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Large-scale growth of C. elegans and isolation of membrane protein complexes


Purification of membrane proteins for biochemical and structural studies is commonly achieved by recombinant overexpression in heterologous cell lines. However, many membrane proteins do not form a functional complex in a heterologous system, and few methods exist to purify sufficient protein from a native source for use in biochemical, biophysical and structural studies. Here, we provide a detailed protocol for the isolation of membrane protein complexes from transgenic Caenorhabditis elegans. We describe how to grow a genetically modified C. elegans line in abundance using standard laboratory equipment, and how to optimize purification conditions on a small scale using fluorescence-detection size-exclusion chromatography. Optimized conditions can then be applied to a large-scale preparation, enabling the purification of adequate quantities of a target protein for structural, biochemical and biophysical studies. Large-scale worm growth can be accomplished in ~9 d, and each optimization experiment can be completed in less than 1 d. We have used these methods to isolate the transmembrane channel-like protein 1 complex, as well as three additional protein complexes (transmembrane-like channel 2, lipid transfer protein and ‘Protein S’), from transgenic C. elegans, demonstrating the utility of this approach in purifying challenging, low-abundance membrane protein complexes.

Key points

  • This protocol outlines the isolation of membrane protein complexes from transgenic C. elegans

  • The primary advantage of the protocol is that it enables isolation of sufficient quantities of low-abundance native membrane protein complexes for use in structural, biochemical or biophysical studies

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Fig. 1: Timeline of large-scale worm growth and protein isolation.
Fig. 2: Diverse membrane protein complexes are amenable to isolation from transgenic C. elegans.
Fig. 3: Identifying regions of TMC-1–mVenus expression using spectral imaging and linear unmixing.
Fig. 4: Optimization of large-scale worm growth.
Fig. 5: Optimization of worm lysis and protein purification.
Fig. 6: Use of FSEC to estimate TMC-1 quality and quantity.

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All data generated or analyzed during this work are included in the published article.


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We thank members of the Gouaux, Aballay and Baconguis laboratories for helpful discussions; S. Petrie and B. Jenkins for help with worm spectral imaging; A. Chinn and M. Frisbie for help with worm growth; R. Hallford for proof reading and M. Freeman for suggesting studies on LPD-3. This work was supported by National Institutes of Health grant 1F32DC017894 to S.C. E.G. gratefully acknowledges J. LaCroute and B. LaCroute for support, and is an investigator of the HHMI.

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Authors and Affiliations



S.C., H.J. and A.G. performed the experiments. S.C., H.J., A.G. and Y.K., together with E.G., designed the project and wrote the manuscript. All authors contributed to manuscript preparation.

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Correspondence to Eric Gouaux.

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Nature Protocols thanks HaoSheng Sun and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Jeong, H. et al. Nature 610, 796–803 (2022):

Extended data

Extended Data Fig. 1 Comparison of sonication and cryo-bead milling methods for lysis of C. elegans.

Representative FSEC trace of affinity purified TMC-2-mVenus isolated from worms that were either lysed with cryo-bead milling (black traces) or sonication (blue traces). Data from three separate worm harvests are shown to illustrate that cryo-bead milling reproducibly results in a higher ratio of TMC-2 dimer to monomer.

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Clark, S., Jeong, H., Goehring, A. et al. Large-scale growth of C. elegans and isolation of membrane protein complexes. Nat Protoc 18, 2699–2716 (2023).

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