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Cited by 231RNA and protein synthesis mechanismsOrigins and Evolution of LifeToxin Mechanisms and Immunotoxins

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Protein synthesis by ribosomes with tethered subunits

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Orelle, Cédric, Carlson, Erik D., Szal, Teresa, Florin, Tanja, Jewett, Michael C., Mankin, Alexander S.·2015·Nature
DOI 10.1038/nature14862
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Abstract

The ribosome is a ribonucleoprotein machine responsible for protein synthesis. In all kingdoms of life it is composed of two subunits, each built on its own ribosomal RNA (rRNA) scaffold. The independent but coordinated functions of the subunits, including their ability to associate at initiation, rotate during elongation, and dissociate after protein release, are an established model of protein synthesis. Furthermore, the bipartite nature of the ribosome is presumed to be essential for biogenesis, since dedicated assembly factors keep immature ribosomal subunits apart and prevent them from translation initiation. Free exchange of the subunits limits the development of specialized orthogonal genetic systems that could be evolved for novel functions without interfering with native translation. Here we show that ribosomes with tethered and thus inseparable subunits (termed Ribo-T) are capable of successfully carrying out protein synthesis. By engineering a hybrid rRNA composed of both small and large subunit rRNA sequences, we produced a functional ribosome in which the subunits are covalently linked into a single entity by short RNA linkers. Notably, Ribo-T was not only functional in vitro, but was also able to support the growth of Escherichia coli cells even in the absence of wild-type ribosomes. We used Ribo-T to create the first fully orthogonal ribosome-messenger RNA system, and demonstrate its evolvability by selecting otherwise dominantly lethal rRNA mutations in the peptidyl transferase centre that facilitate the translation of a problematic protein sequence. Ribo-T can be used for exploring poorly understood functions of the ribosome, enabling orthogonal genetic systems, and engineering ribosomes with new functions.

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References 39 · Cited by 231

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Metadata

Authors
Orelle, C., Carlson, E. D., Szal, T., Florin, T., Jewett, M. C., & Mankin, A. S.
Affiliations
  • University of Illinois Chicago (US)
  • Northwestern University (US)
Corresponding author
  • Jewett, Michael C.
  • Mankin, Alexander S.
ORCID
Venue
Nature
Publisher
Springer Science and Business Media LLC
Published
2015-07-29
Volume
524
Issue
7563
Pages
119-124
ISSN
  • 0028-0836 (ISSN-L)
  • 1476-4687
DOI
10.1038/nature14862
OpenAlex
W2162778008
Cited by
231
References
39
Citations by year
2026: 10 · 2025: 11 · 2024: 21 · 2023: 22 · 2022: 16 · 2021: 21 · 2020: 26 · 2019: 25 · 2018: 20 · 2017: 26 · 2016: 24 · 2015: 9
FWCI
10.39
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature (publishedVersion)
  • PubMed (publishedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • Zenodo (CERN European Organization for Nuclear Research) (submittedVersion, other-oa)
Funders
  • National Science Foundation (1244455, N66001-12-C-4211, 0943393)
  • Advanced Research Projects Agency
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
  • Physical Sciences › Physics and Astronomy › Astronomy and Astrophysics › Origins and Evolution of Life
  • Life Sciences › Immunology and Microbiology › Immunology › Toxin Mechanisms and Immunotoxins
Keywords
ribosome engineering, protein synthesis, ribosome biogenesis, peptidyl transferase center, ribosome evolution, Escherichia coli, translation initiation, ribosomal subunit assembly
MeSH terms
Amino Acid Sequence, Base Sequence, Escherichia coli, Models, Molecular, Molecular Conformation, Mutation, Peptidyl Transferases, Ribosomes, RNA, Messenger, RNA, Ribosomal, 23S, Protein Biosynthesis, Protein Subunits, Bioengineering

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