Developing novel lipid architectures as a platform for membrane biophysics
Abstract/Contents
- Abstract
- Many cellular processes including cell-cell communications and regulated membrane transport are mediated by membrane proteins and depend upon the ability of lipid membranes to be a differentially permeable barrier. However, the roles and function of membrane proteins are often difficult to study due to the complexity of the native membranes and lack of reliable and flexible artificial model lipid membranes. Supported lipid bilayers (SLB) have been used as a model system to study biological membrane behavior and the structure and function of membrane proteins and receptors in a simpler context apart from the complex cellular environment. Although SLBs have the advantages of simple formation, easy handling and are well-suited for investigation by a suite of surface sensitive methods due to their planar geometry, the close proximity of the lower leaflet to the solid support often leads to unfavorable interactions with integral membrane proteins. This causes distortion of the protein conformation and possible loss of its reactivity and function. Moreover, this interaction with the substrate often traps proteins and reduces their mobility in the membranes. Recognizing this limitation, we have developed a new model membrane architecture in which the DNA-tethered lipid bilayer is either to fixed DNA on a surface or to laterally mobile DNA displayed on a supported bilayer. This separates the lipid membranes from surface interactions and provides a more favorable environment for integral membrane protein with large globular domains. With mobile DNA hybrid tethers, stable tethered bilayers were made with specific lipid composition, while those with fixed tethers are stable regardless of membrane composition. The mobile tethers between a tethered and a supported lipid bilayer offer a particularly interesting architecture for studying the dynamics of membrane-membrane interactions. By careful choice of composition, improved stability was obtained and we can investigate the lateral segregation of DNA hybrids when different lengths are present. Based on a theoretical model, the effects of population, length and affinity of DNA complexes are simulated and described. This model system captures some of the essential physics of synapse formation and is a step toward understanding lipid membrane behavior in a cell-to-cell junction. To demonstrate the excellent environment provided by DNA-tethered membranes for studying transmembrane proteins free from any surface interactions, the behavior of a transmembrane protein, the photosynthetic reaction center, reconstituted in the DNA-tethered membranes is investigated. Inspired by DNA-mediated membrane fusion studies of our group, we applied the DNA-machinery to achieve fusion of small (~ 100 nm) proteoliposomes for delivery of membrane proteins to either giant vesicles or DNA-tethered planar lipid membrane patches. The diffusion behavior of delivered proteins is measured and compared with those in supported bilayers. Also, the protein activity and orientation before and after fusion is analyzed. This will offer a feasible method to incorporate intact membrane proteins to already formed model membranes. In addition, the behavior of proteins during the fusion event will provide insight into the mechanism of DNA-mediated lipid membrane fusion. The geometry of our model membrane system directly mimics that of a neuronal synapse. We expect that this architecture will be readily transferable to other model membrane fusion systems, including systems using reconstituted SNARE proteins. Consequently, it will be of considerable interest to a wide range of researchers.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2011 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Chung, Minsub |
|---|---|
| Associated with | Stanford University, Department of Chemical Engineering |
| Primary advisor | Boxer, Steven G. (Steven George), 1947- |
| Primary advisor | Fuller, Gerald G |
| Thesis advisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Fuller, Gerald G |
| Thesis advisor | Swartz, James R |
| Advisor | Swartz, James R |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Minsub Chung. |
|---|---|
| Note | Submitted to the Department of Chemical Engineering. |
| Thesis | Ph.D. Stanford University 2011 |
| Location | electronic resource |
Access conditions
- Copyright
- © 2011 by Minsub Chung
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