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oai:www.ideals.illinois.edu:2142/18580 2023-07-10 col_2142_5131 col_2142_8859 com_2142_5130 com_2142_8858 com_2142_234

Ha, Taekjip Chemla, Yann R. Ha, Taekjip Rogers, John A. Aksimentiev, Aleksei Diao, Jiajie 2011-01-21T22:50:35Z 2011-01-21T22:50:35Z 2013-01-22T11:00:19Z 2011-01-21T22:50:35Z 2010-12 This is a comprehensive study of protein-mediated membrane fusion through single-molecule fluorescence resonance energy transfer (smFRET). Membrane fusion is one of the important cellular processes by which two initially distinct lipid bilayers merge their hydrophobic cores, resulting in one interconnected structure. For example, exocytosis, fertilization of an egg by a sperm and communication between neurons are a few among many processes that rely on some form of fusion. Proteins called soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) play a central role in fusion processes which is also regulated by many accessory proteins, such as synaptotagmin, complexin and Munc18. By a new lipid mixing method at the single-vesicle level, we are able to accurately detect different stages of SNARE-mediated membrane fusion including docking, hemi and full fusion via FRET value of single donor/acceptor vesicle pair. Through this single-vesicle lipid mixing assay, we discovered the vesicle aggregation induced by C2AB/Ca2+, the dual function of complexin, and the fusion promotion role of Munc18/SNARE-core binding mode. While this new method provides the information regarding the extent of the ensemble lipid mixing, the fusion pore opening between two vesicular cavities and the interaction between proteins cannot be detected. In order to overcome these limitations, we then developed a single-vesicle content mixing method to reveal the key factor of pore expansion by detecting the FRET change of dual-labeled DNA probes encapsulated in vesicles. Through our single-vesicle content mixing assay, we found the fusion pore expansion role of yeast SNAREs as well as neuronal SNAREs plus synaptotagmin 1. Item withdrawn by Mark Zulauf (zulauf@illinois.edu) on 2010-11-08T15:45:55Z Item was in collections: University of Illinois Theses & Dissertations (ID: 1) No. of bitstreams: 2 Diao Jiajie.pdf: 4309171 bytes, checksum: 65d9782fac1d6a4b4853f1993e865311 (MD5) Diao_Jiajie.pdf: 4309171 bytes, checksum: 65d9782fac1d6a4b4853f1993e865311 (MD5) Made available in DSpace on 2011-01-21T22:50:35Z (GMT). No. of bitstreams: 2 Diao_Jiajie.pdf: 4279897 bytes, checksum: a0207d384d559375e58c63df5afff8fd (MD5) license.txt: 4059 bytes, checksum: cba7c4eaf068deaf7a7954c7b1160617 (MD5) Item marked as restricted to the 'Administrator' Group (id=1) by William Ingram (wingram2@illinois.edu) on 2011-01-21T22:53:35Z Item is restricted until 2013-01-21T22:53:34Z Item reinstated by Sarah Shreeves (sshreeve@illinois.edu) on 2013-01-22T11:00:19Z Item was in collections: University of Illinois Dissertations and Theses (ID: 204) Dissertations and Theses - Physics (ID: 445) No. of bitstreams: 3 Diao_Jiajie.pdf.txt: 147213 bytes, checksum: 40c8edce430bca169ea9b45480859994 (MD5) Diao_Jiajie.pdf: 4279897 bytes, checksum: a0207d384d559375e58c63df5afff8fd (MD5) license.txt: 4059 bytes, checksum: cba7c4eaf068deaf7a7954c7b1160617 (MD5) Item released from any restrictions by Sarah Shreeves (sshreeve@illinois.edu) on 2013-01-22T11:00:19Z http://hdl.handle.net/2142/18580 en Copyright 2010 Jiajie Diao Single molecule Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) Membrane fusion Single-molecule fluorescence resonance energy transfer study of SNARE-mediated membrane fusion Physics 1244 Physics 0240 University of Illinois at Urbana-Champaign Dissertation Ph.D. PHD:Physics -UIUC 10KS0240PHD