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Streptomycin interferes with conformational coupling between codon recognition and GTPase activation on the ribosome

Nature Structural & Molecular Biology volume 11, pages 316–322 (2004)Cite this article

Abstract

Aminoacyl-tRNAs (aa-tRNAs) are selected by the ribosome through a kinetically controlled induced fit mechanism. Cognate codon recognition induces a conformational change in the decoding center and a domain closure of the 30S subunit. We studied how these global structural rearrangements are related to tRNA discrimination by using streptomycin to restrict the conformational flexibility of the 30S subunit. The antibiotic stabilized aa-tRNA on the ribosome both with a cognate and with a near-cognate codon in the A site. Streptomycin altered the rates of GTP hydrolysis by elongation factor Tu (EF-Tu) on cognate and near-cognate codons, resulting in almost identical rates of GTP hydrolysis and virtually complete loss of selectivity. These results indicate that movements within the 30S subunit at the streptomycin-binding site are essential for the coupling between base pair recognition and GTP hydrolysis, thus modulating the fidelity of aa-tRNA selection.

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Figure 1: Kinetic scheme of EF-Tu-dependent aa-tRNA binding to the ribosomal A site.
Figure 2: Effect of streptomycin on the kinetics of A-site binding.
Figure 3: Effect of streptomycin and paromomycin on GTP hydrolysis.
Figure 4: Discrimination of aa-tRNA.

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Acknowledgements

We thank W. Wintermeyer for critical reading and discussions and V. Ramakrishnan for valuable comments on the manuscript. We thank A. Kubarenko for doing the footprinting experiment, V. Katunin and Y. Semenkov for tRNA preparations, U. Kothe for donating EF-Tu H84A, P. Striebeck, A. Böhm, C. Schillings and S. Möbitz for expert technical assistance. The work was supported by the Deutsche Forschungsgemeinschaft, the Alfried Krupp von Bohlen und Halbach-Stiftung and the Fonds der Chemischen Industrie.

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  1. Institute of Physical Biochemistry, University of Witten/Herdecke, Witten, D-58448, Germany

    Kirill B Gromadski & Marina V Rodnina

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Correspondence to Marina V Rodnina.

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Gromadski, K., Rodnina, M. Streptomycin interferes with conformational coupling between codon recognition and GTPase activation on the ribosome. Nat Struct Mol Biol 11, 316–322 (2004). https://doi.org/10.1038/nsmb742

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