Biological nanopores are an emerging class of biosensors with high-end precision owing to their reproducible fabrication at the nanometer scale. Most notably, nanopore-based DNA sequencing applications are currently being commercialized, while nanopore-based proteomics may become a reality in the near future. Although membrane proteins often prove to be difficult to purify, we describe a straightforward protocol for the preparation of Fragaceatoxin C (FraC) nanopores, which may have applications for DNA analysis and nanopore-based proteomics. Recombinantly expressed FraC nanopores are purified via two rounds of Ni-NTA affinity chromatography before and after oligomerization on sphingomyelin-containing liposomes. Starting from a plasmid vector containing the FraC gene, our method allows the production of purified nanopores within a week. Afterward, the FraC nanopores can be stored at +4 °C for several months, or frozen.
CITATION STYLE
Mutter, N. L., Huang, G., van der Heide, N. J., Lucas, F. L. R., Galenkamp, N. S., Maglia, G., & Wloka, C. (2021). Preparation of fragaceatoxin C (Fra C) nanopores. In Methods in Molecular Biology (Vol. 2186, pp. 3–10). Humana Press Inc. https://doi.org/10.1007/978-1-0716-0806-7_1
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