Skip to main content

Thank you for visiting 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.

Structure of the 30S translation initiation complex


Translation initiation, the rate-limiting step of the universal process of protein synthesis, proceeds through sequential, tightly regulated steps. In bacteria, the correct messenger RNA start site and the reading frame are selected when, with the help of initiation factors IF1, IF2 and IF3, the initiation codon is decoded in the peptidyl site of the 30S ribosomal subunit by the fMet-tRNAfMet anticodon. This yields a 30S initiation complex (30SIC) that is an intermediate in the formation of the 70S initiation complex (70SIC) that occurs on joining of the 50S ribosomal subunit to the 30SIC and release of the initiation factors1,2,3. The localization of IF2 in the 30SIC has proved to be difficult so far using biochemical approaches, but could now be addressed using cryo-electron microscopy and advanced particle separation techniques on the basis of three-dimensional statistical analysis. Here we report the direct visualization of a 30SIC containing mRNA, fMet-tRNAfMet and initiation factors IF1 and GTP-bound IF2. We demonstrate that the fMet-tRNAfMet is held in a characteristic and precise position and conformation by two interactions that contribute to the formation of a stable complex: one involves the transfer RNA decoding stem which is buried in the 30S peptidyl site, and the other occurs between the carboxy-terminal domain of IF2 and the tRNA acceptor end. The structure provides insights into the mechanism of 70SIC assembly and rationalizes the rapid activation of GTP hydrolysis triggered on 30SIC–50S joining2,3 by showing that the GTP-binding domain of IF2 would directly face the GTPase-activated centre of the 50S subunit.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Get just this article for as long as you need it


Prices may be subject to local taxes which are calculated during checkout

Figure 1: One sample—several structures.
Figure 2: Fitting and map interpretation of the 30SIC.
Figure 3: Comparisons of the tRNA and IF2 positions in different translation initiation complexes, and model of ribosomal subunit joining.

Accession codes

Primary accessions


Data deposits

The electron density map of the 30SIC complex has been deposited to the EM Data Bank under the accession number EMD-1523.


  1. Gualerzi, C. O. et al. Initiation factors in the early events of mRNA translation in bacteria. Cold Spring Harb. Symp. Quant. Biol. 66, 363–376 (2001)

    Article  CAS  Google Scholar 

  2. Tomšic, J. et al. Late events of translation initiation in bacteria: a kinetic analysis. EMBO J. 19, 2127–2136 (2000)

    Article  Google Scholar 

  3. Grigoriadou, C., Marzi, S., Kirillov, S., Gualerzi, C. O. & Cooperman, B. S. A quantitative kinetic scheme for 70 S translation initiation complex formation. J. Mol. Biol. 373, 562–572 (2007)

    Article  CAS  Google Scholar 

  4. Caserta, E. et al. Translation initiation factor IF2 interacts with the 30 S ribosomal subunits via two separate binding sites. J. Mol. Biol. 362, 787–799 (2006)

    Article  CAS  Google Scholar 

  5. Simonetti, A. et al. Nature Protocols 10.1038/nprot.2008.130 (2008)

  6. Klaholz, B. P., Myasnikov, A. G. & van Heel, M. Visualization of release factor 3 on the ribosome during termination of protein synthesis. Nature 427, 862–865 (2004)

    Article  ADS  CAS  Google Scholar 

  7. Wimberly, B. T. et al. Structure of the 30S ribosomal subunit. Nature 407, 327–339 (2000)

    Article  ADS  CAS  Google Scholar 

  8. Schluenzen, F. et al. Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution. Cell 102, 615–623 (2000)

    Article  CAS  Google Scholar 

  9. Korostelev, A., Trakhanov, S., Laurberg, M. & Noller, H. F. Crystal structure of a 70S ribosome-tRNA complex reveals functional interactions and rearrangements. Cell 126, 1065–1077 (2006)

    Article  CAS  Google Scholar 

  10. Yusupova, G., Jenner, L., Rees, B., Moras, D. & Yusupov, M. Structural basis for messenger RNA movement on the ribosome. Nature 444, 391–394 (2006)

