Skip to main content
SpringerLink
Log in

Anticodon and acceptor stem nucleotides in tRNAGln are major recognition elements for E. coli glutaminyl-tRNA synthetase

  • Letter
  • Published:

From Nature

View current issue Submit your manuscript

Abstract

THE correct attachment of amino acids to their corresponding (cognate) transfer RNA catalysed by aminoacyl-tRNA synthetases is a key factor in ensuring the fidelity of protein biosynthesis. Previous studies have demonstrated that the interaction of Escherichia coli tRNAGln with glutaminyl-tRNA synthetase (GlnRS) provides an excellent system1 to study this highly specific recognition process, also referred to as 'tRNA identity'2. Accurate acylation of tRNA depends mainly on two principles: a set of nucleotides in the tRNA molecule (identity elements) responsible for proper discrimination by aminoacyl-tRNA synthetases1–3 and competition between different synthetases for tRNAs4–6. Elements of glutamine identity are located in the anticodon2, 7–9 and in the acceptor stem region, including the discriminator base5, 10–13. We report here the production of more than 20 tRNAGln2 mutants at positions likely to be involved in tRNA discrimination by the enzyme. Unmodified tRNA, containing the wild-type anticodon and U or G at its 5′-terminus, can be aminocylated by GlnRS with similar kinetic parameters to native tRNAGln2. By in vitro aminoacylation the mutant tRNAs showed decreases of up to 3 x 105-fold in the specificity constant (kcat/KM)14 with the major contribution of kcat. Despite these large changes, some of these mutant tRNAs are efficient amber suppressors in vivo. Our results show that strong elements for glutamine identity reside in the anticodon region and in positions 2 and 3 of the acceptor stem, and that the contribution of different identity elements to the overall discrimination varies significantly. We discuss our data in the light of the crystal structure of the GlnRS :tRNAGln complex15, 16.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Söll, D. Experientia 46, 1089–1096 (1990).

