Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1998 May 15;17(10):2947–2960. doi: 10.1093/emboj/17.10.2947

The crystal structure of asparaginyl-tRNA synthetase from Thermus thermophilus and its complexes with ATP and asparaginyl-adenylate: the mechanism of discrimination between asparagine and aspartic acid.

C Berthet-Colominas 1, L Seignovert 1, M Härtlein 1, M Grotli 1, S Cusack 1, R Leberman 1
PMCID: PMC1170635  PMID: 9582288

Abstract

The crystal structure of Thermus thermophilus asparaginyl-tRNA synthetase has been solved by multiple isomorphous replacement and refined at 2.6 A resolution. This is the last of the three class IIb aminoacyl-tRNA synthetase structures to be determined. As expected from primary sequence comparisons, there are remarkable similarities between the tertiary structures of asparaginyl-tRNA synthetase and aspartyl-tRNA synthetase, and most of the active site residues are identical except for three key differences. The structure at 2.65 A of asparaginyl-tRNA synthetase complexed with a non-hydrolysable analogue of asparaginyl-adenylate permits a detailed explanation of how these three differences allow each enzyme to discriminate between their respective and very similar amino acid substrates, asparagine and aspartic acid. In addition, a structure of the complex of asparaginyl-tRNA synthetase with ATP shows exactly the same configuration of three divalent cations as previously observed in the seryl-tRNA synthetase-ATP complex, showing that this a general feature of class II synthetases. The structural similarity of asparaginyl- and aspartyl-tRNA synthetases as well as that of both enzymes to the ammonia-dependent asparagine synthetase suggests that these three enzymes have evolved relatively recently from a common ancestor.

