Poster Presentation The 47th Lorne Conference on Protein Structure and Function 2022

Structural studies of the active state assembly and inhibition mechanism of SARM1-TIR domain (#168)

Forhad K Saikot 1 , Thomas Ve 2 , Jeffrey D Nanson 1 , Robert O Hughes 3 , Weixi Gu 1 , Philip S Kerry 4 , Michael J Landsberg 1 , Bostjan Kobe 1 5 6
  1. School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD, Australia
  2. Institute for Glycomics, Griffith University, Southport, Qld, Australia
  3. Disarm Therapeutics, Cambridge, MA, USA
  4. Evotec, Abingdon, Oxfordshire, UK
  5. Institute for Molecular Biosciences, The University of Queensland, Brisbane, Qld, Australia
  6. Australian Infectious Disease Research Centre, University of Queensland, Brisbane, Qld, Australia

SARM1 is a pro-axon degenerative NADase protein. Injury or stress to CNS leads to an increase in the NMN/NAD+ ratio, which triggers SARM1 activation. Upon activation, SARM1 rapidly depletes NAD+ in the affected cells and thereby causes axon degeneration. Recently, the inactive structure of SARM1 was solved. Both NMN and NAD+ can bind to the autoinhibitory ARM domain of SARM1; however, NMN activates and NAD+ retains the inhibitory state of the protein. In stressed cells, these two molecules compete, and NMN replaces NAD+ to activate the protein. In the active form, the catalytic pockets of the effector TIR domains bind and cleave NAD+ into Nam (nicotinamide) and ADPR/cADPR. This NADase activity of the TIR domain is self-assembly dependent. Although the molecular mechanism of SARM1 dependent axon degeneration is being unravelled, there are still a few gaps. The structural basis of the interaction of TIR with NAD+ has been elusive. In this study, we aimed to understand the assembly of the active form of SARM1-TIR through cryo-electron microscopy (cryo-EM). To achieve this, we are using a compound we termed the “adduct”. The adduct is the product of a base-exchange reaction of a novel small molecule inhibitor, 5-iodoisoquinoline, with the Nam portion of NAD+ in the presence of TIR domains. These adducts can orthosterically bind to the catalytic pocket and stabilize the oligomeric active state of the TIR domain. Therefore, the TIR oligomeric active state assembly becomes imageable using cryo-EM and we were able to solve the structure. The structure of oligomeric TIR reveals a double-stranded architecture similar to a previously solved crystal structure of TIR domains. These together not only explain the activation mechanism of the TIR, but also pave a way of post-injury inhibition of this executor of axon degeneration.