Structures and mechanism of E2‐CBASS anti‐phage system

Bacteria deploy diverse innate immune systems to combat bacteriophage infections. The cyclic‐oligonucleotide‐based anti‐phage signaling system (CBASS) is a type of innate prokaryotic immune system. CBASS synthesizes cyclic‐oligonucleotide through cGAS/DncV‐like nucleotidyltransferases (CD‐NTases) to activate downstream effectors, which kill bacteriophage‐infected bacteria, thereby stopping phage spread. One major class of CBASS contains a homolog of eukaryotic ubiquitin‐conjugating enzymes, either as an E1‐E2 fusion or a single E2 enzyme. Both enzymes function by regulating CD‐NTase activity. Currently, many structures of CD‐NTases have been reported, but there are only a few reports of structures where CD‐NTases form complexes with the associated E2. In this study, we analyzed the length and classification of the CD‐NTase in two types of type II CBASS—E1E2/JAB‐CBASS and E2‐CBASS. We found that the CD‐NTase in E2‐CBASS is longer and predominantly belongs to clade G. We also present the structure of the Sm CdnG‐ Sm E2 complex with the bound GTP substrate, which indicates the conservation of the donor binding pattern. Interestingly, we discovered that Sm CdnG contains a conserved C‐terminal α‐helix and β‐sheet structure, which is uniquely involved in forming a complex with Sm E2. We also found that the structure of the E2 protein in the E2‐CBASS system is highly conserved. Altogether, we provide mechanistic insights into the E2‐CBASS system.