Nucleotide signalling pathways play an important role in both bacterial defence systems (1) and mammalian innate immunity pathways (2), and are therefore potential targets for the development of new biotechnology tools and therapeutics. The aim of this thesis is to increase the understanding of the biology of a novel class of bacterial enzymes that use nicotinamide adenine dinucleotide (NAD+) as a substrate (3,4) known as Toll-Interleukin Receptors (TIR) domains. Structural and biochemical approaches were used to gain insight into the function of bacterial TIR domain containing enzymes. Several TIR domain containing proteins from animals, plants and bacteria with important immune functions have now been demonstrated to utilise glycohydrolase activity as part of their signalling mechanism cleaving NAD+ into nicotinamide (Nam) and the nucleotides Adenosine diphosphate ribose (ADPR), cyclic ADPR (cADPR), and two recently described cADPR isomers called 2’cADPR and 3’cADDPR (5–11). Structural studies provided in this thesis allowed for comparison of a new TIR domain structure from the bacterium Leptolyngbya sp. (LeTIR) with available bacterial and non-bacterial TIR domains. Nuclear Magnetic Resonance (NMR) studies conducted as part of this research provide a better understanding on how is the NADase function of LeTIR and a second bacterial TIR domain from Lacunisphaera limnophila (LiTIR). […] This thesis provides new structural and biochemical information on bacterial TIR domains which potentially will help explain how they can produce multiple nucleotide products and provide insight into the structural determinants for their production. I produced a crystal structure in ligand-free state of LeTIR, one bacterial TIR domain that has not been solved until now. In addition, biochemical characterisation with NMR based NADase assays were performed with two bacterial TIR domain-containing proteins (LeTIR and LiTIR).