The ribosomal stalk binds to translation factors IF2, EF-Tu, EF-G and RF3 via a conserved region of the L12 C-terminal domain
(2007) In Journal of Molecular Biology 365(2). p.468-479- Abstract
- Efficient protein synthesis in bacteria requires initiation factor 2 (IF2), elongation factors Tu (EF-Tu) and G (EF-G), and release factor 3 (RF3), each of which catalyzes a major step of translation in a GTP-dependent fashion. Previous reports have suggested that recruitment of factors to the ribosome and subsequent GTP hydrolysis involve the dimeric protein L12, which forms a flexible "stalk" on the ribosome. Using heteronuclear NMR spectroscopy we demonstrate that L12 binds directly to the factors IF2, EF-Tu, EF-G, and RF3 from Escherichia coli, and map the region of L12 involved in these interactions. Factor-dependent chemical shift changes show that all four factors bind to the same region of the C-terminal domain of L12. This region... (More)
- Efficient protein synthesis in bacteria requires initiation factor 2 (IF2), elongation factors Tu (EF-Tu) and G (EF-G), and release factor 3 (RF3), each of which catalyzes a major step of translation in a GTP-dependent fashion. Previous reports have suggested that recruitment of factors to the ribosome and subsequent GTP hydrolysis involve the dimeric protein L12, which forms a flexible "stalk" on the ribosome. Using heteronuclear NMR spectroscopy we demonstrate that L12 binds directly to the factors IF2, EF-Tu, EF-G, and RF3 from Escherichia coli, and map the region of L12 involved in these interactions. Factor-dependent chemical shift changes show that all four factors bind to the same region of the C-terminal domain of L12. This region includes three strictly conserved residues, K70, L80, and E82, and a set of highly conserved residues, including V66, A67, V68 and G79. Upon factor binding, all NMR signals from the C-terminal domain become broadened beyond detection, while those from the N-terminal domain are virtually unaffected, implying that the C-terminal domain binds to the factor, while the N-terminal domain dimer retains its rotational freedom mediated by the flexible hinge between the two domains. Factor-dependent variations in linewidths further reveal that L12 binds to each factor with a dissociation constant in the millimolar range in solution. These results indicate that the L12-factor complexes will be highly populated on the ribosome, because of the high local concentration of ribosome-bound factor with respect to L12. (c) 2006 Elsevier Ltd. All rights reserved. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/679314
- author
- Helgstrand, Magnus LU ; Mandava, Chandra S. ; Mulder, Frans LU ; Liljas, Anders LU ; Sanyal, Suparna LU and Akke, Mikael LU
- organization
- publishing date
- 2007
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- NMR spectroscopy, GTPase, L12, ribosome, protein synthesis
- in
- Journal of Molecular Biology
- volume
- 365
- issue
- 2
- pages
- 468 - 479
- publisher
- Elsevier
- external identifiers
-
- wos:000243243200015
- scopus:33751532962
- pmid:17070545
- ISSN
- 1089-8638
- DOI
- 10.1016/j.jmb.2006.10.025
- language
- English
- LU publication?
- yes
- id
- 2a2aa3a4-6507-4f71-aaeb-73f77bb25ae6 (old id 679314)
- date added to LUP
- 2016-04-01 15:37:18
- date last changed
- 2022-03-14 19:05:26
@article{2a2aa3a4-6507-4f71-aaeb-73f77bb25ae6, abstract = {{Efficient protein synthesis in bacteria requires initiation factor 2 (IF2), elongation factors Tu (EF-Tu) and G (EF-G), and release factor 3 (RF3), each of which catalyzes a major step of translation in a GTP-dependent fashion. Previous reports have suggested that recruitment of factors to the ribosome and subsequent GTP hydrolysis involve the dimeric protein L12, which forms a flexible "stalk" on the ribosome. Using heteronuclear NMR spectroscopy we demonstrate that L12 binds directly to the factors IF2, EF-Tu, EF-G, and RF3 from Escherichia coli, and map the region of L12 involved in these interactions. Factor-dependent chemical shift changes show that all four factors bind to the same region of the C-terminal domain of L12. This region includes three strictly conserved residues, K70, L80, and E82, and a set of highly conserved residues, including V66, A67, V68 and G79. Upon factor binding, all NMR signals from the C-terminal domain become broadened beyond detection, while those from the N-terminal domain are virtually unaffected, implying that the C-terminal domain binds to the factor, while the N-terminal domain dimer retains its rotational freedom mediated by the flexible hinge between the two domains. Factor-dependent variations in linewidths further reveal that L12 binds to each factor with a dissociation constant in the millimolar range in solution. These results indicate that the L12-factor complexes will be highly populated on the ribosome, because of the high local concentration of ribosome-bound factor with respect to L12. (c) 2006 Elsevier Ltd. All rights reserved.}}, author = {{Helgstrand, Magnus and Mandava, Chandra S. and Mulder, Frans and Liljas, Anders and Sanyal, Suparna and Akke, Mikael}}, issn = {{1089-8638}}, keywords = {{NMR spectroscopy; GTPase; L12; ribosome; protein synthesis}}, language = {{eng}}, number = {{2}}, pages = {{468--479}}, publisher = {{Elsevier}}, series = {{Journal of Molecular Biology}}, title = {{The ribosomal stalk binds to translation factors IF2, EF-Tu, EF-G and RF3 via a conserved region of the L12 C-terminal domain}}, url = {{http://dx.doi.org/10.1016/j.jmb.2006.10.025}}, doi = {{10.1016/j.jmb.2006.10.025}}, volume = {{365}}, year = {{2007}}, }