Abstract
SIR2-HerA, a bacterial two-protein anti-phage defense system, induces bacterial death by depleting NAD+ upon phage infection. Biochemical reconstitution of SIR2, HerA, and the SIR2-HerA complex reveals a dynamic assembly process. Unlike other ATPases, HerA can form various oligomers, ranging from dimers to nonamers. When assembled with SIR2, HerA forms a hexamer and converts SIR2 from a nuclease to an NAD+ hydrolase, representing an unexpected regulatory mechanism mediated by protein assembly. Furthermore, high concentrations of ATP can inhibit NAD+ hydrolysis by the SIR2-HerA complex. Cryo-EM structures of the SIR2-HerA complex reveal a giant supramolecular assembly up to 1 MDa, with SIR2 as a dodecamer and HerA as a hexamer, crucial for anti-phage defense. Unexpectedly, the HerA hexamer resembles a spiral staircase and exhibits helicase activities toward dual-forked DNA. Together, we reveal the supramolecular assembly of SIR2-HerA as a unique mechanism for switching enzymatic activities and bolstering anti-phage defense strategies.