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Cost-effective large-scale expression of proteins for NMR studies
Authors:Julia Klopp  Aurélie Winterhalter  Rémy Gébleux  Daniela Scherer-Becker  Christian Ostermeier  Alvar D Gossert
Institution:1.Novartis AG,Basel,Switzerland;2.Institute for Molecular Biology and Biophysics,ETH Zürich,Zurich,Switzerland;3.Biomolecular NMR Spectroscopy Platform,ETH Zürich,Zurich,Switzerland
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.
Keywords:
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