Investigation of Neuronal Membrane Fusion Using Fluorescence Correlation Spectroscopy

Vennekate, Wensi

dc.contributor.advisor	Walla, Peter Jomo Prof. Dr.	de
dc.contributor.author	Vennekate, Wensi	de
dc.date.accessioned	2013-01-20T13:16:49Z	de
dc.date.available	2013-01-30T23:50:57Z	de
dc.date.issued	2013-01-15	de
dc.identifier.uri	http://hdl.handle.net/11858/00-1735-0000-000D-F0EB-E	de
dc.identifier.uri	http://dx.doi.org/10.53846/goediss-3360
dc.description.abstract	Die Neurotransmission erfordert die schnelle Fusion zwischen den synaptischen Vesikeln und der presynaptischen Membran. Die Membranfusion wird durch den Calciumeinstrom ausgelöst und benötigt die SNARE-Proteine. Das Vesikelprotein, Synaptotagmin-1, dient als ein Calciumsensor, und die Bindung des Synaptotagmin-1 an die Plasmamembran ist ein essentieller Schritt für die Membranfusion. In dieser Arbeit wurden einige Details der Membranfusion untersucht und diskutiert. Als Hauptergebnis wird die cis- und trans-Membraninteraktion von Synaptotagmin-1 präsentiert.	de
dc.format.mimetype	application/pdf	de
dc.language.iso	eng	de
dc.rights.uri	http://creativecommons.org/licenses/by-nc-nd/3.0/	de
dc.title	Investigation of Neuronal Membrane Fusion Using Fluorescence Correlation Spectroscopy	de
dc.type	doctoralThesis	de
dc.title.translated	Untersuchung der neuronalen Membranfusion mit der Fluoreszenz Korrelations Spektroskopie	de
dc.contributor.referee	Walla, Peter Jomo Prof. Dr.	de
dc.date.examination	2012-11-08	de
dc.subject.dnb	540 Chemie	de
dc.subject.gok	Molekulare Biophysik. Biophysikalische Chemie (PPN619462841)	de
dc.description.abstracteng	Neurotransmission requires rapid fusion of synaptic vesicles with the presynaptic membrane. This fusion is triggered by calcium influx and promoted by SNARE assembly. The vesicular protein synaptotagmin-1 acts as a calcium sensor and its trans-tethering to the plasma membrane is an essential step in the membrane fusion process. In this study, several details in membrane fusion have been investigated and discussed. First, the molecular requirements of the trans- and cis-membrane interaction of synaptotagmin-1 have been analyzed. All of the three binding sites of synaptotagmin-1 (the C2A domain, the C2B domain and the polylysine patch) are involved in both of these processes. Cis-binding to the host membrane containing 20% phosphatidylserine seems to be a stable state and can be overcome by neither trans-binding between synaptotagmin-1 and the target membrane containing acidic phospholipids, nor by the interaction between synaptotagmin-1 and SNAREs inserted into the acceptor membrane. Similarly, with 12% phosphatidylserine in the host membrane-a situation comparable to that in wild-type vesicles-cis-binding is found nearly exclusively. With 5% phosphatidylserine, however, the presence of either phosphatidylinositol-4,5-bisphosphate in the target membrane or calcium in the solution can invoke conversion to trans-binding with great efficiency. Second, liposome cross-linking can be mediated by soluble C2AB fragments, when all the binding sites for C2AB are saturated on the liposome surface. To rationalize this result, it is assumed that binding with only the C2A- or the C2B domain might occur under these conditions. The other-unbound-domain of the same synaptotagmin molecule will eventually lead to cross-linking, if it attaches to a different liposome. This finding confirms the observation of cis-binding and explains some of the conflicts in the existing literature. Apparently, a subtle balance between cis- and trans-binding of synaptotagmin-1 may play an important role in the regulation of neuronal membrane fusion. Moreover, first data for the characterization of mouse synaptic vesicles, in terms of average protein mass per vesicle, was measured in this study. The results obtained for mouse synaptic vesicles are similar to those of rat synaptic vesicles and offer a single vesicle basis for further investigating mouse synaptic vesicles as well as membrane fusion in a quantitative fashion. Finally, details of the inhibiting effect of α-SNAP on the fusion of chromaffin granules with larger liposomes were also investigated in the present study. The experimental findings suggest that α-SNAP inhibits fusion of chromaffin granules and stops the SNARE assembly at the C-terminal site, so that partial SNARE zippering can still mediate docking of chromaffin granules.	de
dc.contributor.coReferee	Reinhard, Jahn Prof. Dr.	de
dc.contributor.thirdReferee	Steinem, Claudia Prof. Dr.	de
dc.subject.topic	Chemistry	de
dc.subject.ger	Synaptotagmin 1	de
dc.subject.ger	trans-bindung	de
dc.subject.ger	cis-bindung	de
dc.subject.ger	synaptische Vesikel	de
dc.subject.ger	Fluoreszenz Korrelations Sprektroskopie	de
dc.subject.ger	-SNAP	de
dc.subject.ger	Docking	de
dc.subject.ger	Liposome	de
dc.subject.ger	Rückbindung	de
dc.subject.ger	Calcium	de
dc.subject.eng	Synaptotagmin 1	de
dc.subject.eng	trans-binding	de
dc.subject.eng	cis-binding	de
dc.subject.eng	synaptic vesicles	de
dc.subject.eng	Fluorescence correlation spectroscopy	de
dc.subject.eng	-SNAP	de
dc.subject.eng	tethering	de
dc.subject.eng	liposome	de
dc.subject.eng	back binding	de
dc.subject.eng	calcium	de
dc.subject.bk	35 .00 Chemie	de
dc.identifier.urn	urn:nbn:de:gbv:7-webdoc-3882-8	de
dc.identifier.purl	webdoc-3882	de
dc.affiliation.institute	Fakultät für Chemie	de
dc.identifier.ppn	737345934	de

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