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.

  • Article
  • Published:

A test of the role of the proximal histidines in the Perutz model for cooperativity in haemoglobin

Nature Structural Biology volume 4, pages 78–83 (1997)Cite this article

Abstract

Human haemoglobin has long been a paradigm for cooperative ligand binding and allostery. Through analysis of the crystal structures of deoxyhaemoglobin and liganded haemoglobin, Perutz proposed a model for cooperativity in which the bond between the proximal histidine and the protein couples haem rearrangements to protein structure rearrangements. Here we test this model by deleting the bonds between the histidine imidazole side chain and the polypeptide. This detachment method allows us to determine directly the contribution of proximal histidine coupling to cooperativity of distal ligand binding. Proximal detachment significantly increases ligand affinity, reduces cooperativity, and prevents quaternary structure switching, in accord with the Perutz model. Residual cooperativity indicates that additional haem communication pathways exist that do not involve the proximal histidine coupling mechanism.

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

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

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

Similar content being viewed by others

References

  1. Wyman, J. & Gill, S.J. Binding and Linkage: Functional Chemistry of Biological Macromolecules. University Science Books, Mill Valley CA (1990).

    Google Scholar 

  2. Perutz, M.F. Mechanisms of cooperativity and allosteric regulation in proteins. Cambridge University Press, Cambridge England (1990).

    Google Scholar 

  3. Bohr, C. Zentralbl. Physiol. 17, 682 (1903).

    Google Scholar 

  4. Hill, A.V. The possible effects of aggregation of the molecules of haemoglobin on its dissociation curve. J. Physiol. 40, iv–vii (1910).

    Google Scholar 

  5. Antonini, E. & Brunori, M. Hemoglobin and Myoglobin in Their Reactions With Ligands, North-Holland Publishing Co., Amsterdam (1971).

    Google Scholar 

  6. Imai, K. Allosteric effects in haemoglobin, Cambridge University Press, New York (1982).

    Google Scholar 

  7. Cullis, A.F., Muirhead, H., Perutz, M.F. & Rossmann, M.G. Proc. Roy. Soc. A 265, 161–187 (1962).

    CAS  Google Scholar 

  8. Perutz, M.F., Muirhead, H., Cox, J.M. & Goaman, L.C.G. Three-dimensional fourier synthesis of horse oxyhaemoglobin at 2.8 Å resolution: the atomic model. Nature 219, 131–139 (1968).

    Article  CAS  Google Scholar 

  9. Dickerson, R.E. & Geis, I. Hemoglobin. The Benjamin/Cummings Publishing Company, Inc., Menlo Park, California (1983).

    Google Scholar 

  10. Perutz, M.F. Stereochemistry of cooperative effects in haemoglobin. Nature 228, 726–734 (1970).

    Article  CAS  Google Scholar 

  11. Perutz, M.F. Hemoglobin Structure and Respiratory Transport. Sci. Amer. 239, 92–125 (1978).

    Article  CAS  Google Scholar 

  12. Warshel, A. Energy-structure correlation in metalloporphyrins and the control of oxygen binding in hemoglobin. Proc. Natl. Acad. Sci. USA 74, 1789–1793 (1977).

    Article  CAS  Google Scholar 

  13. Gelin, B.R. & Karplus, M. Mechanism of tertiary structural change in hemoglobin. Proc. Natl. Acad. Sci. USA 74, 801–805 (1977).

    Article  CAS  Google Scholar 

  14. Baldwin, J. & Chothia, C. Hemoglobin: The structural changes related to ligand binding and its allosteric mechanism. J. Mol. Biol. 129, 175–220 (1979).

    Article  CAS  Google Scholar 

  15. Vuk-Pavlovic, S. Evolution of the haem-haem interaction in vertebrate haemoglobins-a hypothesis. J. Mol. Evol. 6, 209–214 (1975).

    Article  CAS  Google Scholar 

  16. Barrick, D. Replacement of the proximal ligand of sperm whale myoglobin with free imidazole in the mutant His93→Gly. Biochemistry 33, 6546–6554 (1994)

    Article  CAS  Google Scholar 

  17. DePillis, G.D., Decatur, S.M., Barrick, D. & Boxer, S.G. Functional cavities in proteins. A general method for proximal ligand substitution in myoglobin. J. Am. Chem. Soc. 116, 6981–6982 (1994).

