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Single-Molecule Fluorescence Measurement of SNARE-Mediated Vesicle Fusion

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SNAREs

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1860))

Abstract

This chapter expounds the single vesicle-vesicle fluorescence resonance energy transfer (FRET) measurement to study the membrane fusion mediated by SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins. The formation of a four-α-helix bundle of SNARE proteins can drive two membranes to a close proximity for fusion. Through single-molecule FRET-based microscopy, the lipid-mixing process at the single-vesicle level can be tracked in real time. This reconstitution system is applicable to study the molecular mechanism of SNAREs during different membrane fusion stages, such as docking, hemifusion, and full fusion. Four main parts are described in this chapter, including SNARE reconstitution, imaging preparation, data collection, and analysis.

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Acknowledgments

This work was supported by the 973-program (2015CB856304) and National Institutes of Health (R35GM128837).

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Authors and Affiliations

  1. Department of Cancer Biology, University of Cincinnati College of Medicine, Cincinnati, OH, USA

    Yachong Hu, Zhiqi Tian & Jiajie Diao

  2. School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, China

    Yachong Hu & Zhiqi Tian

Authors
  1. Yachong Hu
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  2. Zhiqi Tian
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  3. Jiajie Diao
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Corresponding author

Correspondence to Jiajie Diao .

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Editors and Affiliations

  1. Department of Biochemistry, University of Illinois Urbana-Champaign, Urbana, IL, USA

    Rutilio Fratti

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Hu, Y., Tian, Z., Diao, J. (2019). Single-Molecule Fluorescence Measurement of SNARE-Mediated Vesicle Fusion. 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_22

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  • DOI: https://doi.org/10.1007/978-1-4939-8760-3_22

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8759-7

  • Online ISBN: 978-1-4939-8760-3

  • eBook Packages: Springer Protocols

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