Abstract
We present protocols for high-level expression of isotope-labelled proteins in E. coli in cost-effective ways. This includes production of large amounts of unlabeled proteins and 13C-methyl methionine labeling in rich media, where yields of up to a gram of soluble protein per liter of culture are reached. Procedures for uniform isotope labeling of 2H, 13C and 15N using auto-induction or isopropyl-β-d-1-thiogalactopyranoside-induction are described, with primary focus on minimal isotope consumption and high reproducibility of protein expression. These protocols are based on high cell-density fermentation, but the key procedures are easily transferred to shake flask cultures.
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Acknowledgements
The authors would like to thank Roger Benoit and Paul Erbel for helpful discussions on the protocols.
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The authors declare that they have no conflict of interest. ADG, JK and AW co-developed the cleaning in place module (LabCIP) for fermenters together with the company Infors HT. There are no financial benefits for any of the named persons connected to Infors HT.
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Klopp, J., Winterhalter, A., Gébleux, R. et al. Cost-effective large-scale expression of proteins for NMR studies. J Biomol NMR 71, 247–262 (2018). https://doi.org/10.1007/s10858-018-0179-0
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DOI: https://doi.org/10.1007/s10858-018-0179-0