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.

  • Research
  • Published:

Soluble Forms of the Human T Cell Receptor CD4 are Efficiently Expressed by Streptomyces lividans

Bio/Technology volume 11, pages 1031–1036 (1993)Cite this article

Abstract

We have developed a new gene expression and secretion system for Streptomyces lividans and used it to produce soluble forms of a human T-cell receptor CD4 at levels greater than 300 mg/l. The system uses the transcription, translation and secretion signals of the serine protease inhibitor gene STI-II which is naturally produced by S. longisporus. Using these signals, soluble derivatives of CD4 were secreted directly into the culture supernatant as correctly processed soluble, biologically active proteins. High level expression of the CD4 proteins depended on the transcription initiation signal, the amino acid sequence surrounding the signal peptide cleavage site and temporally controlled protease activities. We discuss these results in the context of the potential of this system for producing other eukaryotic proteins in Streptomyces.

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

Similar content being viewed by others

References

  1. Brawner, M., Poste, G., Rosenberg, M. and Westpheling, J. 1991. Streptomyces: a host for heterologous gene expression. Current Opinion in Biotechnology 2: 674–681.

    Article  CAS  PubMed  Google Scholar 

  2. Engels, J.W. and Koller, K.-P. 1992. Gene expression and secretion of eukaryotic foreign proteins in Streptomyces, p. 31–53. In: Transgenesis. Murray, J. A. H. (Ed.). John Wiley & Sons Ltd, NY.

    Google Scholar 

  3. Gay, D., Maddon, P., Sekely, R., Talle, M.A., Godfrey, M., Long, E., Goldstein, G., Chess, L., Axel, R., Kappler, J. and Marrack, J. 1987. Functional interaction between human T-cell protein CD4 and the major histocom-patibility complex HLADR antigen. Nature 328: 626–629.

    Article  CAS  PubMed  Google Scholar 

  4. Sleckman, B.P., Peterson, A., Jones, W.K., Foran, J.A., Greenstein, J.L., Seed, B. and Burakoff, S.J. 1987. Expression and function of CD4 in a murine T-cell hybridoma. Nature 328: 351–353.

    Article  CAS  PubMed  Google Scholar 

  5. Doyle, C. and Strominger, J.L. 1987. Interaction between CD4 and class II MHC molecules mediates cell adhesion. Nature 330: 256–259.

    Article  CAS  PubMed  Google Scholar 

  6. Arthos, J., Deen, K.C., Chaikin, M.A., Fornwald, J.A., Sathe, G., Sattentau, Q.J., Clapham, P.R., Weiss, R.A., McDougal, J.S., Pietropaolo, C., Axel, R., Truneh, A., Maddon, P.J. and Sweet, R.W. 1989. Identification of the residues in human CD4 critical for the binding of HIV. Cell 57: 69–181

    Article  Google Scholar 

  7. Garlick, R.L., Kirschner, R.J., Eckenrade, F.M., Tarpley, W.G. and Tomich, C.-S.C. 1990. Escherchia coli expression, purification and biological activity of a truncated soluble CD4. AIDS Res. and Hum. Retroviruses 6: 65–79.

    Article  Google Scholar 

  8. Smith, D.H., Byrn, R.A., Marsters, S.A., Gregory, T., Groopman, J.E. and Capon, D.J. 1987. Blocking of HIV-1 infectivity by a soluble, secreted form of the CD4 antigen. Science 238: 1704–1707.

    Article  CAS  PubMed  Google Scholar 

  9. Fisher, R.A., Bertonis, J.M., Meier, W., Johnson, V.A., Costopoulos, D.S., Liu, T., Tizard, R., Walker, B.D., Hirsch, M.S., Schooley, R.T. and Flavell, R.A. 1988. HIV infection is blocked in vitro by recombinant soluble CD4. Nature 331: 76–78.

    Article  CAS  PubMed  Google Scholar 

  10. Hussey, R.E., Richardson, N.E., Kowalski, M., Brown, N.R., Chang, H.-C., Siliciano, R.R., Dorman, T., Walker, B., Sodroski, J. and Reinherz, E.L. 1988. A soluble CD4 protein selectively inhibits HIV replication and syncytium formation. Nature 331: 78–81.

    Article  CAS  PubMed  Google Scholar 

  11. Deen, K.C., McDougal, J.S., Inacker, R., Folena-Wasserman, G., Arthos, J., Rosenberg, J., Maddon, P.J., Axel, R. and Sweet, R.W. 1988. A soluble form of CD4 (T4) protein inhibits AIDS virus infection. Nature 331: 82–84.

    Article  CAS  PubMed  Google Scholar 

  12. Strickler, J.E., Berka, T.R., Gorniak, J., Rosenberg, J., Keys, R., Rowland, J.J., Rosenberg, M. and Taylor, D.P. 1992. Two novel Slreptomyces protein protease inhibitors. J. Biol. Chem. 267: 3236–3241.

