{"id":{"repo_id":"tdl","oai_identifier":"oai:tdl-ir.tdl.org:2152.5/227"},"canonical_url":"https://search.dev.ndltd.org/etd/tdl/oai:tdl-ir.tdl.org:2152.5/227","repository":{"repo_id":"tdl","name":"Texas Digital Library","base_url":"https://tdl-ir.tdl.org/server/oai/request"},"display":{"title":"Mechanism of Synaptotagmin Action in Neurotransmitter Release","abstract":"Neurotransmitter release occurs by fusion of the synaptic vesicle membrane with the plasma membrane. Formation of a highly stable complex, known as the SNARE (soluble NSF-attachment protein receptors) complex, brings the two membranes close in space. SNARE complex formation is required but probably not sufficient for fusion to occur. An increase in the local Ca2+ concentration at the synaptic terminal rapidly triggers neurotransmitter release. The mechanism of Ca2+ action is still unknown. Synaptotagmin 1, a brain-specific vesicular transmembrane protein, is the Ca2+ sensor in neurons. It has two cytoplasmic C2 domains (C2A and C2B) that bind Ca2+. Both C2 domains interact with negatively charged phospholipids in a Ca2+ dependent manner. The interaction of synaptotagmin 1 with the SNARE complex is also reported. We investigated whether the interaction of synaptotagmin 1 with membranes or with the SNARE complex is critical for membrane fusion. A new method to detect protein-protein interactions by 1D NMR spectroscopy was developed. Either the 13C signal of the SNARE complex or synaptotagmin 1 was monitored to perform competition experiments between SNAREs and lipid vesicles for binding to synaptotagmin 1. In the presence of both lipids and the SNARE complex, synaptotagmin 1 binds to lipids but cannot bind to the SNARE complex. This result suggests that Ca2+-dependent membrane binding is the primary activity of synaptotagmin 1. We investigated the mechanism of Ca2+-dependent phospholipid binding to synaptotagmin 1 C2 domains. A combination of crosslinking and FRET experiments showed that synaptotagmin 1 does not oligomerize upon Ca2+-dependent binding to phospholipid vesicles. Intriguingly, it binds to two membranes simultaneously and brings them into close proximity as visualized by cryo-EM experiments. We showed that the isolated C2B domain is sufficient to induce close membrane proximity. Mutational analysis suggested that the abundance of basic residues around the C2B surface, which generates a highly positive electrostatic potential together with the bound Ca2+ ions, is essential for this activity. We suggest that the ability of the C2B domain to bring membranes into close proximity can explain why the C2B domain has a more critical function in vivo than the C2A domain.nn","abstract_html":"Neurotransmitter release occurs by fusion of the synaptic vesicle membrane with the plasma membrane. Formation of a highly stable complex, known as the SNARE (soluble NSF-attachment protein receptors) complex, brings the two membranes close in space. SNARE complex formation is required but probably not sufficient for fusion to occur. An increase in the local Ca2+ concentration at the synaptic terminal rapidly triggers neurotransmitter release. The mechanism of Ca2+ action is still unknown. Synaptotagmin 1, a brain-specific vesicular transmembrane protein, is the Ca2+ sensor in neurons. It has two cytoplasmic C2 domains (C2A and C2B) that bind Ca2+. Both C2 domains interact with negatively charged phospholipids in a Ca2+ dependent manner. The interaction of synaptotagmin 1 with the SNARE complex is also reported. We investigated whether the interaction of synaptotagmin 1 with membranes or with the SNARE complex is critical for membrane fusion. A new method to detect protein-protein interactions by 1D NMR spectroscopy was developed. Either the 13C signal of the SNARE complex or synaptotagmin 1 was monitored to perform competition experiments between SNAREs and lipid vesicles for binding to synaptotagmin 1. In the presence of both lipids and the SNARE complex, synaptotagmin 1 binds to lipids but cannot bind to the SNARE complex. This result suggests that Ca2+-dependent membrane binding is the primary activity of synaptotagmin 1. We investigated the mechanism of Ca2+-dependent phospholipid binding to synaptotagmin 1 C2 domains. A combination of crosslinking and FRET experiments showed that synaptotagmin 1 does not oligomerize upon Ca2+-dependent binding to phospholipid vesicles. Intriguingly, it binds to two membranes simultaneously and brings them into close proximity as visualized by cryo-EM experiments. We showed that the isolated C2B domain is sufficient to induce close membrane proximity. Mutational analysis suggested that the abundance of basic residues around the C2B surface, which generates a highly positive electrostatic potential together with the bound Ca2+ ions, is essential for this activity. We suggest that the ability of the C2B domain to bring membranes into close proximity can explain why the C2B domain has a more critical function in vivo than the C2A domain.nn","abstract_has_math":false,"creators":["Arac-Ozkan, Demet"],"institution":null,"degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Rizo-Rey, José"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2005,"date_issued":"2005-12-19","date_published":"2005-12-19","updated_at":"2026-07-27T21:19:37Z","subjects":["SNARE Proteins","Synaptotagmin I","Cytoplasm"],"languages":["en"],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["65338227"],"render_values":[{"text":"65338227","href":null,"code":true}]}]},"links":{"outbound_url":"https://hdl.handle.net/2152.5/227","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Rizo-Rey, José"]},{"key":"dc:creator","label":"Author","values":["Arac-Ozkan, Demet"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2010-07-12T16:22:15Z","2026-03-25T17:04:14Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2010-07-12T16:22:15Z"]},{"key":"dc:date.issued","label":"Date","values":["2005-12-19"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["SNARE