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Computational design of an integrin I domain stabilized in the open high affinity conformation

Nature Structural Biology volume 7, pages 674–678 (2000)Cite this article

Abstract

We have taken a computational approach to design mutations that stabilize a large protein domain of ∼200 residues in two alternative conformations. Mutations in the hydrophobic core of the αMβ2 integrin I domain were designed to stabilize the crystallographically defined open or closed conformers. When expressed on the cell surface as part of the intact heterodimeric receptor, binding of the designed open and closed I domains to the ligand iC3b, a form of the complement component C3, was either increased or decreased, respectively, compared to wild type. Moreover, when expressed in isolation from other integrin domains using an artificial transmembrane domain, designed open I domains were active in ligand binding, whereas designed closed and wild type I domains were inactive. Comparison to a human expert designed open mutant showed that the computationally designed mutants are far more active. Thus, computational design can be used to stabilize a molecule in a desired conformation, and conformational change in the I domain is physiologically relevant to regulation of ligand binding.

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Figure 1: Location of mutations in the Mac-1 I domain open structure.
Figure 2: Intact Mac-1 molecules with computationally designed I domains are more active than wild type in binding ligands when transiently expressed in 293T cells.
Figure 3: αMβ2 heterodimers with computationally designed open and closed I domains are active in binding iC3b, and resistant to activation, respectively, when stably expressed in K562 cells.
Figure 4: Isolated computationally designed open I domain mutants bind ligands, whereas wild type and designed closed I domains do not.

