Reconstitution of 30S ribosomal subunits in vitro using ribosome biogenesis factors
- 1Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8562, Japan
- 2Laboratory for Cell-Free Protein Synthesis, RIKEN Quantitative Biology Center, Suita, Osaka 565-0874, Japan
- 3Institute for Medical Physics and Biophysics, Charité-Universitätsmedizin Berlin, 10117 Berlin, Germany
- Corresponding author: amikura{at}edu.k.u-tokyo.ac.jp
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↵4 These authors contributed equally to this work.
Abstract
Reconstitution of ribosomes in vitro from individual ribosomal proteins provides a powerful tool for understanding the ribosome assembly process including the sequential incorporation of ribosomal proteins. However, conventional assembly methods require high-salt conditions for efficient ribosome assembly. In this study, we reconstituted 30S ribosomal subunits from individually purified ribosomal proteins in the presence of ribosome biogenesis factors. In this system, two GTPases (Era and YjeQ) facilitated assembly of a 30S subunit exhibiting poly(U)-directed polyphenylalanine synthesis and native protein synthesis under physiological conditions. This in vitro system permits a study of the assembly process and function of ribosome biogenesis factors, and it will facilitate the generation of ribosomes from DNA without using cells.
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Article is online at http://www.rnajournal.org/cgi/doi/10.1261/rna.065615.118.
- Received March 2, 2018.
- Accepted July 25, 2018.
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