Syt1 (synaptotagmin 1), a major Ca2+ sensor for fast neurotransmitter release, contains tandem Ca2+-binding C2 domains (C2AB), a single transmembrane α-helix and a highly charged 60-residue-long linker in between. Using single-vesicle-docking and content-mixing assays we found that the linker region of Syt1 is essential for its two signature functions: Ca2+-independent vesicle docking and Ca2+-dependent fusion pore opening. The linker contains the basic-amino-acid-rich N-terminal region and the acidic-amino-acid-rich C-terminal region. When the charge segregation was disrupted, fusion pore opening was slowed, whereas docking was unchanged. Intramolecular disulfide cross-linking between N- and C-terminal regions of the linker or deletion of 40 residues from the linker reduced docking while enhancing pore opening, although the changes were subtle. EPR analysis showed Ca2+-induced line broadening reflecting a conformational change in the linker region. Thus the results of the present study suggest that the electrostatically bipartite linker region may extend for docking and fold to facilitate pore opening.
The synaptotagmin 1 linker may function as an electrostatic zipper that opens for docking but closes for fusion pore opening
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Ying Lai, Xiaochu Lou, Yongseok Jho, Tae-Young Yoon, Yeon-Kyun Shin; The synaptotagmin 1 linker may function as an electrostatic zipper that opens for docking but closes for fusion pore opening. Biochem J 15 November 2013; 456 (1): 25–33. doi: https://doi.org/10.1042/BJ20130949
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