Logo of pnasPNASInfo for AuthorsSubscriptionsAboutThis Article
Proc Natl Acad Sci U S A. Aug 16, 1994; 91(17): 8132–8136.
PMCID: PMC44559

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.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.3M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Yarden Y, Ullrich A. Growth factor receptor tyrosine kinases. Annu Rev Biochem. 1988;57:443–478. [PubMed]
  • Ullrich A, Schlessinger J. Signal transduction by receptors with tyrosine kinase activity. Cell. 1990 Apr 20;61(2):203–212. [PubMed]
  • Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science. 1987 Jan 9;235(4785):177–182. [PubMed]
  • Slamon DJ, Godolphin W, Jones LA, Holt JA, Wong SG, Keith DE, Levin WJ, Stuart SG, Udove J, Ullrich A, et al. Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science. 1989 May 12;244(4905):707–712. [PubMed]
  • Plowman GD, Whitney GS, Neubauer MG, Green JM, McDonald VL, Todaro GJ, Shoyab M. Molecular cloning and expression of an additional epidermal growth factor receptor-related gene. Proc Natl Acad Sci U S A. 1990 Jul;87(13):4905–4909. [PMC free article] [PubMed]
  • Kraus MH, Issing W, Miki T, Popescu NC, Aaronson SA. Isolation and characterization of ERBB3, a third member of the ERBB/epidermal growth factor receptor family: evidence for overexpression in a subset of human mammary tumors. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9193–9197. [PMC free article] [PubMed]
  • Kraus MH, Fedi P, Starks V, Muraro R, Aaronson SA. Demonstration of ligand-dependent signaling by the erbB-3 tyrosine kinase and its constitutive activation in human breast tumor cells. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2900–2904. [PMC free article] [PubMed]
  • Hanks SK, Quinn AM, Hunter T. The protein kinase family: conserved features and deduced phylogeny of the catalytic domains. Science. 1988 Jul 1;241(4861):42–52. [PubMed]
  • Knighton DR, Zheng JH, Ten Eyck LF, Ashford VA, Xuong NH, Taylor SS, Sowadski JM. Crystal structure of the catalytic subunit of cyclic adenosine monophosphate-dependent protein kinase. Science. 1991 Jul 26;253(5018):407–414. [PubMed]
  • Gibbs CS, Zoller MJ. Rational scanning mutagenesis of a protein kinase identifies functional regions involved in catalysis and substrate interactions. J Biol Chem. 1991 May 15;266(14):8923–8931. [PubMed]
  • Moran MF, Koch CA, Sadowski I, Pawson T. Mutational analysis of a phosphotransfer motif essential for v-fps tyrosine kinase activity. Oncogene. 1988 Dec;3(6):665–672. [PubMed]
  • Tan JC, Nocka K, Ray P, Traktman P, Besmer P. The dominant W42 spotting phenotype results from a missense mutation in the c-kit receptor kinase. Science. 1990 Jan 12;247(4939):209–212. [PubMed]
  • Knighton DR, Cadena DL, Zheng J, Ten Eyck LF, Taylor SS, Sowadski JM, Gill GN. Structural features that specify tyrosine kinase activity deduced from homology modeling of the epidermal growth factor receptor. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5001–5005. [PMC free article] [PubMed]
  • Guy PM, Carraway KL, 3rd, Cerione RA. Biochemical comparisons of the normal and oncogenic forms of insect cell-expressed neu tyrosine kinases. J Biol Chem. 1992 Jul 15;267(20):13851–13856. [PubMed]
  • Anostario M, Jr, Harrison ML, Geahlen RL. Immunochemical detection of adenine nucleotide-binding proteins with antibodies to 5'-p-fluorosulfonylbenzoyladenosine. Anal Biochem. 1990 Oct;190(1):60–65. [PubMed]
  • Greenfield C, Patel G, Clark S, Jones N, Waterfield MD. Expression of the human EGF receptor with ligand-stimulatable kinase activity in insect cells using a baculovirus vector. EMBO J. 1988 Jan;7(1):139–146. [PMC free article] [PubMed]
  • Koland JG, Cerione RA. Growth factor control of epidermal growth factor receptor kinase activity via an intramolecular mechanism. J Biol Chem. 1988 Feb 15;263(5):2230–2237. [PubMed]
  • Koland JG, Cerione RA. Activation of the EGF receptor tyrosine kinase by divalent metal ions: comparison of holoreceptor and isolated kinase domain properties. Biochim Biophys Acta. 1990 May 22;1052(3):489–498. [PubMed]
  • Buhrow SA, Cohen S, Staros JV. Affinity labeling of the protein kinase associated with the epidermal growth factor receptor in membrane vesicles from A431 cells. J Biol Chem. 1982 Apr 25;257(8):4019–4022. [PubMed]
  • Songyang Z, Shoelson SE, Chaudhuri M, Gish G, Pawson T, Haser WG, King F, Roberts T, Ratnofsky S, Lechleider RJ, et al. SH2 domains recognize specific phosphopeptide sequences. Cell. 1993 Mar 12;72(5):767–778. [PubMed]
  • Carraway KL, 3rd, Sliwkowski MX, Akita R, Platko JV, Guy PM, Nuijens A, Diamonti AJ, Vandlen RL, Cantley LC, Cerione RA. The erbB3 gene product is a receptor for heregulin. J Biol Chem. 1994 May 13;269(19):14303–14306. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Koland JG, O'Brien KM, Cerione RA. Expression of epidermal growth factor receptor sequences as E. coli fusion proteins: applications in the study of tyrosine kinase function. Biochem Biophys Res Commun. 1990 Jan 15;166(1):90–100. [PubMed]
  • Soltoff SP, Carraway KL, 3rd, Prigent SA, Gullick WG, Cantley LC. ErbB3 is involved in activation of phosphatidylinositol 3-kinase by epidermal growth factor. Mol Cell Biol. 1994 Jun;14(6):3550–3558. [PMC free article] [PubMed]
  • Sliwkowski MX, Schaefer G, Akita RW, Lofgren JA, Fitzpatrick VD, Nuijens A, Fendly BM, Cerione RA, Vandlen RL, Carraway KL., 3rd Coexpression of erbB2 and erbB3 proteins reconstitutes a high affinity receptor for heregulin. J Biol Chem. 1994 May 20;269(20):14661–14665. [PubMed]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Compound
    Compound
    PubChem Compound links
  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...