Abstract
In this chapter, we introduce a nanodisc-based experimental platform to study Ca2+-triggered membrane interaction of synaptotagmin-1. We describe and discuss in detail how to assemble this soluble mimetic of the docked vesicle–plasma membrane junction, with fluorescently labeled synaptotagmin-1 bound to trans SNAREpins assembled between nanodiscs and present the stopped-flow rapid mixing method used to monitor the conformational dynamics of Ca2+-activation process on a millisecond timescale.
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Acknowledgments
This work was supported by National Institute of Health (NIH) grant DK027044. We thank Jeff Coleman for critical inputs and suggestions.
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Stroeva, E., Krishnakumar, S.S. (2019). Using Nanodiscs to Probe Ca2+-Dependent Membrane Interaction of Synaptotagmin-1. In: Fratti, R. (eds) SNAREs. Methods in Molecular Biology, vol 1860. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8760-3_14
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DOI: https://doi.org/10.1007/978-1-4939-8760-3_14
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