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.

  • Review Article
  • Published:

Genetic Approaches to Protein Structure and Function: Point Mutations as Modifiers of Protein Function

Bio/Technology volume 10, pages 1002–1006 (1992)Cite this article

Abstract

In this review, I summarize data in the biological literature which underscore the utility of a genetic approach to protein structure/function problems, with emphasis on binding phenomena, particularly of cytokine and growth factor/receptor interactions. Useful parallels or contrasts to chemical ligand/receptor systems and DNA binding protein interactions are examined where they simplify the analysis of protein ligand/receptor interactions. This approach was prompted by the fact that purely rational approaches, based on resolution of the three dimensional structure of proteins, are limited because such data is available for fewer than 3% of the 17,000 proteins for which the amino acid sequence has been deduced by molecular biology techniques.

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

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to the full article PDF.

USD 39.95

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

Similar content being viewed by others

References

  1. Zurawski, S.M., lmler, J.-L and Zurawski, G. 1990. Partial agonist/antagonist mouse interleukin 2 proteins indicate that a 3rd component of the receptor complex functions in signal transduction. EMBO J. 9: 3899–3955.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Labriola-Tomkins, E., Chandron, C., Kaffka, K.L., Biondi, D., Graves, B.J., Hatada, M., Madison, V.S., Karas, J., Killian, P.L. and Ju, G. 1991. Identification of the discontinuous binding site in human interleukin 1 beta for type 1 interleukin 1 receptor. Proc. Nat. Acad. Sci. USA 8: 11182–11186.

    Article  Google Scholar 

  3. Ju, G., Labriola-Tompkins, E., Campen, C., Benjamin, W.R., Karas, J., Plocinski, J., Biondi, D., Killian, P.L., Eisenberg, S.P. and Evans, J. 1991. Conversion of the IL1 receptor antagonist into an agonist by site specific mutagenesis. Proc. Nat. Acad. Sci. USA 88: 2652–2662.

    Article  Google Scholar 

  4. Matzuk, M., Keene, J.L. and Boime, I. 1989. Site specificity of the chorionic gonadotropin N-linked oligosaccharides in signal transduction. J. Biol. Chem. 264: 2409–2414.

    CAS  PubMed  Google Scholar 

  5. Douglas, W.W. 1985. Polypeptides—Angiotensin, plasma kinins, and others, p. 647. In: The Pharmacological Basis of Experimental Therapeutics, 7th ed. Goodman, L.S., Gilman, A.G., Rall, T. and Murad, F. (Eds.). Macmillan Publishing Co., NY.

    Google Scholar 

  6. Roussel, M.F., Downing, J.R. and Sherr, C.J. 1990. Transforming activities of human CSF receptors with different point mutations at codon 301 in the extracellular domain. Ontogene 5: 25–30.

    CAS  Google Scholar 

  7. Yoshimura, A., Longmore, G. and Lodish, H.F. 1990. Point mutation in the exoplasmic domain of the erythropoietin receptor resulting in hormone independent activation and tumorigenicity. Nature 348: 647–649.

    Article  CAS  PubMed  Google Scholar 

  8. Bargmann, C.I., Hung, M.-C. and Weinberg, R.A. 1986. Multiple independent activations of the neu oncogene by a point mutation altering the transmembrane domain of p185. Cell 45: 649–657.

    Article  CAS  PubMed  Google Scholar 

  9. Kjelsberg, M.A., Coteccia, S., Ostrowski, J., Caron, M.G. and Lefkowitz, R.J. 1992. Constitutive activation of the α1B adrenergic receptor by all amino acid substitutions at a single site. Evidence for a region which constrains receptor activation. J. Biol. Chem. 267: 1430–1433.

    CAS  PubMed  Google Scholar 

  10. Revah, F., Bertrand, D., Galzi, J.L., Devillers-Thiery, A., Mulle, C., Hussy, N., Bertrand, S., Ballivet, M. and Changeux, J.P. 1991. Mutations in the channel domain alter desensitization of a neuronal nicotinic receptor. Nature 353: 846–849.

    Article  CAS  PubMed  Google Scholar 

  11. Kleina, L. and Miller, J. 1990. Genetic studies of the lac represser XIII: Extensive amino acid replacement generated by the use of natural and synthetic nonsense suppressors. J. Mol. Biol. 212: 295–318.

    Article  CAS  PubMed  Google Scholar 

  12. Kelly, R.J. and Yanofsky, C. 1985. Mutational studies within the trp repressor of Escherichia coli support the helix-turn-helix model of repressor recognition of operator DNA. Proc. Nat. Acad. Sci. 82: 483–487.

    Article  Google Scholar 

  13. Benhamon, B., Garcia, T., Lerouge, T., Vergezac, A., Gufflo, D., Bigogne, C., Chambon, P. and Gronenmeyer, H. 1992. A single amino acid determines the sensitivity of the progesterone receptor to RU486. Science 255: 206–209.

