Extended Data Fig. 5: Structural analysis.

a, Comparison of GsSir2/Ago, CcSir2/Ago and PgSir2-Ago AlphaFold models with the X-ray structures of long pAgos. Structures are coloured by domains, schematic domain architecture is given above each structure. Guide RNA and target DNA strands are coloured red and blue, respectively. PDB ID codes for long pAgo structures are given in parentheses. RsAgo represents long-B group, TtAgo – longA group of long pAgos (based on Ryazansky et al. classification⁸). MpAgo has a distinct OH-type MID domain that is specific to 5’-OH instead of phosphate (MID domain classification – Ryazansky et al.⁸, MpAgo MID domain biochemical assay - Kaya et al.⁵²). In Sir2/Ago models the N, L1 and L2-like domains previously identified as the APAZ domain correspond to the analogous domains of long pAgos. b, Gs, Cc and Pg Sir2 structural models (cut from full-length models) compared to canonical Sir2 deacetylase TmSir2 and the Sir2 domain of Thoeris defence system protein ThsA. Structures are coloured based on secondary structure. Positions corresponding to ThsA Sir2 N112 and H152 are indicated in green. These residues have been shown to be critical for NAD⁺ hydrolysis in ThsA²⁴. GsSir2 D230 and corresponding positions in other structures are indicated in cyan. NAD⁺ was also superimposed on the ThsA Sir2 structure from TmSir2.