Mechanistic basis for the allosteric activation of NADase activity in the Sir2-HerA antiphage defense system

Sir2-HerA is a widely distributed antiphage system composed of a RecA-like ATPase (HerA) and an effector with potential NADase activity (Sir2). Sir2-HerA is believed to provide defense against phage infection in Sir2-dependent NAD⁺ depletion to arrest the growth of infected cells. However, the detailed mechanism underlying its antiphage activity remains largely unknown. Here, we report functional investigations of Sir2-HerA from Staphylococcus aureus (SaSir2-HerA), unveiling that the NADase function of SaSir2 can be allosterically activated by the binding of SaHerA, which then assembles into a supramolecular complex with NADase activity. By combining the cryo-EM structure of SaSir2-HerA in complex with the NAD⁺ cleavage product, it is surprisingly observed that Sir2 protomers that interact with HerA are in the activated state, which is due to the opening of the α15-helix covering the active site, allowing NAD⁺ to access the catalytic pocket for hydrolysis. In brief, our study provides a comprehensive view of an allosteric activation mechanism for Sir2 NADase activity in the Sir2-HerA immune system.