Acidic pH irreversibly activates the signaling enzyme SARM1

SARM1, an executioner in axon degeneration, is an autoinhibitory NAD-consuming enzyme, composed of multiple domains. NMN and its analogs, CZ-48 and VMN, are the only known activators, which can release the inhibitory ARM domain from the enzymatic TIR domain. Here, we document that acid can also activate SARM1, even more efficiently than NMN, possibly via the protonation of the negative residues. Systematic mutagenesis revealed that a single mutation, E689Q in TIR, led to the constitutive activation of SARM1. It forms a salt bridge with R216 in the neighboring ARM, maintaining the autoinhibitory structure. Using this 'acid activation' protocol, mutation K597E was found to inhibit activation, while H685A eliminated SARM1 catalytic activity, revealing two distinct inhibitory mechanisms. The protocol has also been applied to differentiate two classes of chemical inhibitors. NAD, dHNN, disulfiram, CHAPS, and TRX-100 mainly inhibited the activation process, while nicotinamide and Tweens mainly inhibited SARM1 catalysis. Taken together, we demonstrate a new mechanism for SARM1 activation and decipher two distinct inhibitory mechanisms of SARM1.

Automated summary

SARM1 can be activated by NMN, CZ-48, VMN and acid, with acid being even more effective through protonation of negative residues. Mutations can lead to constitutive activation of SARM1, with the E689Q mutation forming a salt bridge with R216 to maintain the autoinhibitory structure. Two inhibitory mechanisms of SARM1, through K597E mutation to inhibit activation and H685A mutation to eliminate catalytic activity, were revealed using an 'acid activation' protocol.