Abstact

2'-5'-Oligoadenylate synthetases (OAS) are crucial innate immune sensors that activate antiviral responses upon detecting viral double-stranded RNA. Understanding the molecular mechanism of OAS is vital for advancing immunomodulatory therapies. This study provides a detailed enzymatic mechanism of the OAS, integrating structural, kinetic, and quantum chemical analyses. Crystallographic data of the OAS1 postreactive complexes shed light on the geometry of OAS1 following product formation and dissociation, the sequential order of product release, and the pivotal role of divalent metal ions in catalysis. Our data reveal the unanticipated involvement of a third metal ion, which may play a transient supporting role in the catalytic cycle. Moreover, they highlight the central role of quantum mechanisms in the OAS function. Strikingly, substituting catalytic Mg(2+) with Mn(2+) ions increases the substrate binding rate 9-fold and activates OAS for catalysis. The results of this study are pertinent to the OAS/cGAS family of innate immune sensors and offer insights that can be applied to a broader class of nucleotidyltransferases, which play key roles in various biological processes.