    Article  Google Scholar 

  14. Hall, B.G. and Zuzel, T. 1980. Evolution of a new enzymatic function by recombination within a gene. Proc. Nat. Acad. Sci. USA 77: 3529–3533.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Hall, B.G., Betts, P.W. and Wooton, J.C. 1989. DNA sequence analysis of artificially evolved ebg enzyme and ebg repressor genes. Genetics 123: 635–648.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Lau, F. and Fersht, A. 1987. Conversion of allosteric inhibition to activation in phosphofructokinase by protein engineering. Nature 326: 811–812.

    Article  CAS  PubMed  Google Scholar 

  17. Patthy, L. 1985. Evolution of the proteases of blood coagulation and fibrinolysis by assembly from modules. Cell 41: 657–663.

    Article  CAS  PubMed  Google Scholar 

  18. Baron, M., Norman, D.G. and Campbell, I.G. 1991. Protein modules. Trends in Bio. Sci. 16: 13–17.

    Article  CAS  Google Scholar 

  19. Eisenberg, S.P., Brewer, M.T., Verderber, E., Heimdal, P., Brandhuber, B.J. and Thompson, R.C. 1991. Interleukin 1 receptor antagonist is a member of the interleukin 1 gene family: evolution of a cytokine control mechanism. Proc. Nat. Acad. Sci. USA 88: 5232–5236.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Gehrke, L., Jobling, S.A., Park, L.S., McDonald, B., Rossenwasser, L.J. and Auron, P.E. 1990. A point mutation uncouples human interleukin 1β biological activity and receptor binding. J. Biol. Ghem. 265: 5922–5925.

    CAS  Google Scholar 

  21. Fane, B. and King, J. 1991. Intragenic suppressors of folding defects in the P22 tailspike protein. Genetics 127: 263–277.

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Hinrichsen, R., Pollock, M., Hennessey, T. and Russell, C. 1991. An intra-genie suppressor of a calmodulin mutation in paramecium: genetic and biochemical characterization. Genetics 129: 717–725.

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Brandhuber, B.J., Boone, T., Kenney, W.C. and McKay, D.B. 1987. Three dimensional structure of IL2. Science 238: 1707–1709.

    Article  CAS  PubMed  Google Scholar 

  24. Ultsch, M., deVos, A.M. and KossikofT, A.A. 1991. Crystals of the complex between human growth hormone and the extracellular domain of the receptor. J. Mol. Biol. 222: 865–868.

    Article  CAS  PubMed  Google Scholar 

  25. Jaffe, J.H. and Martin, W.R. 1985. Opioid analgesics and antagonists, p. 494. In: The Pharmacological Basis of Experimental Therapeutics, 7th ed. op. cit

    Google Scholar 

  26. Snider, R., Constantine, J., Lowe, J. III, Longo, K., Lebel, W., Woody, H., Drozda, S., Desai, M., Vinick, E., Spencer, R. and Hess, H. 1991. A potent nonpeptide antagonist of the substance P (NK1) receptor. Science 251: 435–437.

    Article  CAS  PubMed  Google Scholar 

  27. Green, F.R., Lynch, B., and Kaiser, E.T. 1987. Biological and physical properties of a model calcitonin containing a glutamate residue interupting the hydrophobic face of the idealized amphiphilic α-helical region. Proc. Nat. Acad. Sci. USA 84: 8340–8344.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Dripps, D., Brandhuber, B.J., Thompson, R.C., and Eisenberg, S. 1991. Interleukin 1 (1L1) receptor antagonist binds to the 80KDa IL1 receptor but does not initiate 1L1 signal transduction. J. Biol. Chem. 266: 10331–10336.

    CAS  PubMed  Google Scholar 

  29. Cohen, J.B. 1978. Ligand binding properties of membrane bound cholinergic receptors of Torpedo marmorata, p. 99–128. In: Molecular Specialization and Symmetry in Membrane Function. Soloman, A.K. and Karnovsky, M. (Eds.). Harvard Univ. Press, Cambridge, MA.

    Google Scholar 

  30. Neubig, R.R. and Cohen, J.B. 1979. Equilibrium binding of [3H] tubo-curarine and [3H]-acetylcholine by Torpedo postsynaptic membranes: stoichiometry and ligand interactions. Biochemistry 18: 5464–5475.

    Article  CAS  PubMed  Google Scholar 

  31. Smee, D.F., Boehme, R., Chernow, M., Binko, B.P. and Matthews, T.R. 1985. Intracellular metabolism and enzymatic phosphorylation of 9- (1,3 dihydroxy-2-propoxymethyl) guanine and acyclovir in herpes simplex virus infected and uninfected cells. Biochemical Pharmacology 34: 1049–1056.

    Article  CAS  PubMed  Google Scholar 

  32. Ling, Z.D., Gillis, S., Hart, L.J. and Matheson, D.D. 1991. Particle concentration fluorescence immunoassay for measuring IL6 receptor numbers. Cytokine 3: 17–20.

    Article  CAS  PubMed  Google Scholar 

  33. King, K., Dohlman, H.G., Thorner, J., Caron, M.G. and Lefkowitz, R.J. 1990. Control of yeast mating signal transduction by a β2 adrenergic receptor and Gsα subunit. Science 250: 121–123.

