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  Positively charged amino acids at the SNAP-25 C terminus determine fusion rates, fusion pore properties, and energetics of tight SNARE complex zippering.

Fang, Q., Zhao, Y., Herbst, A. D., Kim, B. N., & Lindau, M. (2015). Positively charged amino acids at the SNAP-25 C terminus determine fusion rates, fusion pore properties, and energetics of tight SNARE complex zippering. The Journal of Neuroscience, 35(7), 3230-3239. doi:10.1523/JNEUROSCI.2905-14.2015.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0025-010D-2 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002A-0E26-8
Genre: Journal Article

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 Creators:
Fang, Q.1, Author              
Zhao, Y.1, Author              
Herbst, A. D., Author
Kim, B. N., Author
Lindau, M.1, Author              
Affiliations:
1Research Group of Nanoscale Cell Biology, MPI for Biophysical Chemistry, Max Planck Society, ou_1832294              

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Free keywords: SNAP-25; amperometry; capacitance; chromaffin cells; exocytosis; fusion pore
 Abstract: SNAP-25 is a Q-SNARE protein mediating exocytosis of neurosecretory vesicles including chromaffin granules. Previous results with a SNAP-25 construct lacking the nine C terminal residues (SNAP-25Δ9) showed changed fusion pore properties (Fang et al., 2008), suggesting a model for fusion pore mechanics that couple C terminal zipping of the SNARE complex to the opening of the fusion pore. The deleted fragment contains the positively charged residues R198 and K201, adjacent to layers 7 and 8 of the SNARE complex. To determine how fusion pore conductance and dynamics depend on these residues, single exocytotic events in bovine chromaffin cells expressing R198Q, R198E, K201Q, or K201E mutants were investigated by carbon fiber amperometry and cell-attached patch capacitance measurements. Coarse grain molecular dynamics simulations revealed spontaneous transitions between a loose and tightly zippered state at the SNARE complex C terminus. The SNAP-25 K201Q mutant showed no changes compared with SNAP-25 wild-type. However, K201E, R198Q, and R198E displayed reduced release frequencies, slower release kinetics, and prolonged fusion pore duration that were correlated with reduced probability to engage in the tightly zippered state. The results show that the positively charged amino acids at the SNAP-25 C terminus promote tight SNARE complex zippering and are required for high release frequency and rapid release in individual fusion events.

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Language(s): eng - English
 Dates: 2015-02-182015-02
 Publication Status: Published in print
 Pages: -
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 Table of Contents: -
 Rev. Method: Peer
 Identifiers: DOI: 10.1523/JNEUROSCI.2905-14.2015
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Title: The Journal of Neuroscience
Source Genre: Journal
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Pages: - Volume / Issue: 35 (7) Sequence Number: - Start / End Page: 3230 - 3239 Identifier: -