This dissertation focuses on the development of biological platforms on which
the function and characterization of transmembrane proteins can be performed
simultaneously utilizing a biomembrane mimic consisting of a solid supported
phospholipid bilayer (SLB). The study centered on the platform development,
biophysical measurements of transmembrane proteins and membrane species
chromatography. Membrane proteins play an essential role in various cellular and
physiological processes. Their normal functions are essential to our health, and many
impaired proteins have been related to serious diseases. Gaining a better understanding
of membrane proteins is an essential step towards the development of more specific and
competent drugs.
This research study is divided into two main parts. The first part centered on the
creation of a new platform for allowing transmembrane proteins to freely move inside
supported lipid bilayers with the same mobility that can be found in vesicle systems.
SLBs have been extensively used as model systems to study cell membrane processes because they maintain the same two-dimensional fluidity of lipids within the membrane
found in live cells. However, one of the most significant limitations of this platform is its
inability to incorporate mobile transmembrane species. Our strategy involves supporting
the lipid bilayer on a double cushion, where we not only create a large space to
accommodate the transmembrane portion of the protein, but also passivate the
underlying substrate to reduce non-physiological protein-substrate interactions. High
diffusion constants and high mobile fractions were obtained for a transmembrane protein
reconstituted within this double cushion system.
The second area of this study focuses on the creation of a new method to rapidly
separate membrane components using electrophoresis in SLBs. This work showed that
even subtly different chemical isomers can be well-separated by a simple electrophoretic
technique when cholesterol is present in the separation matrix. As a first step towards the
purification of proteins, this work showed that streptavidin proteins doubly bound to a
bilayer by a biotinylated lipid can be separated from streptavidin proteins which are
singly bounded.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/ETD-TAMU-2652 |
| Date | 15 May 2009 |
| Creators | Diaz Vazquez, Arnaldo Joel |
| Contributors | Cremer, Paul S., Hu, James C. |
| Source Sets | Texas A and M University |
| Language | en_US |
| Detected Language | English |
| Type | Book, Thesis, Electronic Dissertation, text |
| Format | electronic, application/pdf, born digital |
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