    Article  CAS  PubMed  Google Scholar 

  34. Arkin, A.P. and Youvan, D.C. 1992. Optimizing nucleotide mixtures to encode specific subsets of amino acids for semi-random mutagenesis. Bio/Technology 10: 297–300.

    CAS  Google Scholar 

  35. Kunkel, T., Roberts, J. and Zakour, R.A. 1987. Rapid and efficient site specific mutagenesis without phenotypic selection. Meth. Enz. 154: 367–382.

    Article  CAS  Google Scholar 

  36. Sayers, J.R., Schmidt, W. and Eckstein, F. 1988. 5′–3′ exonuclease in phosphothioale-based oligonucleotide directed mutagenesis. Nucl. Acids Res. 16: 791–802.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Vandeyar, M., Weiner, M., Hutton, C. and Batt, C. 1988. A simple and rapid method for the selection of oligonucleotide directed mutants. Gene 65: 129–133.

    Article  CAS  PubMed  Google Scholar 

  38. Normanly, J., Masson, J.-M., Kleina, L.G., Abelson, J. and Miller, J.H. 1986. Construction of two Escherichia coli amber suppressor genes: tRNAPhe and tRNACys . Proc. Nat. Acad. Sci. USA 83: 6548–6552.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Kleina, L., Masson, J., Normanly, J., Abelson, J. and Miller, J.H. 1990. Construction of E. coli amber suppressor tRNA genes. II. Synthesis of additional tRNA genes and improvement of suppressor efficiency. J. Mol. Biol. 213: 705–717.

    Article  CAS  PubMed  Google Scholar 

  40. Normanly, J., Kleina, L., Masson, J., Abelson, J. and Miller, J.H. 1990. Construction of amber suppressor tRNA genes. III. Determination of tRNA specificity. J. Mol. Biol. 213: 719–726.

    Article  CAS  PubMed  Google Scholar 

  41. Medynski, D. and Coffino, P. 1989. Rapid generation and identification of multiple mutants of ornithine decarboxylase utilizing a colony color screen and tRNA amber nonsense suppressors. Amer. Soc. of Microbiol. Biotechnology Conf. 2: 12.

    Google Scholar 

  42. Ellman, J.A., Mandel, D. and Schultz, P.G. 1992. Site specific incorporation of novel backbone structures into proteins. Science 255: 197–200.

    Article  CAS  PubMed  Google Scholar 

  43. Bass, S., Greene, R. and Wells, J. 1990. Hormone phage: An enrichment method for variant proteins with altered binding properties. Proteins 8: 309–314.

    Article  CAS  PubMed  Google Scholar 

  44. Nagai, K. and Thogerson, H.C. 1984. Generation of β globulin by sequence-specific proteolysis of a hybrid protein produced in E. coli. Nature 309: 810–812.

    Article  CAS  PubMed  Google Scholar 

  45. Cwirla, P., Barrett, W. and Dower, W.J. 1990. Peptides on phage: A vast library of peptides identifying ligands. Proc. Nat. Acad. Sci. USA 87: 6378–6382.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. McCafferty, J., Griffiths, R.D., Winter, G. and Chiswell, D.J. 1990. Phage antibodies: filamentous phage displaying antibody variable domains. Nature 348: 552–554.

    Article  CAS  PubMed  Google Scholar 

  47. Yamasaki, K., Taga T., Hirata, Y., Yawata, H., Kawasaki, Y., Seed, B., Taniguchi, T., Hirano.T. and Kishimoto, T. 1988. Cloning and expression of the human interleukin 6 receptor. Science 241: 825–828.

    Article  CAS  PubMed  Google Scholar 

  48. Maliszewski, C.R., Sato, T.A., Van den Bos, T., Waugh, S., Dower, S.K., Slack, J., Beckman, M. and Grabstein, K. 1990. Cytokine receptors and B cell function: Recombinant soluble receptors specifically inhibit IL-1 and IL-4 induced B cell activities. J. Immunology 19: 3028–3033.

    Google Scholar 

  49. Goodwin, R.G., Friend, D., Ziegler, S.F., Jerzy, R., Falk, B.A., Gumpel, S., Cosman, D., Dower, S.K., March, C.J. and Namen, A.E. 1990. Cloning of the human and murine 1L7 receptors: demonstration of a soluble form and homology to a new receptor superfamily. Cell 60: 941–951.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Author notes

  1. Dan Medynski: Address correspondence to the author at 2040 Polk St. Ste. ♯204, San Francisco, CA 94109.

Authors and Affiliations

  1. Department of Pathology, Harvard Medical School, Boston, MA, 02115

    Dan Medynski

  2. Dana Farber Cancer Institute, 44 Binney Street, Boston, MA, 02115

    Dan Medynski

Authors
  1. Dan Medynski
    View author publications

    Search author on:PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Medynski, D. Genetic Approaches to Protein Structure and Function: Point Mutations as Modifiers of Protein Function. Nat Biotechnol 10, 1002–1006 (1992). https://doi.org/10.1038/nbt0992-1002

Download citation

  • Issue date:

  • DOI: https://doi.org/10.1038/nbt0992-1002

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