Degree Type

Dissertation

Date of Award

2008

Degree Name

Doctor of Philosophy

Department

Biochemistry, Biophysics and Molecular Biology

First Advisor

Yeon-kyun Shin

Abstract

Membrane fusion is a fundamental life process to transfer cargos between distinct biological compartments. It was widely accepted that the SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) proteins play a central role in driving membrane fusion in eukaryotes. The assembly of SNARE complex brings the opposing membrane together to form a lipidic `stalk', a hemifusion intermediate where the outer leaflets from both membrane was fused and the inner leaflets maintain intact. A zippering model predicted that the formation of the SNARE complex started from the N-terminal and zipped forward to the membrane proximal C-terminal of the SNARE motif. The energy released during the assembly of the complete complex will induce the membrane fusion.

Combining EPR (Electron Paramagnetic Resonance) and FRET (Fluorescence Resonance Energy Transfer) techniques, we investigated the structures and functions of the SNARE proteins from yeast. We found a partially assembled SNARE complex intermediate, in which the C-terminal segment of Sso1pHT SNARE motif and the whole SN2 of Sec9 were flexible and uncomplexed. Furthermore, the studies of mutants in SN2 implied that the SN2 of Sec9 might not be related to the docking of the vesicles, but it plays an important role in the membrane fusion. Unexpectedly, the energy difference between the partially assembled complex and the complete four-helix bundle is close to zero. We suggested that the assembly of the SNARE motif might change the curvature of the lipid membrane and decrease the energy barrier of the membrane fusion.

The membrane fusion was controlled exquisitely by a network of regulators. Up to now, more than twenty regulatory proteins were identified. Some of them, such as synaptotagmin, Munc-18 and complexin, were extensively studied. However, the results from different laboratories are contradicted. In the course of my studies, I tried to investigate these proteins using EPR and single molecular techniques.

DOI

https://doi.org/10.31274/etd-180810-1982

Copyright Owner

Zengliu Su

Language

en

Date Available

2012-04-30

File Format

application/pdf

File Size

121 pages

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