    CAS  PubMed  Google Scholar 

  13. Brawner, M., Fornwald, J., Rosenberg, M., Poste, G. and Westpheling, J. 1990. Heterologous gene expression in Streptomyces, p. 85–93. In: Proceedings of the 6th International Symposium on Genetics of Industrial Microorganisms. Heslot, H., Davies, J., Florent, J., Bobichon, L., Durand, G. and Penasse, L. (Eds.). Société Francaise de Microbiologie, Strasbourg, France.

    Google Scholar 

  14. Li, P., Beckwith, J. and Inouye, H. 1988. Alteration of the amino terminus of the mature sequence of a periplasmic protein can severely affect protein export in Escherichia coli. Proc. Natl. Acad. Sci. USA 85: 7685–7689.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Summers, R.G., Harris, C.R. and Knowles, J.R. 1989. A conservative amino acid substitution, argimne for lysme, abolishes export of a hybrid protein in Escherichia coli. J. Biol. Chem. 264: 20082–20088.

    CAS  PubMed  Google Scholar 

  16. Liss, L.R., Johnson, B.L. and Oliver, D.B. 1985. Export defect adjacent to the processing site of staphylococcal nuclease is suppressed by a prlA mutation. J. Bacteriol. 164: 925–928.

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Yamane, K. and Mizushima, S. 1988. Introduction of basic amino acid residues after the signal peptide inhibits protein translocation across the cytoplasmic membrane of Escherichia coli. J. Biol. Chem. 263: 19690–19696.

    CAS  PubMed  Google Scholar 

  18. Eckhardt, T., Strickler, J., Gorniak, L., Burnett, W.V. and Fare, L.J. 1987. Characterization of the promoter, signal sequence and amino acid terminus of a secreted β-galactosidase from Streptomyces lividans. J. Bacteriol. 169: 4249–4256.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Classon, B.J., Tsagaratos, J., Kirszbaum, L., Maddox, J., Mackay, C.R., Brandon, M., McKenzie, I.F.C. and Walker, I.D. 1986. The L3T4 antigen in mouse and the sheep equivalent are immunoglobulin like. Immunogenetics 23: 129–132.

    Article  CAS  PubMed  Google Scholar 

  20. Peterson, A. and Seed, B. 1988. Genetic analysis of monoclonal antibody and HIV binding sites on the human lymphocyte antigen CD4. Cell 54: 65–72.

    Article  CAS  PubMed  Google Scholar 

  21. Sattentau, Q.J., Dalgleish, A.G., Weiss, R.A. and Beverly, P.C.L. 1986. Epitopes of the CD4 antigen and HIV infection. Science 234: 1120–1123.

    Article  CAS  PubMed  Google Scholar 

  22. Dalgleish, A.G., Beverly, P.C.L., Clapham, P.R., Crawford, D.H., Greaves, M.F. and Weiss, R.A. 1984. The CD4 (T4) antigen is an essential component of the receptor for the AIDS retrovirus. Nature 312: 763–766.

    Article  CAS  PubMed  Google Scholar 

  23. Klatzman, D., Champagne, E., Chamaret, S., Gruest, J., Guetard, P., Hercend, T., Gluckman, J.-C. and Montagnier, L. 1984. T-lymphocyte T4 molecule behaves as the receptor for human retrovirus LAV. Nature 312: 767–768.

    Article  Google Scholar 

  24. Maddon, P.J., Dalgleish, A.G., McDougal, J.S., Clapham, P.R., Weiss, R.A. and Axel, R. 1986. The T4 gene encodes the AIDS virus receptor and is expressed in the immune system and the brain. Cell 47: 333–348.

    Article  CAS  PubMed  Google Scholar 

  25. McDougal, J.S., Kennedy, M.S., Sligh, J.M., Cort, S.P., Mawle, A. and Nicholson, J.K.A. 1986. Binding of HTLV-III/LAV to T4+ T cells by a complex of the 110K viral protein and the T4 molecule. Science 231: 382–385.

    Article  CAS  PubMed  Google Scholar 

  26. Watt, R.A., Shatzman, A.R. and Rosenberg, M. 1985. Expression and characterization of the human c-myc DNA binding protein. Mol. Cell. Biol. 5: 448–456.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Buell, G., Schulz, M.-F., Selzer, G., Chollet, A., Movva, N.R., Semon, D., Escanez, S. and Kawashima, E. 1985. Optimizing the expression in E. coli of a synthetic gene encoding somatomedin-C (IGF-I). Nuc. Acids. Res. 13: 1923–1938.

    Article  CAS  Google Scholar 

  28. Sloma, A., Rufo, G.A., Theriault, K.A., Dwyer, M., Wilson, S.W. and Pero, J. 1991. Cloning and chracterization of the gene for an additional extracellular serine protease of Bacillus subtilis. J. Bacteriol. 173: 6889–6895.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Staunton, D.E., Marlin, S.D., Stratowa, C., Dustin, M.L. and Springer, T.A. 1988. Primary structure of ICAM-1 demonstrates interaction between members of the immunoglobulin and integrin supergene families. Cell 52: 925–933.

