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Structural basis for substrate binding, cleavage and allostery in the tRNA maturase RNase Z

Nature volume 433, pages 657–661 (2005)Cite this article

Abstract

Transfer RNAs (tRNAs) are synthesized as part of longer primary transcripts that require processing of both their 3′ and 5′ extremities in every living organism known. The 5′ side is processed (matured) by the ubiquitously conserved endonucleolytic ribozyme, RNase P1, whereas removal of the 3′ tails can be either exonucleolytic2,3 or endonucleolytic4. The endonucleolytic pathway is catalysed by an enzyme known as RNase Z, or 3′ tRNase5,6. RNase Z cleaves precursor tRNAs immediately after the discriminator base (the unpaired nucleotide 3′ to the last base pair of the acceptor stem, used as an identity determinant by many aminoacyl-tRNA synthetases) in most cases6,7,8, yielding a tRNA primed for addition of the CCA motif by nucleotidyl transferase. Here we report the crystal structure of Bacillus subtilis RNase Z at 2.1 Å resolution, and propose a mechanism for tRNA recognition and cleavage. The structure explains the allosteric properties of the enzyme, and also sheds light on the mechanisms of inhibition by the CCA motif and long 5′ extensions. Finally, it highlights the extraordinary adaptability of the metallo-hydrolase domain of the β-lactamase family for the hydrolysis of covalent bonds.

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Figure 1: Ribbon representation of RNase Z at 2.1 Å resolution.
Figure 2: Structural comparison of members of the metallo-β-lactamase family.
Figure 3: Model of RNase Z–tRNA complex and amino acids important for interaction.
Figure 4: Active sites of RNase Z subunits and the proposed cleavage mechanism.

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References

  1. Frank, D. N. & Pace, N. R. Ribonuclease P: unity and diversity in a tRNA processing ribozyme. Annu. Rev. Biochem. 67, 153–180 (1998)

    Article  CAS  Google Scholar 

  2. Garber, R. L. & Altman, S. In vitro processing of B. mori transfer RNA precursor molecules. Cell 17, 389–397 (1979)

    Article  CAS  Google Scholar 

  3. Li, Z. & Deutscher, M. P. Maturation pathways for E. coli tRNA precursors: a random multienzyme process in vivo. Cell 86, 503–512 (1996)

    Article  CAS  Google Scholar 

  4. Garber, R. L. & Gage, L. P. Transcription of a cloned Bombyx mori tRNA2Ala gene: nucleotide sequence of the tRNA precursor and its processing in vitro. Cell 18, 817–828 (1979)

    Article  CAS  Google Scholar 

  5. Schiffer, S., Rosch, S. & Marchfelder, A. Assigning a function to a conserved group of proteins: the tRNA 3′-processing enzymes. EMBO J. 21, 2769–2777 (2002)

    Article  CAS  Google Scholar 

  6. Nashimoto, M. Distribution of both lengths and 5′ terminal nucleotides of mammalian pre-tRNA 3′ trailers reflects properties of 3′ processing endoribonuclease. Nucleic Acids Res. 25, 1148–1154 (1997)

    Article  CAS  Google Scholar 

  7. Kunzmann, A., Brennicke, A. & Marchfelder, A. 5′ end maturation and RNA editing have to precede tRNA 3′ processing in plant mitochondria. Proc. Natl Acad. Sci. USA 95, 108–113 (1998)

    Article  ADS  CAS  Google Scholar 

  8. Schierling, K., Rosch, S., Rupprecht, R., Schiffer, S. & Marchfelder, A. tRNA 3′ end maturation in archaea has eukaryotic features: the RNase Z from Haloferax volcanii. J. Mol. Biol. 316, 895–902 (2002)

    Article  CAS  Google Scholar 

  9. Pellegrini, O., Nezzar, J., Marchfelder, A., Putzer, H. & Condon, C. Endonucleolytic processing of CCA-less tRNA precursors by RNase Z in Bacillus subtilis. EMBO J. 22, 4534–4543 (2003)

    Article  CAS  Google Scholar 

  10. Tavtigian, S. V. et al. A candidate prostate cancer susceptibility gene at chromosome 17p. Nature Genet. 27, 172–180 (2001)

    Article  CAS  Google Scholar 

  11. Minagawa, A., Takaku, H., Takagi, M. & Nashimoto, M. A novel endonucleolytic mechanism to generate the CCA 3′ termini of tRNA molecules in Thermotoga maritima. J. Biol. Chem. 279, 15688–15697 (2004)

    Article  CAS  Google Scholar 

  12. Mohan, A., Whyte, S., Wang, X., Nashimoto, M. & Levinger, L. The 3′ end CCA of mature tRNA is an antideterminant for eukaryotic 3′-tRNase. RNA 5, 245–256 (1999)

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  14. Schiffer, S., Helm, M., Theobald-Dietrich, A., Giege, R. & Marchfelder, A. The plant tRNA 3′ processing enzyme has a broad substrate spectrum. Biochemistry 40, 8264–8272 (2001)

    Article  CAS  Google Scholar 

  15. Cameron, A. D., Ridderstrom, M., Olin, B. & Mannervik, B. Crystal structure of human glyoxalase II and its complex with a glutathione thiolester substrate analogue. Struct. Fold. Des. 7, 1067–1078 (1999)