    Article  CAS  Google Scholar 

  18. Lu, Y., Casmiro, D.R., Bren, K.L., Richards, J.H. & Gray, H.B. Structurally engineered cytochromes with unusual ligand-binding properties: expression of Saccharomyces cerevisiae Met-80→Ala iso-1-cytochrome c. Proc. Natl. Acad. Sci. USA 90, 11456–11459 (1993).

    Article  CAS  Google Scholar 

  19. McRee, D.E., Jesen, G.M., Fitzgerald, M.M., Siegel, H.A. & Goodin, D.B. Construction of a bisaquo heme enzyme and binding by exogenous ligands. Proc. Natl. Acad. Sci. USA 91, 12847–12851 (1994).

    Article  CAS  Google Scholar 

  20. Wilks, A., Sun, J., Loehr, T.M. & Ortiz de Montellano, P.R. Heme oxygenase His25Ala mutant: replacement of the proximal histidine iron ligand by exogenous bases restores the catalytic activity. J. Am. Chem. Soc. 117, 2925–2926 (1995).

    Article  CAS  Google Scholar 

  21. Newmyer, S., Sun, J., Loehr, T.M. & Ortiz de Montellano, P.R. Rescue of the Horseradish Peroxidase His-170→Ala Mutant Activity by Imidazole: Importance of Proximal Ligand Tethering. Biochemistry 35, 12788–12795 (1996).

    Article  CAS  Google Scholar 

  22. Barrick, D. Depletion and replacement of protein metal ligands. Curr. Op. Biotechnology 6, 411–418 (1995).

    Article  CAS  Google Scholar 

  23. St. George, R.C.C. & Pauling, L. The combining power of hemoglobin for alkyl isocyanides, and the nature of the heme-heme interactions in hemoglobin. Science 114, 629–634 (1951).

    Article  Google Scholar 

  24. Reisberg, P.I. & Olson, J.S. Equilibrium binding of alkyl isocyanides to human hemoglobin. J. Biol. Chem 255, 4144–4150 (1980).

    CAS  PubMed  Google Scholar 

  25. Reisberg, P.I. & Olson, J.S. Kinetic and cooperative mechanisms of ligand binding to hemoglobin. J. Biol. Chem 255, 4159–4169 (1980).

    CAS  PubMed  Google Scholar 

  26. Mims, M.P. et al. Proton nuclear magnetic resonance studies of isonitrile-heme protein complexes. J. Biol. Chem. 258, 6125–6134 (1983).

    CAS  PubMed  Google Scholar 

  27. Dahlquist, F.W. The meaning of Scatchard and Hill plots. Meth. Enzymol. 48, 270–299 (1978).

    Article  CAS  Google Scholar 

  28. Pulsinelli, P.D., Perutz, M.F. & Nagel, R.L. Structure of Hemoglobin M Boston, a Variant with a Five-Coordinated Ferric Heme. Proc. Acad. Sci. USA 70, 3870–3874 (1973).

    Article  CAS  Google Scholar 

  29. Takahashi, S., Lin, A.K.-L.C. & Ho, C. Proton nuclear magnetic resonacne studies of hemoglobins M. Boston (α58E7 His→Tyr) and M Milwaukee (β67E11 Val→Glu): spectral assignments of hyperfine-shifted proton resonances and of proximal histidine (E7) NH resonances to the α and β chains of normal human adult hemoglobin. Biochemistry 19, 5196–5202 (1980).

    Article  CAS  Google Scholar 

  30. Monod, J., Wyman, J. & Changeux, J.-P. On the nature of allosteric transitions: a plausible model. J. Mol. Biol. 12, 88–118 (1965).

    Article  CAS  Google Scholar 

  31. Koshland, D.E. Jr., Nemethy, G. & Filmer, D.L. Comparison of experimental and theoretical models in proteins containing subunits. Biochemistry 5, 365–385 (1966).

    Article  CAS  Google Scholar 

  32. Wyman, J. Linked functions and reciprocal effects in hemoglobin: a second look. Adv. Prot. Chem. 19, 223–286 (1964).

    CAS  Google Scholar 

  33. Olson, J.S. & Gibson, Q.H. The functional properties of hemoglobin Bethesda (α2β2145His) J. Biol. Chem. 247, 3662–3670 (1972).