    Article  Google Scholar 

  2. Normanly, J. & Abelson, J. A. Rev. Biochem. 58, 1029–1049 (1989).

    Article  CAS  Google Scholar 

  3. Schimmel, P. Biochemistry 28, 2747–2759 (1989).

    Article  CAS  Google Scholar 

  4. Swanson, R. et al. Science 242, 1548–1551 (1988).

    Article  ADS  CAS  Google Scholar 

  5. Rogers, M. J. & Söll, D. Proc. natn. Acad. Sci. U.S.A. 85, 6627–6631 (1988).

    Article  ADS  CAS  Google Scholar 

  6. Hou, Y. M. & Schimmel, P. Biochemistry 28, 4942–4947 (1989).

    Article  CAS  Google Scholar 

  7. Schulman, L. H. & Pelka, H. Biochemistry 24, 7309–7314 (1985).

    Article  CAS  Google Scholar 

  8. Seno, T., Agris, P. F. & Söll, D. Biochim. biophys. Acta 349, 328–338 (1974).

    Article  CAS  Google Scholar 

  9. Yaniv, M., Folk, W. R., Berg, P. & Söll, L. J. molec. Biol. 86, 245–260 (1974).

    Article  CAS  Google Scholar 

  10. Crothers, D. M., Seno, T. & Söll, D. Proc. natn. Acad. Sci. U.S.A. 69, 3063–3067 (1972).

    Article  ADS  CAS  Google Scholar 

  11. Shimura, Y. et al. FEBS Lett. 22, 144–148 (1972).

    Article  CAS  Google Scholar 

  12. Hooper, J. L., Russell, R. L. & Smith, J. D. FEBS Lett. 22, 149–155 (1972).

    Article  CAS  Google Scholar 

  13. Perona, J. J., Swanson, R. N., Rould, M. A., Steitz, T. A. & Söll, D. Science 246, 1152–1154 (1989).

    Article  ADS  CAS  Google Scholar 

  14. Fersht, A. Enzyme Structure and Mechanism (Freeman, 1985).

    Google Scholar 

  15. Rould, M. A., Perona, J. J., Söll, D. & Steitz, T. A. Science 246, 1135–1142 (1989).

    Article  ADS  CAS  Google Scholar 

  16. Rould, M. A., Perona, J. J. & Steitz, T. A. Nature III, XXX–XXX (this issue).

  17. Inokuchi, H., Hoben, P., Yamao, F., Ozeki, H. & Söll, D. Proc. natn. Acad. Sci. U.S.A. 81, 5076–5080 (1984).

    Article  ADS  CAS  Google Scholar 

  18. Sampson, J. R. & Uhlenbeck, O. C. Proc. natn. Acad. Sci. U.S.A. 85, 1033–1037 (1988).

    Article  ADS  CAS  Google Scholar 

  19. Himeno, H., Hasegawa, T., Ueda, T., Watanabe, K. & Shimizu, M. Nucleic Acids Res. 18, 6815–6819 (1990).

    Article  CAS  Google Scholar 

  20. Hou, Y.-M. & Schimmel, P. Nature 333, 140–145 (1988).

    Article  ADS  CAS  Google Scholar 

  21. Schulman, L. H. & Pelka, H. Science 242, 765–768 (1988).

    Article  ADS  CAS  Google Scholar 

  22. Schulman, L. H. & Pelka, H. Science 246, 1595–1597 (1989).

    Article  ADS  CAS  Google Scholar 

  23. Hasegawa, T., Himeno, H., Ishikura, H. & Shimizu, M. Biochem. biophys. Res. Commun. 163, 1534–1538 (1989).

    Article  CAS  Google Scholar 

  24. Himeno, H. et al. Nucleic Acids Res. 17, 7855–7863 (1989).

    Article  CAS  Google Scholar 

  25. Schulman, L. H. & Pelka, H. Nucleic Acids Res. 18, 285–289 (1990).

    Article  CAS  Google Scholar 

  26. Perret, V. et al. Nature 344, 787–789 (1990).

    Article  ADS  CAS  Google Scholar 

  27. Hall, K. B., Sampson, J. R. & Uhlenbeck, O. C. Biochemistry 28, 5794–5801 (1989).

    Article  CAS  Google Scholar 

  28. Seong, B. L., Lee, C.-P. & RajBhandary, U. L. J. biol. Chem. 246, 6504–6508 (1989).

    Google Scholar 

  29. Redfield, A. G. et al. in Structure and Dynamics of RNA (eds van Knippenberg, P. H. & Hilbers, C. W.) 99–112 (Plenum, New York, 1986).

    Book  Google Scholar 

  30. Yarus, M. Cell 55, 739–741 (1988).

    Article  CAS  Google Scholar 

  31. Hoben, P. J. thesis, Yale Univ. (1984).

  32. Perona, J. J., Swanson, R., Steitz, T. A. & Söll D. J. molec. Biol. 202, 121–126 (1988).

    Article  CAS  Google Scholar 

  33. Milligan, J. F., Groebe, D. R., Witherell, G. W. & Uhlenbeck, O. C. Nucleic Acids Res. 15, 8783–8798 (1987).

    Article  CAS  Google Scholar 

  34. Jahn, M. J. Jahn, D., Kumar, A. W. & Söll, D. Nucleic Acids Res. 19, 2786 (1991).

    Article  CAS  Google Scholar 

  35. Normanly, J., Ogden, R. C., Horvath, S. J. & Abelson, J. Nature 321, 213–219 (1986).

    Article  ADS  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, 06511, USA

    Martina Jahn, M. John Rogers & Dieter Söll

Authors
  1. Martina Jahn
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. M. John Rogers
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Dieter Söll
    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

Jahn, M., Rogers, M. & Söll, D. Anticodon and acceptor stem nucleotides in tRNAGln are major recognition elements for E. coli glutaminyl-tRNA synthetase. Nature 352, 258–260 (1991). https://doi.org/10.1038/352258a0

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/352258a0

  • Springer Nature Limited

This article is cited by

Search

Navigation