Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.582246
Title: The use of recently developed mass spectrometry-based proteomic approaches for the study of Methylocella silvestris BL2
Author: Patel, Nisha A.
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 2012
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Abstract:
The study of the protein complement, termed proteomics, has advanced over the last twenty years as a consequence of developments in mass spectrometry. Currently, improvements in mass spectrometry-based approaches are targeted towards achieving information on both the identity and abundance of proteins. Increased numbers of protein identifications are obtained by simplifying the analyte of interest. This can be achieved with the use of separation techniques, including two-dimensional liquid chromatography (2D-LC). Ion mobility coupled to mass spectrometry has recently been shown to be a useful post-ionisation separation tool for proteomic studies. The utility of these technologies for obtaining both qualitative and quantitative information is not extensively addressed in the current literature. The use of a recently developed 2D-LC system, together with a method of ion mobility separation and a label-free quantitative approach for proteomic studies has been evaluated here for characterising the proteome of the bacterium Methylocella silvestris. This bacterium is the first methane-utilising bacteria also discovered to grow on substrates containing carbon-carbon bonds, and has great biotechnological potential. The metabolism of this bacterium was studied by obtaining information on its soluble proteome when grown with methane, propane, succinate, acetate, methanol, methylamine or trimethylamine. The benefits and limitations of 2D-LC and ion mobility for profiling and labelfree quantitative studies were demonstrated for simple mixtures and complex bacterial extracts. The combination of both 2D-LC and ion mobility was also achieved, resulting in wider proteome coverage when compared to the respective stand-alone approaches. A cluster of expressed genes that were greatly up-regulated under trimethylamine growth and monomethylamine growth were proposed to be involved in the indirect pathway for trimethylamine metabolism. It was further verified that one of these genes expresses the previously unidentified trimethylamine monooxygenase. A propane assimilation route was proposed, based on information obtained on the levels of primary oxidation enzymes and downstream central metabolic pathways.
Supervisor: Not available Sponsor: Engineering and Physical Sciences Research Council (EPSRC) ; Royal Society of Chemistry (Great Britain) (RSC)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.582246  DOI: Not available
Keywords: QD Chemistry ; QP Physiology ; QR Microbiology
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