Neisseria gonorrhoeae (Ng) is the causative agent of the second most common STI, gonorrhoea. In Australia, the notification rates of gonorrhoea have increased by 80% in the past decade. Trigger Factor (TF) is a 48 kDA protein with three domains, the N-terminal ribosome binding domain, the central PPIase domain and the C-terminal chaperone domain. Being part of the immunophilin superfamily, these proteins have PPIase activity, meaning they catalyse the cis-trans isomerisation of proline residues, which is a rate-limiting step in protein folding. E. coli TF is associated with the bacterial ribosome and cups nascent polypeptides as they exit the ribosome. In E. coli, removal of TF in a DnaK mutant background results in lethality that is not seen in a TF or DnaK mutant background. In Listeria monocytogenes, deletion of TF resulted in reduced tolerance to heat and ethanol stress and attenuated survival in mice models.
To determine if gonococcal TF (NgTF) is an active PPIase, NgTF was recombinantly expressed and purified and enzymatic activity was confirmed using a PPIase assay. To investigate the role of TF in gonococcal stress response and homeostasis the TF mutant strain NgΔtig was created in gonococcal strain FA19, through a replacement of the TF gene (tig) with a kanamycin resistance cassette. A complement strain NgΔtig:tig was also created through the addition of an ectopic copy of tig and an erythromycin resistance cassette into a housekeeping region on an alternate locus on the mutant genome. Growth of these strains at 37°C and 30°C was not affected by deletion of tig. Deletion of tig also did not affect the ability of these strains to survive in murine macrophages. To test if TF is responsible for various stress responses, susceptibility of the mutant and complement strains to hydrogen peroxide was assessed by exposure to 5mM hydrogen peroxide. Interestingly deletion of TF increased gonococcal resistance to 5mM hydrogen peroxide during exponential growth. Further work will aim to explore the association between TF and other stress factors such as heat, pH, and antimicrobial susceptibility to determine if TF plays a role in gonococcal stress response.