    Article  CAS  Google Scholar 

  16. Nashimoto, M., Wesemann, D. R., Geary, S., Tamura, M. & Kaspar, R. L. Long 5′ leaders inhibit removal of a 3′ trailer from a precursor tRNA by mammalian tRNA 3′ processing endoribonuclease. Nucleic Acids Res. 27, 2770–2776 (1999)

    Article  CAS  Google Scholar 

  17. Vogel, A., Schilling, O., Niecke, M., Bettmer, J. & Meyer-Klaucke, W. ElaC encodes a novel binuclear zinc phosphodiesterase. J. Biol. Chem. 277, 29078–29085 (2002)

    Article  CAS  Google Scholar 

  18. Daiyasu, H., Osaka, K., Ishino, Y. & Toh, H. Expansion of the zinc metallo-hydrolase family of the β-lactamase fold. FEBS Lett. 503, 1–6 (2001)

    Article  ADS  CAS  Google Scholar 

  19. Carfi, A. et al. The 3-D structure of a zinc metallo-beta-lactamase from Bacillus cereus reveals a new type of protein fold. EMBO J. 14, 4914–4921 (1995)

    Article  CAS  Google Scholar 

  20. Frazao, C. et al. Structure of a dioxygen reduction enzyme from Desulfovibrio gigas. Nature Struct. Biol. 7, 1041–1045 (2000)

    Article  CAS  Google Scholar 

  21. Matthews, B. W. Solvent content of protein crystals. J. Mol. Biol. 33, 491–497 (1968)

    Article  CAS  Google Scholar 

  22. CCP4. The CCP4 Suite: Programs for protein crystallography. Acta Crystallogr. D 50, 760–763 (1994)

    Article  Google Scholar 

  23. Otwinowski, Z. & Minor, W. in Methods in Enzymology Vol. 276, Macromolecular Crystallography Part A (eds Carter, C. W. Jr & Sweet, R. M.) 307–326 (Academic, New York, 1997)

    Book  Google Scholar 

  24. Uson, I. & Sheldrick, G. M. Advances in direct methods for protein crystallography. Curr. Opin. Struct. Biol. 9, 643–648 (1999)

    Article  CAS  Google Scholar 

  25. Schneider, T. R. & Sheldrick, G. M. Substructure solution with SHELXD. Acta Crystallogr. D 58, 1772–1779 (2002)

    Article  Google Scholar 

  26. Lu, G. FINDNCS: A program to detect non-crystallographic symmetries in protein crystals from heavy atom sites. J. Appl. Crystallogr. 32, 365–368 (1999)

    Article  CAS  Google Scholar 

  27. Perrakis, A., Morris, R. & Lamzin, V. S. Automated protein model building combined with iterative structure refinement. Nature Struct. Biol. 6, 458–463 (1999)

    Article  CAS  Google Scholar 

  28. Kleywegt, G. J. & Jones, T. A. in From First Map to Final Model (eds Bailey, S. R. H. & Waller, D.) 59–66 (SERC Daresbury Laboratory, Daresbury, UK, 1994)

    Google Scholar 

  29. Berman, H. M. et al. The Protein Data Bank. Nucleic Acids Res. 28, 235–242 (2000)

    Article  ADS  CAS  Google Scholar 

Download references

Acknowledgements

We thank M. Springer, D. Picot, R. Giégé, F. Allemand, V. Arluison and F. Dardel for discussions, S. Fieulaine and M. Pirocchi for help with the beam-line BM30A at the European Synchrotron Radiation Facility, and J.L. Popot for use of crystallography facilities and X-ray generator at the IBPC. This work was supported by the CNRS (UPR 9073), Université Paris VII-Denis Diderot, PRFMMIP 2001/2003, and ACI Jeunes Chercheurs from the Ministère de l'Education Nationale.

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Authors and Affiliations

  1. CNRS UPR 9073, affiliated with Université de Paris 7—Denis Diderot,

    Olivier Pellegrini & Ciarán Condon

  2. FRC550, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005, Paris, France

    Inés Li de la Sierra-Gallay

Authors
  1. Inés Li de la Sierra-Gallay
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  3. Ciarán Condon
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Corresponding author

Correspondence to Ciarán Condon.

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Supplementary information

Supplementary Figure S1

Shows an alignment COGs representing different members of the metallo -lactamase family. (PPT 1058 kb)

Supplementary Figure S2

Shows both the secondary structure features of RNase Z superimposed on its amino acid sequence and the secondary structure topology of the protein. (PPT 52 kb)

Supplementary Figure S3

Shows the crystal packing of RNase Z. (PPT 122 kb)

Supplementary Methods

Describes the seleno-methionine labelling of RNase Z. (RTF 3 kb)

Supplementary Table

Shows data-collection, phasing and refinement statistics for resolution of RNase Z structure. (RTF 304 kb)

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de la Sierra-Gallay, I., Pellegrini, O. & Condon, C. Structural basis for substrate binding, cleavage and allostery in the tRNA maturase RNase Z. Nature 433, 657–661 (2005). https://doi.org/10.1038/nature03284

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