Proteins","Synaptotagmin I","Cytoplasm"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://hdl.handle.net/2152.5/227","65338227"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://hdl.handle.net/2152.5/227"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Neurotransmitter release occurs by fusion of the synaptic vesicle membrane with the plasma membrane. Formation of a highly stable complex, known as the SNARE (soluble NSF-attachment protein receptors) complex, brings the two membranes close in space. SNARE complex formation is required but probably not sufficient for fusion to occur. An increase in the local Ca2+ concentration at the synaptic terminal rapidly triggers neurotransmitter release. The mechanism of Ca2+ action is still unknown. Synaptotagmin 1, a brain-specific vesicular transmembrane protein, is the Ca2+ sensor in neurons. It has two cytoplasmic C2 domains (C2A and C2B) that bind Ca2+. Both C2 domains interact with negatively charged phospholipids in a Ca2+ dependent manner. The interaction of synaptotagmin 1 with the SNARE complex is also reported. We investigated whether the interaction of synaptotagmin 1 with membranes or with the SNARE complex is critical for membrane fusion. A new method to detect protein-protein interactions by 1D NMR spectroscopy was developed. Either the 13C signal of the SNARE complex or synaptotagmin 1 was monitored to perform competition experiments between SNAREs and lipid vesicles for binding to synaptotagmin 1. In the presence of both lipids and the SNARE complex, synaptotagmin 1 binds to lipids but cannot bind to the SNARE complex. This result suggests that Ca2+-dependent membrane binding is the primary activity of synaptotagmin 1. We investigated the mechanism of Ca2+-dependent phospholipid binding to synaptotagmin 1 C2 domains. A combination of crosslinking and FRET experiments showed that synaptotagmin 1 does not oligomerize upon Ca2+-dependent binding to phospholipid vesicles. Intriguingly, it binds to two membranes simultaneously and brings them into close proximity as visualized by cryo-EM experiments. We showed that the isolated C2B domain is sufficient to induce close membrane proximity. Mutational analysis suggested that the abundance of basic residues around the C2B surface, which generates a highly positive electrostatic potential together with the bound Ca2+ ions, is essential for this activity. We suggest that the ability of the C2B domain to bring membranes into close proximity can explain why the C2B domain has a more critical function in vivo than the C2A domain.nn"]},{"key":"dc:title","label":"Title","values":["Mechanism of Synaptotagmin Action in Neurotransmitter Release"]}]}],"canonical_facts":{"dc:contributor":["Rizo-Rey, José"],"dc:creator":["Arac-Ozkan, Demet"],"dc:date.accessioned":["2010-07-12T16:22:15Z","2026-03-25T17:04:14Z"],"dc:date.available":["2010-07-12T16:22:15Z"],"dc:date.issued":["2005-12-19"],"dc:description.abstract":["Neurotransmitter release occurs by fusion of the synaptic vesicle membrane with the plasma membrane. Formation of a highly stable complex, known as the SNARE (soluble NSF-attachment protein receptors) complex, brings the two membranes close in space. SNARE complex formation is required but probably not sufficient for fusion to occur. An increase in the local Ca2+ concentration at the synaptic terminal rapidly triggers neurotransmitter release. The mechanism of Ca2+ action is still unknown. Synaptotagmin 1, a brain-specific vesicular transmembrane protein, is the Ca2+ sensor in neurons. It has two cytoplasmic C2 domains (C2A and C2B) that bind Ca2+. Both C2 domains interact with negatively charged phospholipids in a Ca2+ dependent manner. The interaction of synaptotagmin 1 with the SNARE complex is also reported. We investigated whether the interaction of synaptotagmin 1 with membranes or with the SNARE complex is critical for membrane fusion. A new method to detect protein-protein interactions by 1D NMR spectroscopy was developed. Either the 13C signal of the SNARE complex or synaptotagmin 1 was monitored to perform competition experiments between SNAREs and lipid vesicles for binding to synaptotagmin 1. In the presence of both lipids and the SNARE complex, synaptotagmin 1 binds to lipids but cannot bind to the SNARE complex. This result suggests that Ca2+-dependent membrane binding is the primary activity of synaptotagmin 1. We investigated the mechanism of Ca2+-dependent phospholipid binding to synaptotagmin 1 C2 domains. A combination of crosslinking and FRET experiments showed that synaptotagmin 1 does not oligomerize upon Ca2+-dependent binding to phospholipid vesicles. Intriguingly, it binds to two membranes simultaneously and brings them into close proximity as visualized by cryo-EM experiments. We showed that the isolated C2B domain is sufficient to induce close membrane proximity. Mutational analysis suggested that the abundance of basic residues around the C2B surface, which generates a highly positive electrostatic potential together with the bound Ca2+ ions, is essential for this activity. We suggest that the ability of the C2B domain to bring membranes into close proximity can explain why the C2B domain has a more critical function in vivo than the C2A domain.nn"],"dc:identifier":["https://hdl.handle.net/2152.5/227","65338227"],"dc:identifier.uri":["https://hdl.handle.net/2152.5/227"],"dc:language":["en"],"dc:subject":["SNARE Proteins","Synaptotagmin I","Cytoplasm"],"dc:title":["Mechanism of Synaptotagmin Action in Neurotransmitter Release"],"dc:type":["Thesis"]},"updated_at":"2026-07-27T21:19:37Z"}