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Structure of a Numb PTB domain–peptide complex suggests a basis for diverse binding specificity

Nature Structural Biology volume 5, pages 1075–1083 (1998)Cite this article

Abstract

The phosphotyrosine-binding (PTB) domain of Numb, a protein involved in asymmetric cell division, has recently been shown to bind to the adapter protein Lnx through an LDNPAY sequence, to the Numb-associated kinase (Nak) through a sequence that does not contain an NPXY motif and to GP(p)Y-containing peptides obtained from library screening. We show here that these diverse peptide sequences bind with comparable affinities to the Numb PTB domain at a common binding site on the surface of the protein. The NMR structure of the Numb PTB domain in complex with a GPpY-containing peptide reveals a novel mechanism of binding with the peptide in a helical turn that does not hydrogen bond to the PTB domain β-sheet. These results suggest that PTB domains can potentially have multiple modes of peptide recognition and provide a structural basis from which the multiple functions of the Numb PTB domain during asymmetric cell division could arise.

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Figure 1: Competition binding data for unlabeled peptides with a complex of the dNumb PTB domain and fluorescein-labeled Nak peptide.
Figure 2: Solution structure of the dNumb PTB domain–GPpY peptide complex.
Figure 3: Structure-based alignment34 of sequences of the dNumb, X1114, Shc8 and IRS-110 PTB domains, with residues of the dNumb PTB domain highlighted in green for helices and yellow for β-strands.
Figure 4: a–d, Surface representations (using MOLMOL47) of PTB domains in complex with target peptides (in yellow), with blue representing positive and red negative electrostatic potential.
Figure 5: a, 15N-1H correlation spectrum of the dNumb PTB domain–GPpY peptide complex.

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Acknowledgements

We thank Y. N. Jan for kindly providing the Drosophila Numb cDNA. This work was supported by grants to J.D.F.-K. and L.E.K. from the National Cancer Institute of Canada (NCIC) and to T.P. from the Human Frontier Science Program, Asashi Chemical Company, the NCIC, and the Medical Research Council of Canada (MRC) and by Howard Hughes International Research Scholar awards to T.P. and L.E.K. T.P. is a Terry Fox Cancer Research Scientist of the NCIC. S.-C.L. is a post-doctoral fellow of the MRC. C.Z. and S.J.F.V. are recipients of Human Frontier Science Program fellowships.

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Author notes

  1. Sébastien J. F. Vincent

    Present address: Nestlé Research Center, Vers-chez-les-Blanc, CH-1000, Lausanne, 26, Switzerland

  2. Shun-Cheng Li and Catherine Zwahlen: These authors contributed equally to the work.

Authors and Affiliations

  1. Program in Molecular Biology and Cancer, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, M5G 1X5, Ontario

    Shun-Cheng Li, Sébastien J. F. Vincent & Tony Pawson

  2. Department of Molecular and Medical Genetics, University of Toronto, Toronto, M5S 1A8, Ontario, Canada

    Shun-Cheng Li, Sébastien J. F. Vincent & Tony Pawson

  3. Structural Biology and Biochemistry, Hospital for Sick Children, Toronto, M5G 1X8, Ontario, Canada

    Catherine Zwahlen & Julie D. Forman-Kay

  4. Department of Biochemistry, University of Toronto, Toronto, M5S 1A8, Ontario, Canada

    Catherine Zwahlen & Julie D. Forman-Kay

  5. Protein Engineering Network Centres of Excellence and Departments of Chemistry, Biochemistry, and Molecular and Medical Genetics, University of Toronto, Toronto, M5S 1A8, Ontario, Canada

    Catherine Zwahlen, Sébastien J. F. Vincent & Lewis E. Kay

  6. Amgen Institute, 620 University Avenue, Toronto, M5G 2C1, Ontario, Canada

    C. Jane McGlade

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Correspondence to Julie D. Forman-Kay.

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Li, SC., Zwahlen, C., Vincent, S. et al. Structure of a Numb PTB domain–peptide complex suggests a basis for diverse binding specificity. Nat Struct Mol Biol 5, 1075–1083 (1998). https://doi.org/10.1038/4185

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