Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Paper
  • Published:

A repertoire library that allows the selection of synthetic SH2s with altered binding specificities

Oncogene volume 20, pages 5186–5194 (2001)Cite this article

Abstract

Tyrosine phosphorylation is one of the major mechanisms involved in the intracellular propagation of external signals. Strategies aimed at interfering with this process might allow the control of several cellular phenotypes. SH2 domains mediate protein–protein interactions by recognizing phosphotyrosine (pY) residues in the context of specific phosphopeptides. We created an SH2-scaffolded repertoire library by randomly mutagenizing five critical amino acid positions in the specificity-determining region of the PLCγ C-terminal SH2 domain. Synthetic SH2 domains were selected from the library using biotinylated phosphopeptides derived from a natural PLCγ-SH2 ligand as well as unrelated SH2 ligands. The isolated SH2s displayed high binding affinity constants for the selecting peptides and were capable of interacting with the corresponding proteins.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3

Similar content being viewed by others

References

  • Arvidsson AK, Rupp E, Nanberg E, Downward J, Ronnstrand L, Wennstrom S, Schlessinger J, Heldin CH, Claesson-Welsh L . 1994 Mol. Cell Biol. 14: 6715–6726

  • Bradshaw JM, Mitaxov V, Waksman G . 2000 J. Mol. Biol. 299: 521–535

  • Breeze AL, Kara BV, Barratt DG, Anderson M, Smith JC, Luke RW, Best JR, Cartlidge SA . 1996 EMBO J. 15: 3579–3589

  • Carpenter G, Ji Q . 1999 Exp. Cell. Res. 253: 15–24

  • Eck MJ, Shoelson SE, Harrison SC . 1993 Nature 362: 87–91

  • Fazioli F, Minichiello L, Matoska V, Castagnino P, Miki T, Wong WT, Di Fiore PP . 1993 EMBO J 12: 3799–3808

  • Hatada MH, Lu X, Laird ER, Green J, Morgenstern JP, Lou M, Marr CS, Phillips TB, Ram MK, Theriault K, Zoller MJ, Karas JL . 1995 Nature 377: 32–38

  • Ikeda M, Arai K, Masai H . 1996 Gene 181: 167–171

  • Kraulis PJ . 1991 J. Appl. Crystallogr. 24: 946–950

  • Kuriyan J, Cowburn D . 1997 Annu. Rev. Biophys. Biomol. Struct. 26: 259–288

  • Larose L, Gish G, Shoelson S, Pawson T . 1993 Oncogene 8: 2493–2499

  • Lee CH, Kominos D, Jacques S, Margolis B, Schlessinger J, Shoelson SE, Kuriyan J . 1994 Structure 2: 423–438

  • Marengere LE, Songyang Z, Gish GD, Schaller MD, Parsons JT, Stern MJ, Cantley LC, Pawson T . 1994 Nature 369: 502–505

  • Merit EA, Murphy MEP . 1994 Acta Crystollogr. D 50: 869–873

  • Morton TA, Myszka DG . 1998 Methods Enzymol. 295: 268–294

  • Narula SS, Yuan RW, Adams SE, Green OM, Green J, Philips TB, Zydowsky LD, Botfield MC, Hatada M, Laird ER, Zoller Mj, Karas JL, Delgarno DC . 1995 Structure 3: 1061–1073

  • Ogura K, Tsuchiya S, Terasawa H, Yuzawa S, Hatanaka H, Mandiyan V, Schlessinger J, Inagaki F . 1999 J. Mol. Biol. 289: 439–445

  • Panayotou G, Gish G, End P, Truong O, Gout I, Dhand R, Fry MJ, Hiles I, Pawson T, Waterfield MD . 1993 Mol. Cell. Biol. 13: 3567–3576

  • Pascal SM, Singer AU, Gish G, Yamazaki T, Shoelson SE, Pawson T, Kay LE, Forman-Kay JD . 1994 Cell 77: 461–472

  • Rahuel J, Gay B, Erdmann D, Strauss A, Garcia-Echeverria C, Furet P, Caravatti G, Fretz H, Schoepfer J, Grutter MG . 1996 Nat. Struct. Biol. 3: 586–589

  • Salcini AE, McGlade J, Pelicci G, Nicoletti I, Pawson T, Pelicci PG . 1994 Oncogene 9: 2827–2836

  • Schneider S, Buchert M, Georgiev O, Catimel B, Halford M, Stacker SA, Baechi T, Moelling K, Hovens CM . 1999 Nat. Biotechnol. 17: 170–175

  • Shoelson SE . 1997 Curr. Opin. Chem. Biol. 1: 227–234

  • Songyang Z, Gish G, Mbamalu G, Pawson T, Cantley LC . 1995 J. Biol. Chem. 270: 26029–26032

  • Songyang Z, Shoelson SE, Chaudhuri M, Gish G, Pawson T, Haser WG, King F, Roberts T, Ratnofsky S, Lechleider RJ, Neel BG, Birge RB, Fajardo JE, Chou MM, Hanafusa H, Schaffhausen B, Cantley LC . 1993 Cell 72: 767–778

  • van der Geer P, Wiley S, Gish GD, Pawson T . 1996 Curr. Biol. 6: 1435–1444

  • Waksman G, Shoelson SE, Pant N, Cowburn D, Kuriyan J . 1993 Cell 72: 779–790

Download references

Acknowledgements

We thank Dr Pascale Romano for critically reviewing the manuscript. This work was supported by grants from Associazione Italiana Ricerca sul Cancro, from the Armenise-Harvard Foundation and from the CNR (Progetto Biotecnologie).

Author information

Author notes

  1. Maria Grazia Malabarba and Elisabetta Milia: MG Malabarba and E Milia contributed equally to this work

Authors and Affiliations

  1. Department of Experimental Oncology, European Institute of Oncology, Via Ripamonti 435, Milan, 20141, Italy

    Maria Grazia Malabarba, Elisabetta Milia, Mario Faretta, Raffaella Zamponi, Pier Giuseppe Pelicci & Pier Paolo Di Fiore

  2. IFOM, the FIRC Institute for Molecular Oncology, Via Serio 21, Milan, 20090, Italy

    Pier Paolo Di Fiore

Authors
  1. Maria Grazia Malabarba
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Elisabetta Milia
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mario Faretta
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Raffaella Zamponi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Pier Giuseppe Pelicci
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Pier Paolo Di Fiore
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Pier Paolo Di Fiore.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Malabarba, M., Milia, E., Faretta, M. et al. A repertoire library that allows the selection of synthetic SH2s with altered binding specificities. Oncogene 20, 5186–5194 (2001). https://doi.org/10.1038/sj.onc.1204654

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.onc.1204654

Keywords

This article is cited by

Search

Advanced search

Quick links