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Biochem Biophys Res Commun. 2015 Oct 2;465(4):864-70. doi: 10.1016/j.bbrc.2015.08.111. Epub 2015 Aug 28.

Dynamic light scattering analysis of SNARE-driven membrane fusion and the effects of SNARE-binding flavonoids.

Author information

1
Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746, South Korea; Biomedical Research Institute, Korea Institute of Science and Technology, Seoul 136-791, South Korea.
2
Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746, South Korea.
3
Biomedical Research Institute, Korea Institute of Science and Technology, Seoul 136-791, South Korea.
4
Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746, South Korea. Electronic address: dhkweon@skku.edu.

Abstract

Soluble N-ethylmaleimide-sensitive-factor attachment protein receptor (SNARE) proteins generate energy required for membrane fusion. They form a parallelly aligned four-helix bundle called the SNARE complex, whose formation is initiated from the N terminus and proceeds toward the membrane-proximal C terminus. Previously, we have shown that this zippering-like process can be controlled by several flavonoids that bind to the intermediate structures formed during the SNARE zippering. Here, our aim was to test whether the fluorescence resonance energy transfer signals that are observed during the inner leaflet mixing assay indeed represent the hemifused vesicles. We show that changes in vesicle size accompanying the merging of bilayers is a good measure of progression of the membrane fusion. Two merging vesicles with the same size D in diameter exhibited their hydrodynamic diameters 2D + d (d, intermembrane distance), 2D and 2D as membrane fusion progressed from vesicle docking to hemifusion and full fusion, respectively. A dynamic light scattering assay of membrane fusion suggested that myricetin stopped membrane fusion at the hemifusion state, whereas delphinidin and cyanidin prevented the docking of the vesicles. These results are consistent with our previous findings in fluorescence resonance energy transfer assays.

KEYWORDS:

Dynamic light scattering; FRET; Flavonoid; Membrane fusion; SNARE

PMID:
26319432
DOI:
10.1016/j.bbrc.2015.08.111
[Indexed for MEDLINE]

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