MemStar: A one-shot Escherichia coli-based approach for high-level bacterial membrane protein production |
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| Authors: | Chiara Lee Hae Joo Kang Anna Hjelm Abdul Aziz Qureshi Emmanuel Nji Hassanul Choudhury Konstantinos Beis Jan-Willem de Gier David Drew |
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| Affiliation: | 1. Division of Molecular Biosciences, Imperial College London, London SW7 2AZ, UK;2. Centre for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden;3. Membrane Protein Laboratory, Diamond Light Source, Harwell Science and Innovation Campus, Didcot, Oxfordshire OX11 0DE, UK;4. Rutherford Appleton Laboratory, Research Complex at Harwell, Didcot, Oxfordshire OX11 0DE, UK |
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| Abstract: | Optimising membrane protein production yields in Escherichiacoli can be time- and resource-consuming. Here, we present a simple and effective Membrane protein Single shot amplification recipe: MemStar. This one-shot amplification recipe is based on the E. coli strain Lemo21(DE3), the PASM-5052 auto-induction medium and, contradictorily, an IPTG induction step. Using MemStar, production yields for most bacterial membrane proteins tested were improved to reach an average of 5 mg L−1 per OD600 unit, which is significantly higher than yields obtained with other common production strategies. With MemStar, we have been able to obtain new structural information for several transporters, including the sodium/proton antiporter NapA. |
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| Keywords: | Membrane protein production X-ray crystallography High-throughput Escherichia coli |
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