    CAS  PubMed  Google Scholar 

  34. Arnone, A. & Perutz, M.F. Structure of inositol hexaphosphate-human deoxyhaemoglobin complex. Nature 249, 34–36 (1974).

    Article  CAS  Google Scholar 

  35. Ho, C. Proton NMR studies on hemoglobin: cooperative interactions and partially ligated intermediates Adv. Prot. Chem. 43, 153–312 (1992).

    CAS  Google Scholar 

  36. Stryer, L. Biochemistry 4th ed., W.H. Freeman & Co., New York (1995).

    Google Scholar 

  37. Voet, D. & Voet, J.G. Biochemistry 2nd ed., John Wiley & Sons, Inc., New York (1995).

    Google Scholar 

  38. Rodgers, K.R. & Spiro, T.G. Nanosecond dynamics of the R→T transition in hemoglobin: ultraviolet Raman studies. Science 265, 1697–1698 (1994).

    Article  CAS  Google Scholar 

  39. Pauling, L. The nature of the chemical bond, 3rd ed., Cornell University Press, 1960.

    Google Scholar 

  40. Ackers, G.K., Doyle, M.L., Myers, D. & Daugherty, M.A. The molecular code for cooperativity in hemoglobin. Science 255, 54–63 (1992).

    Article  CAS  Google Scholar 

  41. Paoli, M., Liddington, R., Tame, J., Wilkinson, A. & Dodson, G. Crystal structure of T state haemoglobin with oxygen bound at all four haems. J. Mol. Biol. 256, 775–792 (1996).

    Article  CAS  Google Scholar 

  42. Viggiano, G. & Ho, C. Proton nuclear magnetic resonance investigations of structural changes associated with cooperative oxygenation of human adult hemoglobin. Proc. Natl. Acad. Sci. USA 76, 3673–3677 (1979).

    Article  CAS  Google Scholar 

  43. Friedman, J.M. Time-resolved resonance Raman spectroscopy as a probe of structure, dynamics, and reactivity in hemoglobin. Meth. Enzymol. 232, 205–231 (1994).

    Article  CAS  Google Scholar 

  44. Liddington, R. X-ray Crystallography of partially liganded structures. Meth. Enzymol. 232, 15–26 (1994).

    Article  CAS  Google Scholar 

  45. Kunkel, T.M. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc. Natl. Acad. Sci. USA 82, 488–492 (1985).

    Article  CAS  Google Scholar 

  46. Shen, T.-J. et al. Production of unmodified human adult hemoglobin in Escherichia coli. Proc. Natl. Acad. Sci. USA 90, 8108–8112 (1993).

    Article  CAS  Google Scholar 

  47. Hayashi, A., Suzuki, T. & Shin, M. An enzyme reduction system for metmyoglobin and methemoglobin, and its application to functional studies of oxygen carriers. Biochem. Biophys. Acta 310, 309–316 (1973).

    CAS  PubMed  Google Scholar 

  48. Johnson, M.L. & Frasier, S.G. Nonlinear least-squares analysis. Meth. Enzymol. 117, 301–342 (1985).

    Article  CAS  Google Scholar 

  49. Plateau, P. & Guéron, M. Exchangeable proton NMR without baseline distortion, using new strong pulse sequences. J. Amer. Chem. Soc. 104, 7310–7311 (1982).

    Article  CAS  Google Scholar 

Download references

Author information

Author notes

  1. Doug Barrick

    Present address: Department of Biophysics & Biophysical Chemistry, School of Medicine, Johns Hopkms University, 725 N. Wolfe St., Baltimore, Maryland, 21205, USA

Authors and Affiliations

  1. Institute of Molecular Biology, University of Oregon, Eugene, Oregon, 97403, USA

    Doug Barrick & Frederick W. Dahlquist

  2. Department of Biological Science, Carnegie Mellon University, 4400 Fifth Avenue, Pittsburgh, Pennsylvania, 15213, USA

    Nancy T. Ho, Virgil Simplaceanu & Chien Ho

Authors
  1. Doug Barrick
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nancy T. Ho
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Virgil Simplaceanu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Frederick W. Dahlquist
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chien Ho
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Barrick, D., Ho, N., Simplaceanu, V. et al. A test of the role of the proximal histidines in the Perutz model for cooperativity in haemoglobin. Nat Struct Mol Biol 4, 78–83 (1997). https://doi.org/10.1038/nsb0197-78

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsb0197-78

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing