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Aminoacylation complex structures of leucyl-tRNA synthetase and tRNALeu reveal two modes of discriminator-base recognition

Nature Structural & Molecular Biology volume 12, pages 915–922 (2005)Cite this article

Abstract

Leucyl-tRNA synthetase (LeuRS) specifically recognizes the characteristic long variable arm and the discriminator base, A73, of tRNALeu in archaea and eukarya. The LeuRS 'editing domain' hydrolyzes misformed noncognate aminoacyl-tRNA. Here we report the crystal structure of the archaeal Pyrococcus horikoshii LeuRS–tRNALeu complex. The protruding C-terminal domain of LeuRS specifically recognizes the bases at the tip of the long variable arm. The editing domain swings from its tRNA-free position to avoid clashing with the tRNA. Consequently the tRNA CCA end can bend and reach the aminoacylation active site. The tRNA 3′ region assumes two distinct conformations that allow A73 to be specifically recognized in different ways. One conformation is the canonical 'aminoacylation state.' The other conformation seems to be the 'intermediate state,' where the misaminoacylated 3′ end has partially relocated to the editing domain.

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Figure 1: Overall structure of the P. horikoshii LeuRS–tRNALeu complex.
Figure 2: Recognition of the long variable arm of tRNA by the C-terminal domain.
Figure 3: C-terminal domain architectures of P. horikoshii (P.h.) LeuRS, S. aureus (S.a.) IleRS and T. thermophilus (T.t.) ValRS.
Figure 4: The two modes of recognition of the discriminator A73 and the two acceptor stem conformations.
Figure 5: Relocation of CCA end toward the editing domain.

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References

  1. Larkin, D.C., Williams, A.M., Martinis, S.A. & Fox, G.E. Identification of essential domains for Escherichia coli tRNA(leu) aminoacylation and amino acid editing using minimalist RNA molecules. Nucleic Acids Res. 30, 2103–2113 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Asahara, H. et al. Recognition nucleotides of Escherichia coli tRNA(Leu) and its elements facilitating discrimination from tRNASer and tRNA(Tyr). J. Mol. Biol. 231, 219–229 (1993).

    Article  CAS  PubMed  Google Scholar 

  3. Small, I. et al. In vivo import of a normal or mutagenized heterologous transfer RNA into the mitochondria of transgenic plants: towards novel ways of influencing mitochondrial gene expression? EMBO J. 11, 1291–1296 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Breitschopf, K., Achsel, T., Busch, K. & Gross, H.J. Identity elements of human tRNA(Leu): structural requirements for converting human tRNA(Ser) into a leucine acceptor in vitro. Nucleic Acids Res. 23, 3633–3637 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Soma, A., Uchiyama, K., Sakamoto, T., Maeda, M. & Himeno, H. Unique recognition style of tRNA(Leu) by Haloferax volcanii leucyl-tRNA synthetase. J. Mol. Biol. 293, 1029–1038 (1999).

    Article  CAS  PubMed  Google Scholar 

  6. Yaremchuk, A., Kriklivyi, I., Tukalo, M. & Cusack, S. Class I tyrosyl-tRNA synthetase has a class II mode of cognate tRNA recognition. EMBO J. 21, 3829–3840 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Biou, V., Yaremchuk, A., Tukalo, M. & Cusack, S. The 2.9 A crystal structure of T. thermophilus seryl-tRNA synthetase complexed with tRNA(Ser). Science 263, 1404–1410 (1994).

    Article  CAS  PubMed  Google Scholar 

  8. Eriani, G., Delarue, M., Poch, O., Gangloff, J. & Moras, D. Partition of tRNA synthetases into two classes based on mutually exclusive sets of sequence motifs. Nature 347, 203–206 (1990).

    Article  CAS  PubMed  Google Scholar 

  9. Rould, M.A., Perona, J.J., Soll, D. & Steitz, T.A. Structure of E. coli glutaminyl-tRNA synthetase complexed with tRNA(Gln) and ATP at 2.8 A resolution. Science 246, 1135–1142 (1989).

    Article  CAS  PubMed  Google Scholar 

  10. Ruff, M. et al. Class II aminoacyl transfer RNA synthetases: crystal structure of yeast aspartyl-tRNA synthetase complexed with tRNA(Asp). Science 252, 1682–1689 (1991).

    Article  CAS  PubMed  Google Scholar 

  11. Jakubowski, H. & Goldman, E. Editing of errors in selection of amino acids for protein synthesis. Microbiol. Rev. 56, 412–429 (1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Schmidt, E. & Schimmel, P. Residues in a class I tRNA synthetase which determine selectivity of amino acid recognition in the context of tRNA. Biochemistry 34, 11204–11210 (1995).

    Article  CAS  PubMed  Google Scholar 

  13. Lin, L., Hale, S.P. & Schimmel, P. Aminoacylation error correction. Nature 384, 33–34 (1996).

    Article  CAS  PubMed  Google Scholar 

  14. Nureki, O. et al. Enzyme structure with two catalytic sites for double-sieve selection of substrate. Science 280, 578–582 (1998).

    Article  CAS  PubMed  Google Scholar 

  15. Lincecum, T.L. et al. Structural and mechanistic basis of pre- and posttransfer editing by leucyl-tRNA synthetase. Mol. Cell 11, 951–963 (2003).

    Article  CAS  PubMed  Google Scholar 

  16. Fukunaga, R., Fukai, S., Ishitani, R., Nureki, O. & Yokoyama, S. Crystal structures of the CP1 domain from Thermus thermophilus isoleucyl-tRNA synthetase and its complex with L–valine. J. Biol. Chem. 279, 8396–8402 (2004).

    Article  CAS  PubMed  Google Scholar 

  17. Fukai, S. et al. Structural basis for double-sieve discrimination of L-valine from L-isoleucine and L-threonine by the complex of tRNA(Val) and valyl-tRNA synthetase. Cell 103, 793–803 (2000).

    Article  CAS  PubMed  Google Scholar 

  18. Silvian, L.F., Wang, J. & Steitz, T.A. Insights into editing from an Ile-tRNA synthetase structure with tRNAIle and mupirocin. Science 285, 1074–1077 (1999).

    Article  CAS  PubMed  Google Scholar 

  19. Chen, J.F., Guo, N.N., Li, T., Wang, E.D. & Wang, Y.L. CP1 domain in Escherichia coli leucyl-tRNA synthetase is crucial for its editing function. Biochemistry 39, 6726–6731 (2000).

    Article  CAS  PubMed  Google Scholar 

  20. Baldwin, A.N. & Berg, P. Transfer ribonucleic acid-induced hydrolysis of valyladenylate bound to isoleucyl ribonucleic acid synthetase. J. Biol. Chem. 241, 839–845 (1966).

    CAS  PubMed  Google Scholar 

  21. Hale, S.P., Auld, D.S., Schmidt, E. & Schimmel, P. Discrete determinants in transfer RNA for editing and aminoacylation. Science 276, 1250–1252 (1997).

    Article  CAS  PubMed  Google Scholar 

  22. Fukunaga, R. & Yokoyama, S. Crystal structure of leucyl-tRNA synthetase from the archaeon Pyrococcus horikoshii reveals a novel editing domain orientation. J. Mol. Biol. 346, 57–71 (2005).

    Article  CAS  PubMed  Google Scholar 

  23. Fukunaga, R. & Yokoyama, S. Structural basis for non-cognate amino acid discrimination by the valyl-tRNA synthetase editing domain. J. Biol. Chem. 280, 29937–29945 (2005).

    Article  CAS  PubMed  Google Scholar 

  24. Sekine, S., Nureki, O., Shimada, A., Vassylyev, D.G. & Yokoyama, S. Structural basis for anticodon recognition by discriminating glutamyl-tRNA synthetase. Nat. Struct. Biol. 8, 203–206 (2001).

    Article  CAS  PubMed  Google Scholar 

  25. Delagoutte, B., Moras, D. & Cavarelli, J. tRNA aminoacylation by arginyl-tRNA synthetase: induced conformations during substrates binding. EMBO J. 19, 5599–5610 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Hauenstein, S., Zhang, C.M., Hou, Y.M. & Perona, J.J. Shape-selective RNA recognition by cysteinyl-tRNA synthetase. Nat. Struct. Mol. Biol. 11, 1134–1141 (2004).

    Article  CAS  PubMed  Google Scholar 

  27. Fukai, S. et al. Mechanism of molecular interactions for tRNA(Val) recognition by valyl-tRNA synthetase. RNA 9, 100–111 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Tukalo, M., Yaremchuk, A., Fukunaga, R., Yokoyama, S. & Cusack, S. The crystal structure of leucyl-tRNA synthetase complexed with tRNALeu in the editing conformation. Nat. Struct. Mol. Biol., advance online publication 11 September 2005 (10.1038/nsmb986)

  29. Cusack, S., Yaremchuk, A. & Tukalo, M. The 2 Å crystal structure of leucyl-tRNA synthetase and its complex with a leucyl-adenylate analogue. EMBO J. 19, 2351–2361 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Fukunaga, R., Ishitani, R., Nureki, O. & Yokoyama, S. Crystallization of leucyl-tRNA synthetase complexed with tRNALeu from the archaeon Pyrococcus horikoshii. Acta Crystallogr. F 61, 30–32 (2005).

    Article  CAS  Google Scholar 

  31. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. in Methods Enzymol. Vol. 276 (eds. Carter, C.W. Jr. & Sweet, R.M.) 307–326 (Academic Press, San Diego, USA, 1997).

    Article  CAS  PubMed  Google Scholar 

  32. Weeks, C.M. & Miller, R. Optimizing Shake-and-Bake for proteins. Acta Crystallogr. D Biol. Crystallogr. 55, 492–500 (1999).

    Article  CAS  PubMed  Google Scholar 

  33. CCP4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  34. Fukunaga, R. & Yokoyama, S. Crystallization and preliminary X-ray crystallographic study of leucyl-tRNA synthetase from the archaeon Pyrococcus horikoshii. Acta Crystallogr. D Biol. Crystallogr. 60, 1916–1918 (2004).

    Article  PubMed  Google Scholar 

  35. Brunger, A.T. et al. Crystallography & NMR system: a new software suite for macromolecular structure determination. Acta Crystallogr. D Biol. Crystallogr. 54, 905–921 (1998).

    Article  CAS  PubMed  Google Scholar 

  36. Jones, T.A., Zou, J.Y., Cowan, S.W. & Kjeldgaard, Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  PubMed  Google Scholar 

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Acknowledgements

We thank M. Tukalo, A. Yaremchuk and S. Cusack for providing the coordinates of the T. thermophilus LeuRS–tRNALeu complex before publication and for helpful discussions. We thank R. Ishitani and O. Nureki for their help with crystal preparation and X-ray data collection. We also thank T. Sengoku, T. Yanagisawa, S. Sekine, M. Kawamoto, H. Sakai and M. Yamamoto for their help with data collection at BL41XU and BL26B1 in SPring-8. This work was supported by Grants-in-Aid for Scientific Research in Priority Areas from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan, the RIKEN Structural Genomics/Proteomics Initiative (RSGI) and the National Project on Protein Structural and Functional Analyses, MEXT. R.F. was supported by Research Fellowships of the Japan Society for the Promotion of Science for Young Scientists and by SPring-8 Budding Researchers Support.

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Authors and Affiliations

  1. Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, Tokyo, Japan

    Ryuya Fukunaga & Shigeyuki Yokoyama

  2. RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, 230-0045, Yokohama, Japan

    Ryuya Fukunaga & Shigeyuki Yokoyama

  3. RIKEN Harima Institute at SPring-8, 1-1-1 Kouto, Mikazuki-cho, Sayo-gun, 679-5148, Hyogo, Japan

    Shigeyuki Yokoyama

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Correspondence to Shigeyuki Yokoyama.

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Supplementary information

Supplementary Fig. 1

Omit electron density maps of tRNA. (PDF 506 kb)

Supplementary Fig. 2

Two modes of discriminator recognition. (PDF 448 kb)

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Fukunaga, R., Yokoyama, S. Aminoacylation complex structures of leucyl-tRNA synthetase and tRNALeu reveal two modes of discriminator-base recognition. Nat Struct Mol Biol 12, 915–922 (2005). https://doi.org/10.1038/nsmb985

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