Full Text

The Full Text of this article is available as a PDF (1.2 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aberg A., Yaremchuk A., Tukalo M., Rasmussen B., Cusack S. Crystal structure analysis of the activation of histidine by Thermus thermophilus histidyl-tRNA synthetase. Biochemistry. 1997 Mar 18;36(11):3084–3094. doi: 10.1021/bi9618373. [DOI] [PubMed] [Google Scholar]
  2. Airas R. K. Differences in the magnesium dependences of the class I and class II aminoacyl-tRNA synthetases from Escherichia coli. Eur J Biochem. 1996 Aug 15;240(1):223–231. doi: 10.1111/j.1432-1033.1996.0223h.x. [DOI] [PubMed] [Google Scholar]
  3. Arnez J. G., Augustine J. G., Moras D., Francklyn C. S. The first step of aminoacylation at the atomic level in histidyl-tRNA synthetase. Proc Natl Acad Sci U S A. 1997 Jul 8;94(14):7144–7149. doi: 10.1073/pnas.94.14.7144. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beaulande M., Tarbouriech N., Härtlein M. Human cytosolic asparaginyl-tRNA synthetase: cDNA sequence, functional expression in Escherichia coli and characterization as human autoantigen. Nucleic Acids Res. 1998 Jan 15;26(2):521–524. doi: 10.1093/nar/26.2.521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Belrhali H., Yaremchuk A., Tukalo M., Berthet-Colominas C., Rasmussen B., Bösecke P., Diat O., Cusack S. The structural basis for seryl-adenylate and Ap4A synthesis by seryl-tRNA synthetase. Structure. 1995 Apr 15;3(4):341–352. doi: 10.1016/s0969-2126(01)00166-6. [DOI] [PubMed] [Google Scholar]
  6. Belrhali H., Yaremchuk A., Tukalo M., Larsen K., Berthet-Colominas C., Leberman R., Beijer B., Sproat B., Als-Nielsen J., Grübel G. Crystal structures at 2.5 angstrom resolution of seryl-tRNA synthetase complexed with two analogs of seryl adenylate. Science. 1994 Mar 11;263(5152):1432–1436. doi: 10.1126/science.8128224. [DOI] [PubMed] [Google Scholar]
  7. Berthet-Colominas C., Seignovert L., Cusack S., Leberman R. Preliminary X-ray diffraction studies on asparaginyl-tRNA synthetase from Thermus thermophilus. Acta Crystallogr D Biol Crystallogr. 1997 Mar 1;53(Pt 2):195–196. doi: 10.1107/S0907444996014096. [DOI] [PubMed] [Google Scholar]
  8. Bult C. J., White O., Olsen G. J., Zhou L., Fleischmann R. D., Sutton G. G., Blake J. A., FitzGerald L. M., Clayton R. A., Gocayne J. D. Complete genome sequence of the methanogenic archaeon, Methanococcus jannaschii. Science. 1996 Aug 23;273(5278):1058–1073. doi: 10.1126/science.273.5278.1058. [DOI] [PubMed] [Google Scholar]
  9. Cavarelli J., Eriani G., Rees B., Ruff M., Boeglin M., Mitschler A., Martin F., Gangloff J., Thierry J. C., Moras D. The active site of yeast aspartyl-tRNA synthetase: structural and functional aspects of the aminoacylation reaction. EMBO J. 1994 Jan 15;13(2):327–337. doi: 10.1002/j.1460-2075.1994.tb06265.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cavarelli J., Rees B., Ruff M., Thierry J. C., Moras D. Yeast tRNA(Asp) recognition by its cognate class II aminoacyl-tRNA synthetase. Nature. 1993 Mar 11;362(6416):181–184. doi: 10.1038/362181a0. [DOI] [PubMed] [Google Scholar]
  11. Curnow A. W., Ibba M., Söll D. tRNA-dependent asparagine formation. Nature. 1996 Aug 15;382(6592):589–590. doi: 10.1038/382589b0. [DOI] [PubMed] [Google Scholar]
  12. Cusack S. Aminoacyl-tRNA synthetases. Curr Opin Struct Biol. 1997 Dec;7(6):881–889. doi: 10.1016/s0959-440x(97)80161-3. [DOI] [PubMed] [Google Scholar]
  13. Cusack S., Berthet-Colominas C., Härtlein M., Nassar N., Leberman R. A second class of synthetase structure revealed by X-ray analysis of Escherichia coli seryl-tRNA synthetase at 2.5 A. Nature. 1990 Sep 20;347(6290):249–255. doi: 10.1038/347249a0. [DOI] [PubMed] [Google Scholar]
  14. Cusack S. Eleven down and nine to go. Nat Struct Biol. 1995 Oct;2(10):824–831. doi: 10.1038/nsb1095-824. [DOI] [PubMed] [Google Scholar]
  15. Cusack S., Yaremchuk A., Tukalo M. The crystal structures of T. thermophilus lysyl-tRNA synthetase complexed with E. coli tRNA(Lys) and a T. thermophilus tRNA(Lys) transcript: anticodon recognition and conformational changes upon binding of a lysyl-adenylate analogue. EMBO J. 1996 Nov 15;15(22):6321–6334. [PMC free article] [PubMed] [Google Scholar]