    Article  ADS  CAS  Google Scholar 

  11. Carter, A. P. et al. Crystal structure of an initiation factor bound to the 30S ribosomal subunit. Science 291, 498–501 (2001)

    Article  ADS  CAS  Google Scholar 

  12. Roll-Mecak, A., Cao, C., Dever, T. E. & Burley, S. K. X-ray structures of the universal translation initiation factor IF2/eIF5B: conformational changes on GDP and GTP binding complex. Cell 103, 781–792 (2000)

    Article  CAS  Google Scholar 

  13. Myasnikov, A. G. et al. Conformational transition of initiation factor 2 from the GTP- to GDP-bound state visualized on the ribosome. Nature Struct. Mol. Biol. 12, 1145–1149 (2005)

    Article  CAS  Google Scholar 

  14. Selmer, M. et al. Structure of the 70S ribosome complexed with mRNA and tRNA. Science 313, 1935–1942 (2006)

    Article  ADS  CAS  Google Scholar 

  15. Szkaradkiewicz, K., Zuleeg, S., Limmer, S. & Sprinzl, M. Interaction of fMet-tRNAfMet and fMet-AMP with the C-terminal domain of Thermus thermophilus translation initiation factor 2. Eur. J. Biochem. 267, 4290–4299 (2000)

    Article  CAS  Google Scholar 

  16. Guenneugues, M. et al. Mapping the fMet-tRNA binding site of initiation factor IF2. EMBO J. 19, 5233–5249 (2000)

    Article  CAS  Google Scholar 

  17. Valle, M. et al. Cryo-EM reveals an active role for aminoacyl-tRNA in the accommodation process. EMBO J. 21, 3557–3567 (2002)

    Article  CAS  Google Scholar 

  18. Moazed, D. & Noller, H. F. Intermediate states in the movement of transfer RNA in the ribosome. Nature 342, 142–148 (1989)

    Article  ADS  CAS  Google Scholar 

  19. Allen, G. S., Zavialov, A., Gursky, R., Ehrenberg, M. & Frank, J. The cryo-EM structure of a translation initiation complex from Escherichia coli . Cell 121, 703–712 (2005)

    Article  CAS  Google Scholar 

  20. Gualerzi, C. O. & Pon, C. L. Initiation of mRNA translation in prokaryotes. Biochemistry 29, 5881–5889 (1990)

    Article  CAS  Google Scholar 

  21. Antoun, A., Pavlov, M. Y., Tenson, T. & Ehrenberg, M. Ribosome formation from subunits studied by stopped-flow and Rayleigh light scattering. Biol. Proceed. Online 6, 35–54 (2004)

    Article  CAS  Google Scholar 

  22. Ali, I. K., Lancaster, L., Feinberg, J., Joseph, S. & Noller, H. F. Deletion of a conserved, central ribosomal intersubunit RNA bridge. Mol. Cell 23, 865–874 (2006)

    Article  CAS  Google Scholar 

  23. Milon, P., Konevega, A. L., Gualerzi, C. O. & Rodnina, M. V. Kinetic checkpoint at a late step in translation initiation. Mol. Cell 30, 712–720 (2008)

    Article  CAS  Google Scholar 

  24. Giuliodori, A. M., Giangrossi, M., Brandi, A., Gualerzi, C. O. & Pon, C. L. Cold-stress-induced de novo expression of infC and role of IF3 in cold-shock translational bias. RNA 13, 1355–1365 (2007)

    Article  CAS  Google Scholar 

  25. Klaholz, B. P. et al. Structure of the Escherichia coli ribosomal termination complex with release factor 2. Nature 421, 90–94 (2003)

    Article  ADS  CAS  Google Scholar 

  26. Marzi, S. et al. Structured mRNAs regulate translation initiation by binding to the platform of the ribosome. Cell 130, 1019–1031 (2007)

    Article  CAS  Google Scholar 

  27. Efron, B. Nonparametric estimates of standard error: The jackknife, the bootstrap and other methods. Biometrika 68, 589–599 (1981)

