Infections of Neisseria gonorrhoeae (gonococcus) and the related Neisseria meningitidis (meningococcus) result in a high worldwide disease burden. The etiological agent of the sexually transmitted infection gonorrhoea, N. gonorrhoeae infects the human urogenital tract, resulting in inflammation and in some cases infertility. By contrast, N. meningitidis survives asymptomatically in the nasopharynx, but can invade the bloodstream, leading to septicaemia and meningitis. While these two pathogenic Neisseria are genetically very similar, there are still many notable differences. One of these is in the putative formaldehyde detoxification operon of adhC-estD, encoding for an alcohol dehydrogenase and an esterase. In the gonococcus, adhC is a pseudogene but estD still appears to be expressed, raising the question of its function. This thesis focuses on the biochemical properties of the AdhC-EstD system, its metabolic role in the cell, and how they are related to the different environmental niches occupied by this pair of pathogenic Neisseria.
Phenotypic characterisation of meningococcal mutants generated in adhC-estD confirmed their role in formaldehyde detoxification. These mutants were also affected in biofilm formation, which pointed to the possible accumulation of endogenous formaldehyde. Biochemical characterisation of EstD from N. meningitidis found it to be highly active toward its physiological substrate S-formylglutathione, further confirming its role in formaldehyde detoxification. Its crystal structure revealed molecular details causing this specificity; and the biochemical and physiological consequences of S-glutathionylation to a conserved Cys54 were also explored.
Even with a defective detoxification system, mutation of the gonococcal estD still resulted in a formaldehyde sensitive and biofilm deficient phenotype. An estD C54A mutant was also found to be deficient in biofilm formation on transformed cervical epithelial cells. With the results pointing to a secondary role for EstD, possible interactions with proteins involved in the folate dependent carbon metabolism were discovered. It was hypothesised that this system may act in a secondary
pathway for formaldehyde detoxification in N. gonorrhoeae, and this may be regulated by EstD. The potential implications of this discovery in relation to pathogenic Neisseria were also discussed.
This thesis proposes that differences between the two bacteria are the result of specific genetic adaptations caused by their colonisation of differing environments. N. meningiditis requires the glutathione dependent dedicated detoxification system due to the formaldehyde-rich environment of the bloodstream; while the non-invasive N. gonorrhoeae survives in an environment that does not require its use. A model for the adaptation of these two Neisseria species to the metabolism of the reactive species S-nitrosoglutathione is also proposed.