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Localization of synaptotagmin-binding domains on syntaxin

Y Kee and RH Scheller
Journal of Neuroscience 15 March 1996, 16 (6) 1975-1981; https://doi.org/10.1523/JNEUROSCI.16-06-01975.1996
Y Kee
Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University Medical Center, California 94305, USA.
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RH Scheller
Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University Medical Center, California 94305, USA.
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Abstract

Synaptotagmin, an abundant calcium- and phospholipid-binding protein of synaptic vesicles, has been proposed to regulate neurotransmitter release at the nerve terminal. To understand better the biochemical mechanism of neurotransmitter release, we have investigated the calcium- dependent and -independent protein-protein interactions between synaptotagmin I and syntaxin 1a, a subunit of the receptor for synaptic vesicles on the presynaptic plasma membrane. Soluble syntaxin 1a binds to synaptotagmin glutathione S-transferase (GST) fusion protein, and the binding was decreased in the presence of calcium. A synaptotagmin fragment containing the second C2 repeat (Syt3–5) had the same binding profile as the whole cytoplasmic domain; however, fragments containing the first C2 repeat (Syt1–3 and Syt2–3) showed calcium-dependent binding to syntaxin. In addition, the soluble full-length cytoplasmic domain of synaptotagmin binds to a syntaxin GST fusion protein in a calcium-dependent manner. Syntaxin domains required for calcium- dependent and -independent synaptotagmin-binding were localized using syntaxin deletion mutants. Amino acids 241–266 of the syntaxin C terminus were required for calcium-independent binding of synaptotagmin. The minimal domain required for calcium-dependent binding of synaptotagmin to syntaxin was localized to amino acids 220– 266. The syntaxin domains required for synaptotagmin binding overlap with the domains for vesicle-associated membrane protein (or VAMP) and alpha-soluble N-ethyl-maleimide-sensitive fusion protein attachment protein (or alphaSNAP) interactions. The data suggest both calcium- dependent and -independent roles of synaptotagmin in regulating synaptic vesicle release and/or recycling.

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The Journal of Neuroscience: 16 (6)
Journal of Neuroscience
Vol. 16, Issue 6
15 Mar 1996
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Localization of synaptotagmin-binding domains on syntaxin
Y Kee, RH Scheller
Journal of Neuroscience 15 March 1996, 16 (6) 1975-1981; DOI: 10.1523/JNEUROSCI.16-06-01975.1996

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Localization of synaptotagmin-binding domains on syntaxin
Y Kee, RH Scheller
Journal of Neuroscience 15 March 1996, 16 (6) 1975-1981; DOI: 10.1523/JNEUROSCI.16-06-01975.1996
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