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References

  1. Springer, T.A. Nature 346, 425–433 (1990).

    Article  CAS  Google Scholar 

  2. Michishita, M., Videm, V. & Arnaout, M.A. Cell 72, 857–867 (1993).

    Article  CAS  Google Scholar 

  3. Diamond, M.S., Garcia-Aguilar, J., Bickford, J.K., Corbi, A.L. & Springer, T.A. J. Cell Biol. 120, 1031–1043 (1993).

    Article  CAS  Google Scholar 

  4. Lee, J.-O., Bankston, L.A., Arnaout, M.A. & Liddington, R.C. Structure 3, 1333–1340 (1995).

    Article  CAS  Google Scholar 

  5. Lee, J.-O., Rieu, P., Arnaout, M.A. & Liddington, R. Cell 80, 631–638 (1995).

    Article  CAS  Google Scholar 

  6. Qu, A. & Leahy, D.J. Proc. Natl. Acad. Sci. USA 92, 10277–10281 (1995).

    Article  CAS  Google Scholar 

  7. Qu, A. & Leahy, D.J. Structure 4, 931–942 (1996).

    Article  CAS  Google Scholar 

  8. Emsley, J., King, S.L., Bergelson, J.M. & Liddington, R.C. J. Biol. Chem. 272, 28512–28517 (1997).

    Article  CAS  Google Scholar 

  9. Baldwin, E.T. et al. Structure 6, 923–935 (1998).

    Article  CAS  Google Scholar 

  10. Nolte, M. et al. FEBS Lett. 452, 379–385 (1999).

    Article  CAS  Google Scholar 

  11. Rich, R.L. et al. J. Biol. Chem. 274, 24906–24913 (1999).

    Article  CAS  Google Scholar 

  12. Huang, C. & Springer, T.A. J. Biol. Chem. 270, 19008–19016 (1995).

    Article  CAS  Google Scholar 

  13. Li, R., Rieu, P., Griffith, D.L., Scott, D. & Arnaout, M.A. J. Cell Biol. 143, 1523–1534 (1998).

    Article  CAS  Google Scholar 

  14. Zhang, L. & Plow, E.F. Biochemistry 38, 8064–8071 (1999).

    Article  CAS  Google Scholar 

  15. Emsley, J., Knight, C.G., Farndale, R.W., Barnes, M.J. & Liddington, R.C. Cell 101, 47–56 (2000).

    Article  CAS  Google Scholar 

  16. Springer, T.A. Proc. Natl. Acad. Sci. USA 94, 65–72 (1997).

    Article  CAS  Google Scholar 

  17. Oxvig, C., Lu, C. & Springer, T.A. Proc. Natl. Acad. Sci. USA 96, 2215–2220 (1999).

    Article  CAS  Google Scholar 

  18. Perutz, M.F. Q. Rev. Biophys. 22, 139–237 (1989).

    Article  CAS  Google Scholar 

  19. Dahiyat, B.I. & Mayo, S.L. Science 278, 82–87 (1997).

    Article  CAS  Google Scholar 

  20. Malakauskas, S.M. & Mayo, S.L. Nature Struct. Biol. 5, 470–475 (1998).

    Article  CAS  Google Scholar 

  21. Street, A.G. & Mayo, S.L. Structure 7, R105–R109 (1999).

    Article  CAS  Google Scholar 

  22. Gordon, D.B., Marshall, S.A. & Mayo, S.L. Curr. Opin. Struct. Biol. 9, 509–513 (1999).

    Article  CAS  Google Scholar 

  23. Lasters, I., De Maeyer, M. & Desmet, J. Protein Eng. 8, 815–822 (1995).

    Article  CAS  Google Scholar 

  24. Harbury, P.B., Plecs, J.J., Tidor, B., Alber, T. & Kim, P.S. Science 282, 1462–1467 (1998).

    Article  CAS  Google Scholar 

  25. Oxvig, C. & Springer, T.A. Proc. Natl. Acad. Sci. USA 95, 4870–4875 (1998).

    Article  CAS  Google Scholar 

  26. Lu, C., Oxvig, C. & Springer, T.A. J. Biol. Chem. 273, 15138–15147 (1998).

    Article  CAS  Google Scholar 

  27. Diamond, M.S. & Springer, T.A. J. Cell Biol. 120, 545–556 (1993).

    Article  CAS  Google Scholar 

  28. Lu, C. & Springer, T.A. J. Immunol. 159, 268–278 (1997).

    CAS  Google Scholar 

  29. Petruzzelli, L., Maduzia, L. & Springer, T. J. Immunol. 155, 854–866 (1995).

    CAS  Google Scholar 

  30. Humphries, M.J. & Newham, P. Trends Cell Biol. 8, 78–83 (1998).

    Article  CAS  Google Scholar 

  31. Loftus, J.C. & Liddington, R.C. J. Clin. Invest. 99, 2302–2306 (1997).

    Article  CAS  Google Scholar 

  32. Knorr, R. & Dustin, M.L. J. Exp. Med. 186, 719–730 (1997).

    Article  CAS  Google Scholar 

  33. Dahiyat, B.I. & Mayo, S.L. Proc. Natl. Acad. Sci. USA 94, 10172–10177 (1997).

    Article  CAS  Google Scholar 

  34. Dill, K.A. Biochemistry 29, 7133–7155 (1990).

    Article  CAS  Google Scholar 

  35. Dahiyat, B.I. & Mayo, S.L. Protein Sci. 5, 895–903 (1996).

    Article  CAS  Google Scholar 

  36. Dahiyat, B.I., Gordon, D.B. & Mayo, S.L. Protein Sci. 6, 1333–1337 (1997).

    Article  CAS  Google Scholar 

  37. Street, A.G. & Mayo, S.L. Fold. Des. 3, 253–258 (1998).

    Article  CAS  Google Scholar 

  38. Casimiro, D.R., Toy-Palmer, A., Blake, R.C.d. & Dyson, H.J. Biochemistry 34, 6640–6648 (1995).

    Article  CAS  Google Scholar 

  39. Prodromou, C. & Pearl, L.H. Protein Eng. 5, 827–829 (1992).

    Article  CAS  Google Scholar 

  40. Kishimoto, T.K., O'Connor, K., Lee, A., Roberts, T.M. & Springer, T.A. Cell 48, 681–690 (1987).

    Article  CAS  Google Scholar 

  41. Kleywegt, G.J. & Jones, T.A. Acta Crystallogr. D 50, 178–185 (1994).

    Article  CAS  Google Scholar 

  42. Carson, M. Methods Enzymol. 277, 493–505 (1997).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank M. Ferzly for technical assistance. This work was supported by the NIH.

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Authors and Affiliations

  1. The Center for Blood Research and Department of Pathology, Harvard Medical School, 200 Longwood Avenue, Boston, 02115, Massachusetts, USA

    Motomu Shimaoka, Hua Jing, Junichi Takagi & Timothy A. Springer

  2. Howard Hughes Medical Institute and Division of Biology, California Institute of Technology, Pasadena, mail-code 147-75, 91125, California, USA

    Julia M. Shifman & Stephen L. Mayo

Authors
  1. Motomu Shimaoka
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  2. Julia M. Shifman
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  3. Hua Jing
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  4. Junichi Takagi
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  5. Stephen L. Mayo
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Correspondence to Stephen L. Mayo or Timothy A. Springer.

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Shimaoka, M., Shifman, J., Jing, H. et al. Computational design of an integrin I domain stabilized in the open high affinity conformation. Nat Struct Mol Biol 7, 674–678 (2000). https://doi.org/10.1038/77978

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