    Article  CAS  PubMed  Google Scholar 

  30. Bevilacqua, M.P., Stengelin, S., Gimbrone, M.A. and Seed, B. 1989. Endotheial leukocyte adhesion molecule: an inducible receptor for neutrophils related to complement regulatory proteins and lectins. Science 243: 1160–1165.

    Article  CAS  PubMed  Google Scholar 

  31. Maniatis, T., Fritsch, E.F. and Sambrook, J. 1982. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.

    Google Scholar 

  32. Yanisch-Perron, C., Vieira, J. and Messing, J. 1985. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33: 103–119.

    Article  CAS  PubMed  Google Scholar 

  33. Katz, E., Thompson, C.J. and Hopwood, D.A. 1983. Cloning and expression of the tyrosinase gene from Streptomyces antibioticus in Streptomyces lividans. J. Gen. Microbiol. 129: 2703–2714.

    CAS  PubMed  Google Scholar 

  34. Kieser, T., Hopwood, D.A., Wright, H.M. and Thompson, C.J. 1982. pIJ101, a multi-copy broad host-range Streptomyces plasmid:functional analysis and development of DNA cloning vectors. Mol. Gen. Genet. 185: 223–238.

    Article  CAS  PubMed  Google Scholar 

  35. Kunkel, T.A. 1985. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc. Natl. Acad. Sci. USA 82: 488–492.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Littman, D.R., Maddon, P.J. and Axel, R. 1988. Corrected CD4 sequence. Cell 55: 541.

    Article  CAS  PubMed  Google Scholar 

  37. Bibb, M.J., Ward, J.M., Kieser, T., Cohen, S.N. and Hopwood, D.A. 1981. Excision of chromosomal DNA sequences from Streptomyces coeticolor forms a novel family of plasmids detectable in Streptomyces lividans. Mol. Gen. Genet. 184: 230–240.

    CAS  PubMed  Google Scholar 

  38. Thompson, C.J., Ward, J.M. and Hopwood, D.A. 1982. Cloning of antibiotic resistance and nutritional genes in streptomycetes. J. Bacteriol. 151: 668–677.

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Laemmli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.

    Article  CAS  PubMed  Google Scholar 

  40. Towbin, H., Staehelin, T. and Gordon, J. 1979. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets:procedure and some applications. Proc. Natl. Acad. Sci. USA 76: 4350–4354.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Brawner, M.E., Auerbach, J.I., Fornwald, J.A., Rosenberg, M. and Taylor, D.P. 1985. Characterization of Streptomyces promoter sequences using the Escherichia coli galactokinase gene. Gene 40: 191–201.

    Article  CAS  PubMed  Google Scholar 

  42. Matsudaira, P. 1987. Sequences from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J. Biol. Chem. 262: 10035–10038.

    CAS  PubMed  Google Scholar 

  43. Thompson, C.J., Ward, J.M. and Hopwood, D.A. 1980. DNA cloning in Streptomyces: resistance genes from antibiotic-producing species. Nature 286: 525–527.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Author notes

  1. Mary Jo Donovan

    Present address: Bristol-Myers Squibb, Microbial Technology, New Brunswick, NJ, 08903

  2. John Keller

    Present address: SmithKline Beecham Pharmaceuticals, Four Falls Corporate Center, P.O. Box 1510, King of Prussia, PA, 19406

  3. Dean P. Taylor

    Present address: PanLabs Inc., 11804 North Creek Pkwy South, Bothell, WA, 98011

  4. Edward J. Arcuri

    Present address: Merck & Co. Inc., P. O. Box 4, WP 28-79, West Point, PA

  5. Mary E. Brawner: Corresponding author.

Authors and Affiliations

  1. SmithKline Beecham Pharmaceuticals, King of Prussia, PA, 19406

    James A. Fornwald, Mary Jo Donovan, Robert Gerber, John Keller, Dean P. Taylor, Edward J. Arcuri & Mary E. Brawner

  2. Gene Expression Sciences, King of Prussia, PA, 19406

    James A. Fornwald, Mary Jo Donovan, Dean P. Taylor, Edward J. Arcuri & Mary E. Brawner

  3. Biological Process Sciences, King of Prussia, PA, 19406

    Robert Gerber & John Keller

Authors
  1. James A. Fornwald
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mary Jo Donovan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Robert Gerber
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. John Keller
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Dean P. Taylor
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Edward J. Arcuri
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Mary E. Brawner
    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

Fornwald, J., Donovan, M., Gerber, R. et al. Soluble Forms of the Human T Cell Receptor CD4 are Efficiently Expressed by Streptomyces lividans. Nat Biotechnol 11, 1031–1036 (1993). https://doi.org/10.1038/nbt0993-1031

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0993-1031

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