Browsing by Subject "Proteome"
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Item Mesoporous silica chips for harvesting the low molecular weight proteome from human serum(2009-12) Hu, Ye; Ferrari, Mauro, 1959-In this dissertation, mesoporous silica thin films with tunable features at the nanoscale were fabricated using the triblock copolymer template pathway, with the aim of specifically harvesting the low molecular weight peptides and proteins from human serum, which has been regarded as a potential source of diagnostic biomarkers for the early detection of disease. The superior properties of mesoporous silica have been demonstrated in applications which include chemical sensing, filtration, catalysis, drug-delivery and selective biomolecular uptake. These properties depend on the architectural, physical and chemical properties of the materials, which in turn are determined by the processing parameters in evaporation-induced self-assembly (EISA). Using the different polymer templates and polymer concentration in the precursor solution, various pore size distributions, pore structures and surface hydrophilicities were obtained and applied for nanotexture-selective recovery of low mass proteins. With the assistance of mass spectrometry and statistic analysis, we demonstrated the correlation between the nanophase characteristics of the mesoporous silica thin film and the specificity and efficacy of low mass proteome harvesting. In addition, to overcome the limitations of the pre-functionalization method in polymer selection, plasma ashing was used for the first time for the treatment of the mesoporous silica surface prior to chemical modification. Opposite surface charges due to the different functional groups used, resulted in a distinctive selectivity of the low molecular weight proteins from the serum sample. The mesoporous silica chips operate with extraordinary rapidity, high reproducibility, no sample pre-processing, and substantial independence from sample acquisition and storage temperature.In conclusion our study demonstrates that the ability to tune the physicochemical properties of mesoporous silica surfaces has the potential to promote the use of this material as a tool for the selective separation and concentration of the low molecular weight proteome from complex biological fluids.Item Systematic analysis of transcriptome and proteome in opportunistic bacterium Pseudomonas aeruginosa(2012-05) Kwon, Taejoon; Marcotte, Edward M.Transcription and translation are the two most important central mechanisms to control gene regulation in living organism. Although these two mechanisms have been studied intensively for last several decades, it is still not clear how all the information encoded on genomic DNA is converted to mRNA and proteins, the molecular functional components that change the characteristics of cells, depending on their needs. Here, I investigated the gene regulation of opportunistic bacterium Pseudomonas aeruginosa, using recently developed high-throughput techniques, microarray for transcriptomics and LC-MS/MS for proteomics. By analyzing transcriptome of 17 strains isolated over time from three individuals with cystic fibrosis, I identified 24 genes showing significant expression changes consistently across all strains, as evidence of parallel evolution of common traits that were beneficial in establishing chronic infection. Also, by analyzing proteome and transcriptome of two reference Pseudomonas aeruginosa strains, PAO1 and PA14, under growth condition mimicking in vivo nutrition environment in cystic fibrosis patients, I revealed that protein abundances are less correlated than mRNA abundance between them, and many proteins known as virulence factors showed different abundances only in protein level, demonstrating that post-transcriptional regulation is important to understand pathogenesis of Pseudomonas aeruginosa. To boost sensitivity both in identification and quantification in shotgun proteomics, I also created a novel integrative database search algorithm, and released freely available software package termed in MSblender. These results would be valuable information for the research community to understand the regulatory mechanism of gene expression both in transcription and translation, especially related to infectious diseases caused by pathogenic bacteria. Also, I present an integrative analysis method would be generally beneficial to extract more information from conventionally used shotgun proteomics experiments.