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Title: Studying the adaptation of Escherichia coli K-12 in response to trimethylamine-N-oxide
Author: Denby, Katie Jane
ISNI:       0000 0004 5993 2582
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2016
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Escherichia coli is a Gram-negative, metabolically versatile facultative anaerobe, which utilises either fermentation, anaerobic respiration or aerobic respiration for energy generation and growth. Trimethylamine-N-oxide (TMAO) is used by E. coli as an alternative terminal electron acceptor, being reduced to trimethylamine (TMA). Although the regulation and operation of the E. coli TMAO respiratory chain (TorCAD) are well characterised, there is no understanding of the dynamic adaptive processes that occur in E. coli during transition from fermentative to TMAO-respiratory growth. Here, glucose-limited chemostat cultures were used to study these adaptive processes. Analyses of the transcriptional and metabolic changes occurring when E. coli K-12 responds to TMAO revealed a number of unexpected components of the adaptive process. Firstly, it was found that growth on a sub-optimal concentration of TMAO resulted in mixed metabolism, with two distinct sub-populations of cells that either activate or do not activate the transcription of the torCAD operon in response to TMAO. DNA methylation was found to contribute to this regulation in response to low concentrations of TMAO. Secondly, it was found that E. coli possesses TMAO demethylase activity, as both the products of this activity, dimethylamine and formaldehyde and the induction of the frmRAB operon was detected when cells were grown in the presence of TMAO. Work was carried out to characterise the regulator protein, FrmR, of the frmRAB operon. Here, it is shown that the FrmR transcriptional repressor specifically reacts with formaldehyde, resulting in the formation of two inter-molecular methylene bridges between the N-terminal Pro2 and Cys35 residues of adjacent FrmR subunits, leading to alleviation of repression. It was also shown that growth yield was higher in the presence of TMAO under aerobic conditions eventhough TMAO reduction to TMA was not detected. Futhermore, the nitrate response regulator NarL does not directly regulate the torCAD operon.
Supervisor: Jeffrey, Green Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available