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Studying protein‐reconstituted proteoliposome fusion with content indicators in vitro
Author(s) -
Diao Jiajie,
Zhao Minglei,
Zhang Yunxiang,
Kyoung Minjoung,
Brunger Axel T.
Publication year - 2013
Publication title -
bioessays
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.175
H-Index - 184
eISSN - 1521-1878
pISSN - 0265-9247
DOI - 10.1002/bies.201300010
Subject(s) - mixing (physics) , fusion , lipid bilayer fusion , chemistry , vesicle , synaptotagmin 1 , lipid vesicle , in vitro , chromatography , liposome , membrane , biophysics , biochemistry , biology , physics , philosophy , linguistics , quantum mechanics , synaptic vesicle
In vitro reconstitution assays are commonly used to study biological membrane fusion. However, to date, most ensemble and single‐vesicle experiments involving SNARE proteins have been performed only with lipid‐mixing, but not content‐mixing indicators. Through simultaneous detection of lipid and small content‐mixing indicators, we found that lipid mixing often occurs seconds prior to content mixing, or without any content mixing at all, during a 50‐seconds observation period, for Ca 2+ ‐triggered fusion with SNAREs, full‐length synaptotagmin‐1, and complexin. Our results illustrate the caveats of commonly used bulk lipid‐mixing fusion experiments. We recommend that proteoliposome fusion experiments should always employ content‐mixing indicators in addition to, or in place of, lipid‐mixing indicators.