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Single reconstituted neuronal SNARE complexes zipper in three distinct stages
Ying Gao1, Sylvain Zorman, Gregory Gundersen
1Department of Cell Biology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
Science (New York, N.Y.)
|August 21, 2012
Summary
Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins mediate membrane fusion. This study reveals a key intermediate in SNARE assembly, suggesting a stepwise, not continuous, zippering process driving fusion.
Area of Science:
- Biochemistry
- Cell Biology
- Biophysics
Background:
- Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins are essential for membrane fusion.
- SNAREs assemble into a four-helix bundle through a zippering mechanism.
Purpose of the Study:
- To observe and characterize a SNARE assembly intermediate in real time.
- To elucidate the step-by-step mechanism of SNARE-mediated membrane fusion.
Main Methods:
- Utilized optical tweezers in a cell-free reconstitution experiment.
- Applied external force to mimic membrane repulsion and stabilize intermediates.
Main Results:
- Observed a stable, half-assembled SNARE four-helix bundle intermediate.
- Demonstrated that SNARE zippering occurs through three sequential binary switches.
- Quantified the free energy released during fusion (36 k(B)T).
Conclusions:
- The zippering hypothesis is supported, but proceeds in discrete steps.
- A half-zippered intermediate plays a crucial role in membrane fusion.
- Externally applied force can stabilize and influence SNARE complex assembly.
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