Mycobacterium tuberculosis is the biggest infectious disease killer worldwide, with 1.6 million deaths per annum and increasing rates of antimicrobial-resistant infections. Strategies to manage tuberculosis require fundamental research to drive innovation in therapeutics, but the World Health Organisation has highlighted the scarcity of new approaches. An opportunity exists to address these points by harnessing “toxin-antitoxin” (TA) systems in bacteria that target essential steps such as translation, DNA replication and cell wall synthesis. The abundance of TA systems throughout bacteria, and M. tuberculosis in particular, suggests these systems represent a rich source of new biochemistry and antibacterial targets.
TA systems encode two components, a toxic protein that targets an essential cellular process, and an antagonistic antitoxin, which blocks toxin activity when cells are growing under favourable conditions. Although the processes that lead to toxin activation remain under debate, it has been proposed that under certain stress conditions, increased toxin transcription and synthesis may lead to activation. This in turn reduces growth rate, which can provide a means to survive with minimal metabolic burden until favourable conditions return. TA systems are remarkably abundant in M. tuberculosis, which encodes more than 80 putative systems that are thought to contribute to the success of M. tuberculosis as a human pathogen. Many of the putative M. tuberculosis toxins tested thus far were shown to inhibit bacterial growth, and the highly toxic nature of some toxins suggests that their antibacterial mechanisms could be developed into antimicrobials.
Research Plan. In our recent work, we established and characterised the MenAT TA family of nucleotidyltransferases, which were proven to be active in killing tuberculosis (Cai et al. (2020), Beck et al. (2020)). Toxin MenT3 targets and modifies all four mycobacterial serine tRNAs. The MenAT family also represents a new class of type VII TA systems, regulated through phosphorylation by the cognate MenA antitoxin.
These discoveries prompt two new routes for controlling the growth of M. tuberculosis; (i) modulating toxin activity and (ii) inhibiting amino-acyl charging of serine tRNAs. This project aims to explore these opportunities through a combination of biochemistry, structural biology and in silico methods encompassing protein dynamics modelling, docking studies and structure-based drug design.
Objective 1 – Toxin inhibition. Use our high resolution structures of toxin MenT3 and MenT4 with VirtualFlow, to screen and identify potential small molecule inhibitors. Hits will be screened for toxin inhibition through established biochemical assays, microbiology and X-ray crystallographic studies.
Objective 2 – Dysregulation of toxin-antitoxin interactions. Structural and biophysical studies will be performed to build models for the control of MenT toxins through phosphorylation by cognate MenA antitoxins.
Objective 3 – Target toxin targets. MenT3 modifies serine tRNAs, indicating the seryl-tRNA synthetase as a potential antibiotic target. Structural studies of the seryl-tRNA synthetase from M. tuberculosis will be combined with in silico screening methods to identify and characterise potential inhibitors.
Training. This project is part of the EPSRC Molecular Sciences for Medicine (MoSMED) Centre for Doctoral Training. Led by Dr Tim Blower (Durham), the project will provide training in molecular microbiology, protein biochemistry and structural biology. Second supervisor Dr Agnieszka Bronowska (Newcastle) will provide training in in silico methods including ligand screening and structure-based drug design. At later stages, Dr Danny Cole (Newcastle) will provide further training to employ hit-to-lead optimisation workflows. This multidisciplinary training will be further supported by external courses (ie. Diamond Light Source), producing a scientist with the broad range of in-demand skills that are needed to tackle ongoing societal problems such as antimicrobial resistance and discovery of novel drug hits.
To apply for this project please visit the Durham University application portal to be found at: https://www.dur.ac.uk/study/pg/apply/ Please select the course code F1A201 for a PhD in Molecular Sciences for Medicine and indicate the project reference or the title of the project in the ‘Field of Study’ section of the application form. Please note that there is no need to submit a Research Proposal with your application however we do require a Cover Letter, CV, an academic transcript, the contact details of two referees and proof of English language proficiency if appropriate. Should you have any queries regarding the application process at Durham University please contact the Durham MoSMed CDT Manager, Emma Worden at: [Email Address Removed].
For more details of how to apply and other available opportunities, please visit the MoSMed website: PhD Studentships | Molecular Sciences for Medicine | Newcastle University (ncl.ac.uk)
Please see https://research.ncl.ac.uk/mosmed/ and https://www.blowerlab.com/ for more information, or contact Dr Tim Blower [Email Address Removed] for informal enquires.
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