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 Article
  • Published:

The similarities of bar and pat gene products make them equally applicable for plant engineers

Nature Biotechnology volume 14, pages 1274–1278 (1996)Cite this article

Abstract

The bar and pat genes, isolated from different Streptomyces species, both encode a phosphinothricin acetyltransferase (PAT) and are widely applied in plant genetic engineering. The genes were expressed in Escherichia coli and the corresponding proteins were purified and used for functional and structural comparison. Both proteins are homodimers regardless of whether they are expressed in microorganisms or in plants. They have comparable molecular weights and show immuno–cross-reactivity to their respective polyclonal antisera. The enzymes have a similar substrate affinity towards L-phosphinothricin and do not acetylate any of the other L-amino acids tested. In model digestion experiments using simulated human gastric fluids, their enzymatic activity is decreased within seconds, accompanied by a rapid and complete breakdown of both proteins. These data demonstrate the structural and functional equivalence of the PAT proteins, which is also reflected in the comparable performance of transgenic plants carrying the bar or pat gene.

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. De Block, M., Botterman, J., Vandewiele, M., Dockx, J., Thoen, C., Gosselé, V., et al. 1987. Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J. 6: 2513–2518.

    Article  CAS  Google Scholar 

  2. Wohlleben, W., Arnold, W., Broer, I., Hillemann, D., Strauch, E., and Pühler, A. 1988. Nucleotide sequence of the phosphinothricin N-acetyltransferase gene from Streptomyces Tü494 and its expression in Nicotiana tabacum . Gene 70: 25–37.

    Article  CAS  Google Scholar 

  3. Thompson, C.J., Movva, N.R., Tichard, R., Crameri, R., Davies, J.E., Lauwereys, M., et al. 1987. Characterization of the herbicide-resistance gene bar from Streptomyces hygroscopicus . EMBO J. 6: 2519–2523.

    Article  CAS  Google Scholar 

  4. Wild, A., Sauer, H., and Rühle, W. 1987. The effect of phosphinothricin (glufosinate) on photosynthesis. I. Inhibition of photosynthesis and accumulation of ammonia. Z. Naturforsch. 42c: 263–269.

    Article  Google Scholar 

  5. Sauer, H., Wild, A., and Rühle, W. 1987. The effect of phosphinothricin (glufosinate) on photosynthesis. II. The causes of inhibition of photosynthesis. Z. Naturforsch. 42c: 270–278.

    Article  Google Scholar 

  6. Givan, C.V., Joy, K.W., and Kleczkowski, L.A. 1988. A decade of photorespiratory nitrogen cycling. TIBS 13: 433–437.

    CAS  PubMed  Google Scholar 

  7. Bayer, E., Gugel, K.H., Hägele, K., Hagenmaier, H., Jessipow, S., König, W.A., et al. 1972. Stoffwechselprodukte von Mikroorganismen. Phosphinothricin und Phosphinothricyl-alanyl-alanin. Helv. Chim. Acta 55: 224–239.

    Article  CAS  Google Scholar 

  8. Kondo, Y., Shomura, T., Ogawa, Y., Tsuruoka, T., Watanabe, H., Totukawa, K., et al. 1973. Studies on a new antibiotic SF-1293.1. Isolation and physico-chemical and biological characterization of SF-1293 substances. Sci. Rep. Meiji Seika 13: 34–41.

    Google Scholar 

  9. Seto, H., Imai, S., Tsuruoka, T., Satoh, A., Kojima, M., Inouye, S., et al. 1982. Studies on the biosynthesis of bialaphos (SF-1293). 1. Incorporation of 13C- and 2H-labeled precursors into bialaphos. J. Antibiot 35: 1719–1721.

    Article  CAS  Google Scholar 

  10. Strauch, E., Wohlleben, W., and Pühler, A. 1988. Cloning of a phosphinothricin N-acetyltransferase gene from Streptomyces viridochromogenes Tü494 and its expression in Streptomyces lividans and Escherichia coli . Gene 63: 65–74.

    Article  CAS  Google Scholar 

  11. Vasil, I.K. 1994. Molecular improvement of cereals. Plant Mol. Biol. 25: 925–937.

    Article  CAS  Google Scholar 

  12. Shen, W.-H. and Hohn, B. 1995. Vectors based on maize streak virus can replicate to high copy numbers in maize plants. J. Gen. Virol. 76: 965–969.

    Article  CAS  Google Scholar 

  13. Botterman, J., Gosselé, V., Thoen, C., and Lauwereys, M. 1991. Characterization of phosphinothricin acetyltransferase and C-terminal enzymatically active fusion proteins. Gene 102: 33–37.

    Article  CAS  Google Scholar 

  14. De Greef, W., Delon, R., De Block, M., Leemans, J., and Botterman, J. 1989. Evaluation of herbicide resistance in transgenic crops under field conditions. Bio/Technology 7: 61–64.

    CAS  Google Scholar 

  15. D'Halluin, K., De Block, M., Denecke, J., Janssens, J., Leemans, J., Reynaerts, A., et al. 1992. The bar gene as selectable and screenable marker in plant engineering. Methods Enzymol. 216: 415–426.

    Article  CAS  Google Scholar 

  16. Rasche, E., Cremer, J., Donn, G., and Zink, J. 1995. The development of glufos-inate ammonium tolerant crops into the market. Brit. Crop Prot Conf. Weeds 3: 791–800.