  16. Delarue M., Poterszman A., Nikonov S., Garber M., Moras D., Thierry J. C. Crystal structure of a prokaryotic aspartyl tRNA-synthetase. EMBO J. 1994 Jul 15;13(14):3219–3229. doi: 10.1002/j.1460-2075.1994.tb06623.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Eriani G., Cavarelli J., Martin F., Dirheimer G., Moras D., Gangloff J. Role of dimerization in yeast aspartyl-tRNA synthetase and importance of the class II invariant proline. Proc Natl Acad Sci U S A. 1993 Nov 15;90(22):10816–10820. doi: 10.1073/pnas.90.22.10816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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. 1990 Sep 13;347(6289):203–206. doi: 10.1038/347203a0. [DOI] [PubMed] [Google Scholar]
  19. Gatti D. L., Tzagoloff A. Structure and evolution of a group of related aminoacyl-tRNA synthetases. J Mol Biol. 1991 Apr 5;218(3):557–568. doi: 10.1016/0022-2836(91)90701-7. [DOI] [PubMed] [Google Scholar]
  20. Ibba M., Curnow A. W., Söll D. Aminoacyl-tRNA synthesis: divergent routes to a common goal. Trends Biochem Sci. 1997 Feb;22(2):39–42. doi: 10.1016/s0968-0004(96)20033-7. [DOI] [PubMed] [Google Scholar]
  21. Kron M., Marquard K., Härtlein M., Price S., Leberman R. An immunodominant antigen of Brugia malayi is an asparaginyl-tRNA synthetase. FEBS Lett. 1995 Oct 23;374(1):122–124. doi: 10.1016/0014-5793(95)01092-s. [DOI] [PubMed] [Google Scholar]
  22. Nakatsu T., Kato H., Oda J. Crystal structure of asparagine synthetase reveals a close evolutionary relationship to class II aminoacyl-tRNA synthetase. Nat Struct Biol. 1998 Jan;5(1):15–19. doi: 10.1038/nsb0198-15. [DOI] [PubMed] [Google Scholar]
  23. Onesti S., Miller A. D., Brick P. The crystal structure of the lysyl-tRNA synthetase (LysU) from Escherichia coli. Structure. 1995 Feb 15;3(2):163–176. doi: 10.1016/s0969-2126(01)00147-2. [DOI] [PubMed] [Google Scholar]
  24. Poterszman A., Delarue M., Thierry J. C., Moras D. Synthesis and recognition of aspartyl-adenylate by Thermus thermophilus aspartyl-tRNA synthetase. J Mol Biol. 1994 Nov 25;244(2):158–167. doi: 10.1006/jmbi.1994.1716. [DOI] [PubMed] [Google Scholar]
  25. Ruff M., Krishnaswamy S., Boeglin M., Poterszman A., Mitschler A., Podjarny A., Rees B., Thierry J. C., Moras D. Class II aminoacyl transfer RNA synthetases: crystal structure of yeast aspartyl-tRNA synthetase complexed with tRNA(Asp). Science. 1991 Jun 21;252(5013):1682–1689. doi: 10.1126/science.2047877. [DOI] [PubMed] [Google Scholar]
  26. Seignovert L., Härtlein M., Leberman R. Asparaginyl-tRNA synthetase from Thermus thermophilus HB8. Sequence of the gene and crystallization of the enzyme expressed in Escherichia coli. Eur J Biochem. 1996 Jul 15;239(2):501–508. doi: 10.1111/j.1432-1033.1996.0501u.x. [DOI] [PubMed] [Google Scholar]
  27. Tomb J. F., White O., Kerlavage A. R., Clayton R. A., Sutton G. G., Fleischmann R. D., Ketchum K. A., Klenk H. P., Gill S., Dougherty B. A. The complete genome sequence of the gastric pathogen Helicobacter pylori. Nature. 1997 Aug 7;388(6642):539–547. doi: 10.1038/41483. [DOI] [PubMed] [Google Scholar]
  28. Ueda H., Shoku Y., Hayashi N., Mitsunaga J., In Y., Doi M., Inoue M., Ishida T. X-ray crystallographic conformational study of 5'-O-[N-(L-alanyl)-sulfamoyl]adenosine, a substrate analogue for alanyl-tRNA synthetase. Biochim Biophys Acta. 1991 Oct 25;1080(2):126–134. doi: 10.1016/0167-4838(91)90138-p. [DOI] [PubMed] [Google Scholar]
  29. Yip K. S., Stillman T. J., Britton K. L., Artymiuk P. J., Baker P. J., Sedelnikova S. E., Engel P. C., Pasquo A., Chiaraluce R., Consalvi V. The structure of Pyrococcus furiosus glutamate dehydrogenase reveals a key role for ion-pair networks in maintaining enzyme stability at extreme temperatures. Structure. 1995 Nov 15;3(11):1147–1158. doi: 10.1016/s0969-2126(01)00251-9. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES