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| Title: | METHODS DEVELOPMENT IN BIOLOGICAL MASS SPECTROMETRY: APPLICATIONS IN SMALL MOLECULE RESEARCH AND PROTEOMICS |
| Authors: | Williams, Taufika Islam |
| Keywords: | Mass Spectrometry
Alzheimers Disease
Aldehydes
Bicine-dSDSPAGE
Membrane Proteins |
| Date Created: | 2005 |
| Publisher: | University of Kentucky |
| Abstract: | Technological developments have enabled mass spectrometry (MS) to evolve as
one of the most versatile, sensitive and widely used analytical methods. Key areas of
research in biological MS include the development of analyte-selective MS
methodologies, along with the design of MS compatible separation technology. Analytes
of interest range from small, biologically active molecules in disease progression
research, to macromolecules such as proteins, in proteomics investigations. Advances in
these areas are vital to maintaining the level of sophistication that has become the
benchmark for MS analyses.
Mass spectrometry has found a permanent station in disease progression studies,
particularly in biomarker discovery. This is especially true for Alzheimer’s disease (AD),
a condition marked by widespread lipid peroxidation (LPO) in the brain. The main
hypothesis of the first part of this dissertation is that LPO produces aldehydes that can
potentially be exploited as AD biomarkers. Design of novel LC-MS/MS methods for
brain aldehyde analysis is described. The methods were applied towards aldehyde
quantification in the hippocampus, superior and middle temporal gyrus and cerebellum of
subjects with early AD (EAD), mild cognitive impairment (MCI) and age-matched
controls. Results obtained indicated elevation of neurotoxic aldehydes in MCI and EAD
brain and suggested that LPO occurred early in AD. Understanding AD progression has
become important for developing diagnostic methods and treatments.
Mass spectrometry is also the major analytical tool in proteomics, where gel
electrophoresis is dominant in pre-MS separations. The main hypothesis of the latter part
of this dissertation is that exposure of microbe fermenters including Clostridium
thermocellum to an external stimulus, such as ethanol, can alter the membrane proteome.
Design of novel doubled-SDS-PAGE (dSDS-PAGE) methods for membrane protein
analysis is described, as these proteins are under-represented in standard 2D-PAGE. The
newly developed Bicine-dSDS-PAGE offered superior separation over other methods and
was applied towards analysis of wild type and ethanol-adapted C. thermocellum cell
membranes. Significant differences in protein expression were observed. An
understanding of ethanol adaptation will promote the design of more ethanol-tolerant
strains. Such an outcome can have dramatic effects in the fuel industry as the trend
towards more efficient fuel development gathers momentum. |
| URI: | http://hdl.handle.net/10225/164 |
| Appears in Collections: | Electronic Theses and Dissertations
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| tiwdis.pdf | | 1340Kb | Adobe PDF | View/Open |
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