Please use this identifier to cite or link to this item: http://dx.doi.org/10.18419/opus-2348
|Title:||Model-based analysis of an adaptive evolution experiment with Escherichia coli in a pyruvate limited continuous culture with glycerol|
|metadata.ubs.publikation.source:||EURASIP journal on bioinformatics and systems biology (2012), 2012:14. URL http://dx.doi.org./10.1186/1687-4153-2012-14|
|Abstract:||Bacterial strains that were genetically blocked in important metabolic pathways and grown under selective conditions underwent a process of adaptive evolution: certain pathways may have been deregulated and therefore allowed for the circumvention of the given block. A block of endogenous pyruvate synthesis from glycerol was realized by a knockout of pyruvate kinase and phosphoenolpyruvate carboxylase in E. coli. The resulting mutant strain was able to grow on a medium containing glycerol and lactate, which served as an exogenous pyruvate source. Heterologous expression of a pyruvate carboxylase gene from Corynebacterium glutamicum was used for anaplerosis of the TCA cycle. Selective conditions were controlled in a continuous culture with limited lactate feed and an excess of glycerol feed. After 200–300 generations pyruvate-prototrophic mutants were isolated. The genomic analysis of an evolved strain revealed that the genotypic basis for the regained pyruvate-prototrophy was not obvious. A constraint-based model of the metabolism was employed to compute all possible detours around the given metabolic block by solving a hierarchy of linear programming problems. The regulatory network was expected to be responsible for the adaptation process. Hence, a Boolean model of the transcription factor network was connected to the metabolic model. Our model analysis only showed a marginal impact of transcriptional control on the biomass yield on substrate which is a key variable in the selection process. In our experiment, microarray analysis confirmed that transcriptional control probably played a minor role in the deregulation of the alternative pathways for the circumvention of the block.|
|Appears in Collections:||04 Fakultät Energie-, Verfahrens- und Biotechnik|
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