Glycosylation of Campylobacter iron transport systems and the role of host stress hormones
thesisposted on 30.05.2013, 13:02 by Susan Louise Hardy
Campylobacter jejuni is the main cause of gastroenteritis in developed countries and the exact nature of the organism’s virulence continues to be the subject of much research. Initially the main area of focus for this project was to establish whether the glycosylation of iron uptake proteins is necessary for their function. We know that protein glycosylation is essential for interaction with host cells and that bacteria also require functioning iron uptake systems in order to colonise their host. Mutants in pglB, pglI and pglK were utilised in growth assays and compared with mutants in all known iron uptake systems. The haem uptake system was investigated and a haem biosynthesis gene hemE targeted, which will be used to establish whether exogenous haem can be used for metabolism. The C. jejuni haem oxygenase ChuZ has been expressed, purified and its viability assessed by absorbance spectrophotometry, with a view to solving the structure through X-ray crystallography. With knowledge of the structure it is anticipated that it will be possible to clarify the exact function of the protein. The catechol noradrenaline has been shown to enhance growth in iron-restricted conditions through the receptor CfrA. This study has investigated the effects of glycosylation on iron uptake via CfrA. The gluconeogenic enzyme GAPDH has been shown to localise to the outer membrane where it interacts with host transferrins. Whilst the effects of glycosylation on the uptake of iron from lactoferrin have been investigated, work has also begun on constructing a gapA mutant which will be used to investigate the interaction with lactoferrin and transferrin. This study has revealed that glycosylation of iron uptake proteins was not necessary for their function. Levels of natural competence were also studied and it was found that mutants in pglB and pglK were compromised with respect to their ability to naturally transform.