Abstract
Non-vesicular lipid transport between bilayers at membrane contact sites plays important physiological roles. Mechanistic insight into the action of lipid-transport proteins localized at these sites requires determination of the distance between bilayers at which this transport can occur. Here we developed DNA-origami nanostructures to organize size-defined liposomes at precise distances and used them to study lipid transfer by the synaptotagmin-like mitochondrial lipid-binding protein (SMP) domain of extended synaptotagmin 1 (E-Syt1). Pairs of DNA-ring-templated donor and acceptor liposomes were docked through DNA pillars, which determined their distance. The SMP domain was anchored to donor liposomes via an unstructured linker, and lipid transfer was assessed via a Förster resonance energy transfer (FRET)-based assay. We show that lipid transfer can occur over distances that exceed the length of an SMP dimer, which is compatible with the shuttle model of lipid transport. The DNA nanostructures developed here can also be adapted to study other processes occurring where two membranes are closely apposed to each other.
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Acknowledgements
We thank Y. Cai for discussion. This work was supported by NIH grants NS036251 and DA018343, the HHMI and the Kavli Foundation to P.D.C.; an NIH Director’s New Innovator Award (GM114830) and a Yale University faculty startup fund to C.L.; a Human Frontier Science Program Long-term Fellowship to X.B.; and an Agency for Science, Technology and Research Graduate Scholarship (Singapore) to Q.X.
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X.B. designed and performed liposome tethering and lipid-transfer assays, and prepared DNA-origami-organized liposomes. Z.Z. designed the DNA-origami structures, prepared DNA-origami-organized liposomes and performed EM studies. Q.X. performed EM studies. X.B., Z.Z., P.D.C. and C.L. initiated the project, analyzed and interpreted data, and wrote the manuscript.
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Bian, X., Zhang, Z., Xiong, Q. et al. A programmable DNA-origami platform for studying lipid transfer between bilayers. Nat Chem Biol 15, 830–837 (2019). https://doi.org/10.1038/s41589-019-0325-3
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DOI: https://doi.org/10.1038/s41589-019-0325-3
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