    Article  MathSciNet  Google Scholar 

  28. Haynor, D. R. & Woods, S. D. Resampling estimates of precision in emission tomography. IEEE Trans. Med. Imaging 8, 337–343 (1989)

    Article  CAS  Google Scholar 

  29. Penczek, P. A., Yang, C., Frank, J. & Spahn, C. M. Estimation of variance in single-particle reconstruction using the bootstrap technique. J. Struct. Biol. 154, 168–183 (2006)

    Article  CAS  Google Scholar 

  30. Rosenthal, P. B. & Henderson, R. Optimal determination of particle orientation, absolute hand, and contrast loss in single-particle electron cryomicroscopy. J. Mol. Biol. 333, 721–745 (2003)

    Article  CAS  Google Scholar 

  31. van Heel, M. & Schatz, M. Fourier shell correlation threshold criteria. J. Struct. Biol. 151, 250–262 (2005)

    Article  CAS  Google Scholar 

  32. Thompson, J. & Dahlberg, A. E. Testing the conservation of the translational machinery over evolution in diverse environments: assaying Thermus thermophilus ribosomes and initiation factors in a coupled transcription-translation system from Escherichia coli . Nucleic Acids Res. 32, 5954–5961 (2004)

    Article  CAS  Google Scholar 

  33. Rodnina, M. V., Semenkov, Y. P. & Wintermeyer, W. Purification of fMettRNA(fMet) by fast protein liquid chromatography. Anal. Biochem. 219, 380–381 (1994)

    Article  CAS  Google Scholar 

  34. Fechter, P. et al. Ribosomal initiation complexes probed by toeprinting and effect of trans-acting translational regulators in bacteria. Methods Mol. Biol. (in the press)

  35. Yusupova, G. Z., Yusupov, M. M., Cate, J. H. & Noller, H. F. The path of the messenger RNA throught the ribosome. Cell 106, 233–241 (2001)

    Article  CAS  Google Scholar 

  36. van Heel, M., Harauz, G., Orlova, E. V., Schmidt, R. & Schatz, M. A new generation of the IMAGIC image processing system. J. Struct. Biol. 116, 17–24 (1996)

    Article  CAS  Google Scholar 

  37. Orlov, I. M., Morgan, D. G. & Cheng, R. H. Efficient implementation of a filtered back-projection algorithm using a voxel-by-voxel approach. J. Struct. Biol. 154, 287–296 (2006)

    Article  CAS  Google Scholar 

Download references


We thank J. Thompson for providing IF2 strains, M. Argentini for mass spectroscopy analysis of IF1 and IF2, and P. Schultz, D. Moras, J.-C. Thierry, V. Mallouh, G. Yusupova and I. Orlov for their constant support and interest. This work was supported by grants from the Centre National pour la Recherche Scientifique (CNRS), the Ministère de la Recherche et de la Technologie, the European Molecular Biology Organization Young Investigator Programme, the Institut du Développement et des Ressources en Informatique Scientifique, and the European Commission as SPINE2-complexes (contract no LSHG-CT-2006-031220). A.S. is a PhD student in a co-tutorial between the Université Louis Pasteur (ULP) and the Università di Camerino, and was supported by SPINE2-complexes, by the Institut National de la Santé et de la Recherche Médicale (INSERM) and by the Fondation de la Recherche Médicale (FRM). S.M. was supported by postdoctoral fellowships from the ULP, the CNRS and from the FRM, and A.G.M. was a recipient of postdoctoral fellowships from the CNRS and the FRM. The electron microscope facility is supported by the Alsace Region, the INSERM, the CNRS and the Association pour la Recherche sur le Cancer.

Author information

Authors and Affiliations


Corresponding author

Correspondence to Bruno P. Klaholz.

Supplementary information

Supplementary Information

This file contains Supplementary Figures S1-S2 with legends, and Supplementary References. (PDF 404 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Simonetti, A., Marzi, S., Myasnikov, A. et al. Structure of the 30S translation initiation complex. Nature 455, 416–420 (2008).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


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