Studying protein-reconstituted proteoliposome fusion with content indicators in vitro
- PMID: 23625805
- PMCID: PMC4453005
- DOI: 10.1002/bies.201300010
Studying protein-reconstituted proteoliposome fusion with content indicators in vitro
Abstract
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.
© 2013 WILEY Periodicals, Inc.
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