    Google Scholar 

  17. Chater, K.F. and Hopwood, D.A. 1993. 6. Streptomyces, pp. 83–90 in Bacillus subtilis and other gram-positive bacteria. Sonenshein, A.L. (ed.). American Society for Microbiology, Washington, D.C.

    Google Scholar 

  18. Logusch, E.W., Walker, D.M., McDonald, J.F., Franz, J.E., Villafranca, J.J., Dilanni, C.L., et al. 1990. Inhibition of Escherichia coli glutamine synthetase by a- and g-substituted phosphinothricins. Biochemistry 29: 366–372.

    Article  CAS  Google Scholar 

  19. Acaster, M.A. and Weitzman, R.D.J. 1985. Kinetic analysis of glutamine syn-thetases from various plants. FEBS Lett. 189: 241–244.

    Article  CAS  Google Scholar 

  20. Logusch, E.W., Walker, D.M., McDonald, J.F., and Franz, J.E. 1991. Inhibition of plant glutamine synthetases by substituted phosphinothricins. Plant Physiol. 95: 1057–1062.

    Article  CAS  Google Scholar 

  21. Buttner, M.J., Chater, K.F., and Bibb, M.J. 1990. Cloning, disruption, and trans-lational analysis of three RNA polymerase sigma factor genes of Streptomyces coelicolor A3(2). J. Bacteriol. 172: 3367–3378.

    Article  CAS  Google Scholar 

  22. Marcos, A.T., Gutiérrez, S., Diez, B., Fernández, F.J., Oguiza, J.A., and Martin, J.F. 1995. Three genes hrdB, hrdD, and hrdT of Streptomyces griseus IMRU 3570, encoding sigma factor-like proteins, are differentially expressed under specific nutritional conditions. Gene 153: 41–48.

    Article  CAS  Google Scholar 

  23. Murakami, T., Anzai, H., Imai, S., Satoh, A., Nagaoka, K., and Thompson C.J. 1986. The bialaphos biosynthetic genes of Streptomyces hygroscopicus: molecular cloning and characterization of the gene cluster. Mol. Gen. Genet. 205: 42–50.

    Article  CAS  Google Scholar 

  24. Wohlleben, W., Alijah, R., Dorendorf, J., Hillemann, D., Nussbaumer, B., and Pelzer, S. 1992. Identification and characterization of phosphinothricin-tripeptide biosynthetic genes in Streptomyces viridochromogenes . Gene 115: 127–132.

    Article  CAS  Google Scholar 

  25. Raibaud, A., Zalacain, M., Holt, T.G., Tizard, R., and Thompson, C.J. 1991. Nucleotide sequence analysis reveals linked N-acetyl hydrolase, thioesterase, transport, and regulatory genes encoded by the bialaphos biosynthetic gene cluster of Streptomyces hygroscopicus . J. Bacteriol. 173: 4454–4463.

    Article  CAS  Google Scholar 

  26. Bedford, D.J., Lewis, C.G., and Buttner, M.J. 1991. Characterization of a gene conferring bialaphos resistance in Streptomyces coelicolor A3(2). Gene 104: 39–45.

    Article  CAS  Google Scholar 

  27. Taylor, S.L., Lemanske, R.F., Jr., Bush, R.K., and Busse, W.W. 1987. Food allergens: structure and immunologic properties. Ann. Allergy 59: 93–99.

    CAS  PubMed  Google Scholar 

  28. Sambrook, J.J., Fritsch, E.F., and Maniatis, T. 1989. Molecular cloning: A laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

    Google Scholar 

  29. Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of micro-gram quantities of protein utilizing the principle of protein dye binding. Anal. Biochem. 72: 248–254.

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  31. Merril, C.R. 1990. Gel-staining techniques. In: Guide to protein purification. Deutscher, M.P. (ed.). Methods Enzymol. 182: 477–488.

    Chapter  Google Scholar 

  32. Blum, H., Beier, H., and Gross, H.J. 1987. Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis 8: 93–99.

    Article  CAS  Google Scholar 

  33. The United States Pharmacopeia, Vol. XXIII. 1995. United States Pharmacopeial Convention, Inc. Rockville, MD. 2053.

  34. Chou, P.Y. and Fasman, G.D. 1978. Prediction of secondary structure of proteins from amino acid sequence. Adv. Enzymol. Relat. Subj. Biochem. 47: 45–148.

    CAS  Google Scholar 

  35. Rost, B. and Sander, D. 1993. Improved prediction of protein secondary structure by use of sequence profiles and neural networks. Proc. Natl. Acad. Sci. USA

Download references

Author information

Author notes

  1. Arno Schulz: Corresponding author (e-mail: arno.schulz@agrevo.com)

Authors and Affiliations

  1. Hoecht Schering AgrEvo GmbH, Forschung Biochemie, D-65926, Frankfurt am Main, Germany

    Axel Wehrmann & Arno Schulz

  2. Forschungszentrum Jülich GmbH, Institut für Biotechnologie 1, D-52425, Jülich, Germany

    Axel Wehrmann

  3. Plant Genetic Systems, N. V., JozefPlateaustraat 22, B-9000, Gent, Belgium

    Adri Van Vliet, Chris Opsomer & Johan Botterman

Authors
  1. Axel Wehrmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Adri Van Vliet
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Chris Opsomer
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Johan Botterman
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Arno Schulz
    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

Wehrmann, A., Vliet, A., Opsomer, C. et al. The similarities of bar and pat gene products make them equally applicable for plant engineers. Nat Biotechnol 14, 1274–1278 (1996). https://doi.org/10.1038/nbt1096-1274

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt1096-1274

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