The bacterium Campylobacter jejuni is the most common cause of food borne disease in the UK, causing a 5-7 day enteritis including profuse watery diarrhoea, abdominal pain, fever, headache and occasionally vomiting. In rare cases leading to the paralysing autoimmune disease, Guillain-Barré syndrome. C. jejuni are highly motile cells, propelled through the environment by flagella, their motility is directed through a behaviour called chemotaxis. Cells are able to detect attractants or repellents and reposition the cell accordingly. Chemotaxis is central to C. jejuni colonisation as non-motile and non-chemotactic mutant strains poorly colonise their usual hosts. In Escherichia coli chemotaxis is regulated by the Che proteins which form a two component phospho relay system. In previous studies In silico comparison of E. coli Che proteins identified homologues in C. jejuni, which display altered chemotactic phenotypes in Δche mutant strains. Studies of interactions between the Che proteins using bacterial and yeast two hybrid systems, suggested ways in which the homologues may interact, but to further discern these mechanisms required in vitro study. For the purpose of this study the C. jejuni Che homologues were cloned, expressed and purified, for use in in vitro experiments. Radiolabelled Phosphotransfer assays confirmed CheA as a histidine kinase, and demonstrated Pi transfer to the response regulators of CheY, CheV and CheA, in that order of preference. Affinity tag pull-down assays found the predicted decrease in affinity between phosphorylated CheY and CheA, but also an increase in the affinity of phosphorylated CheV for the receptor, TLP[subscript 1]. The results of this study confirm the two component backbone of the C. jejuni Che model, and suggest how CheV may regulate methylation adaption in a system devoid of a CheB response regulator.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:593779 |
| Date | January 2014 |
| Creators | Ainsworth, Paul |
| Contributors | Ketley, Julian |
| Publisher | University of Leicester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/2381/28667 |
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