation are at least 2 orders of magnitude less efficient. However, p180erbB3 is capable of binding the ATP analog 5'-p-fluorosulfonylbenzoyladenosine, indicating that the lack of observed kinase activity is probably not due to nonfunctional or denatured receptors expressed by the insect cells. On the basis of these results, we propose that p180erbB3 possesses an impaired intrinsic tyrosine kinase activity.
authors.affiliation: Department of Pharmacology, Cornell University, Ithaca, NY 14853.
authors.forename: P M
authors.initials: PM
authors.lastname: Guy
authors.forename: J V
authors.initials: JV
authors.lastname: Platko
authors.forename: L C
authors.initials: LC
authors.lastname: Cantley
authors.forename: R A
authors.initials: RA
authors.lastname: Cerione
authors.forename: K L
authors.initials: KL
authors.lastname: Carraway
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007313:Insecta; D007700:Kinetics; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D020893:Receptor, ErbB-3; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D004815:Epidermal Growth Factor; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D066246:ErbB Receptors; D020893:Receptor, ErbB-3