Ikeda, Naomi (2014) Molecular mechanisms of coccoid form formation in 'Campylobacter jejuni'. (PhD thesis), Kingston University, .
Abstract
It is known that under stress conditions Campylobacter jejuni can change morphology from a characteristic spiral to coccoid form (CF). Genetic and biochemical mechanisms of CF formation (CFF) in C. jejuni remain unclear, however cell shape maintenance in other microorganisms is known to be genetically determined. The focus of this study is the fiinctional analysis of spoT and amiA genes in C. jejuni. Inactivation of the spoT gene stimulated CFF in strain 11168H and 81116, but the direct role of this gene in CFF is unclear due to the pleiotropic effects of spoT mutation. The amiA gene, encoding a putative peptidoglycan amidase, is involved in CFF in the closely related bacterium H. pylori. Comparative genomics studies revealed conservation of the amiA gene clusters among diverse bacteria. Amino acid sequence analysis of AmiA orthologs suggests a possible dual function of these proteins due to high variability in their N-terminal domains, as opposed to high conservation of the C-terminal domains responsible for amidase activity. The inactivation of this gene in H. pylori affected the accumulation of a dipeptide motif peptidoglycan unit suggesting additional (peptidase) activity of this enzyme. In contrast to H. pylori, attempts to inactivate amiA in C. jejuni strains 11168H and 81116 proved unsuccessful suggesting the gene is essential for bacterial viability. Over expression of C. jejuni AmiA in E. coli resulted in decreased viability due to loss of membrane integrity as revealed by BacLight LIVE/DEAD staining. C. jejuni AmiA overexpression also lead to a change in the E. coli muropeptide profile compared to native levels. Results suggest that, similar to AmiA proteins in other bacteria, AmiA of C. jejuni may also be involved in peptidoglycan maintenance and in CFF.
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