Magnetic Resonance
Magnetic Resonance
MR
Articles | Volume 4, issue 2
Articles | Volume 4, issue 2
https://doi.org/10.5194/mr-4-187-2023
© Author(s) 2023. This work is distributed under
the Creative Commons Attribution 4.0 License.
https://doi.org/10.5194/mr-4-187-2023
© Author(s) 2023. This work is distributed under
the Creative Commons Attribution 4.0 License.
Articles | Volume 4, issue 2
Research article
 | 
19 Jul 2023
Research article |  | 19 Jul 2023

Cell-free synthesis of proteins with selectively 13C-labelled methyl groups from inexpensive precursors

Damian Van Raad, Gottfried Otting, and Thomas Huber

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Cited articles

Amero, C., Asunción Durá, M., Noirclerc-Savoye, M., Perollier, A., Gallet, B., Plevin, M. J., Vernet, T., Franzetti, B., and Boisbouvier, J.: A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies, J. Biomol. NMR, 50, 229–236, https://doi.org/10.1007/s10858-011-9513-5, 2011. 
Apponyi, M. A., Ozawa, K., Dixon, N. E., and Otting, G.: Cell-free protein synthesis for analysis by NMR spectroscopy, Method. Mol. Biol., 426, 257–268, https://doi.org/10.1007/978-1-60327-058-8_16, 2008. 
Behera, S. P., Dubey, A., Chen, W. N., De Paula, V. S., Zhang, M., Sgourakis, N. G., Bermel, W., Wagner, G., Coote, P. W., and Arthanari, H.: Nearest-neighbor NMR spectroscopy: categorizing spectral peaks by their adjacent nuclei, Nat. Commun., 11, 5547, https://doi.org/10.1038/s41467-020-19325-4, 2020. 
Boswell, Z. K. and Latham, M. P.: Methyl-based NMR spectroscopy methods for uncovering structural dynamics in large proteins and protein complexes, Biochemistry, 58, 144-155, https://doi.org/10.1021/acs.biochem.8b00953, 2018. 
Caschera, F. and Noireaux, V.: A cost-effective polyphosphate-based metabolism fuels an all E. coli cell-free expression system, Metabol. Engin., 27, 29–37, https://doi.org/10.1016/j.ymben.2014.10.007, 2015. 
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Short summary
A novel cell-free protein synthesis system called eCells produces amino acids based on specific isotopes using low-cost precursors. The system selectively labels methyl groups, i.e valine and leucine, with high efficiency. eCells achieve high levels of 13C incorporation and deuteration in protein preparations, making them suitable for NMR experiments of large protein complexes. They are easy to prepare, can be scaled up in volume and are a promising tool for protein production and NMR studies.
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