In plant innate immunity, recognition of pathogen effectors generally occurs through the family of nucleotide-binding leucine-rich repeat receptors (NLRs). NLRs can be characterised into different groups based on their N-terminal domain, the Toll/interleukin-1 receptor (TIR), coiled coil (CC), and resistance to powdery mildew 8 (RPW8) domain. For TIR-NLRs (TNLs) and CC-NLRs (CNLs), either direct or indirect detection utilises the leucine rich repeats (LRR) domain. Plant genomes also encode multiple TN proteins that are naturally truncated TNLs lacking the LRR domain and are implicated in innate immunity. Recent studies have also shown that the TIR domains of TNLs or TN/TIR-only proteins possess NADase activity.
My project aims to investigate the TN gene cluster (TN4-12) from Arabidopsis thaliana though biophysical, functional and structural characterisation of these TN proteins as well as their TIR domains.
Currently, small scale expression using Escherichia coli Rosetta2 strain and purification of N-6HIS TN 7, 8, 11 constructs have shown that these protein were insoluble or poorly expressed. In order to improve protein expression, the cloning of different fusion partners to improve protein folding and solubility is currently ongoing. TN11TIR was successfully purified and TN8TIR purification is currently being optimised.