• Anette Daniela Volfa Vestergaard
4. term, Sustainable Biotechnology, Master (Master Programme)
Infections are often initiated though complex protein-protein interactions (PPIs) between pathogen and host. Antibiotics previously used to treat infectious diseases are unable to co-evolve at the rate in which bacteria become multi-drug resistant. Bacteriophage therapy has become an emerging approach in the pursuit of alternative treatments. Researching PPIs between bacteriophages and bacteria could reveal novel drug targets and support the development of bacteriophage-directed treatment. Although promising, the research of the viral mechanisms in both bacteria and in phages is only in its infancy. To close this gap, this paper looks into the PPIs between the stringent stress response in Escherichia coli (E. coli), specifically proteins relA and spoT, with the T7 bacteriophage proteins 1.5, 4.2 and 6.5. The aim of the current research was to identify which relA and spoT domains interacted with the phage proteins in vivo, and investigate what effect these phage proteins had on the catalytic activity of relA and spoT. The domain interactions were determined through bacterial two-hybrid assay (B2H) testing. After 48 hours, the assays indicated that the strongest interaction were found between phage proteins 4.2 and 6.5, and the spoT hydrolysis-synthesis (HS) and ThrRS, GTPase and SpoT (TGS) domains. For relA, the strongest interactions were observed between 4.2 and 6.5 and the relA domains TGS, CC- and ACT. Only weak interactions arose between T7 protein 1.5 and the spoT and relA truncations. In effort to understand how the phage proteins influenced the enzymatic activity of relA and spoT, the plan was to purify the E. coli and T7 phage proteins and run them through a biochemical assay using radioactive pppGpp and GTP. Due to time constraints (three months), that was not fully realized. The results found in the B2H could be understood as preliminary indicators of the interactions between the phage proteins 4.2 and 6.5 with relA and spoT. Furthermore, it is speculated that 4.2 and 6.5 inhibit the host defensive mechanisms by targeting the proteins involved in the stringent stress response. Further testing is needed confirm these preliminary results.
Publication date7 Jun 2022
Number of pages70
External collaboratorYong Zhang
Yong Zhang yong.zhang@bio.ku.dk
ID: 472264984