The structure of NAD+ consuming protein Acinetobacter baumannii TIR domain shows unique kinetics and conformations.

Klontz, E.; Obi, J. O.; Wang, Y.; Glendening, G.; Carr, J.; Tsibouris, C.; Buddula, S.; Nallar, S.; Soares, A.; Beckett, D.; Redzic, J. S.; Eisenmesser, E.; Palm, C.; Schmidt, K.; Scudder, A.; Obiorah, T.; Essuman, K.; Milbrandt, J.; Diantonio, A.; Ray, K.; Snyder, M. L.; Deredge, D.; Snyder, G. A.

doi:10.1101/2023.05.19.541320

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bioRxiv · 10.1101/2023.05.19.541320

The structure of NAD+ consuming protein Acinetobacter baumannii TIR domain shows unique kinetics and conformations.

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Abstract

Toll-like and Interleukin-1/18 receptor resistance (TIR) domain-containing proteins function as important signaling and immune regulatory molecules. TIR domain-containing proteins identified in eukaryotic and prokaryotic species also exhibit NAD+ hydrolase activity in select bacteria, plants, and mammalian cells. We report the crystal structure of the Acinetobacter baumannii TIR domain protein (AbTir-TIR) with confirmed NAD+ hydrolysis and map the conformational effects of its interaction with NAD+ using HDX-MS. NAD+ results in mild decreases in deuterium uptake at the dimeric interface. In addition, AbTir-TIR exhibits EX1 kinetics indicative of large cooperative conformational changes which are slowed down upon substrate binding. Additionally, we have developed label-free imaging using 2pFLIM which shows differences in bacteria expressing native and mutant NAD+ hydrolase-inactivated AbTir-TIREA protein. Our observations are consistent with substrate-induced conformational changes reported in other TIR model systems with NAD+ hydrolase activity. These studies provide further insight into bacterial TIR protein mechanisms and their varying roles in biology.

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Klontz, E., Obi, J. O., Wang, Y., Glendening, G., Carr, J., Tsibouris, C., Buddula, S., Nallar, S., Soares, A., Beckett, D., Redzic, J. S., Eisenmesser, E., Palm, C., Schmidt, K., Scudder, A., Obiorah, T., Essuman, K., Milbrandt, J., Diantonio, A., Ray, K., Snyder, M. L., Deredge, D., Snyder, G. A.. 2023-05-20. The structure of NAD+ consuming protein Acinetobacter baumannii TIR domain shows unique kinetics and conformations.. https://doi.org/10.1101/2023.05.19.541320

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