And Type ¢ó Effect The Protein Hopu1 Structure Its Substrate Grp7, Adp-ribose Methylation,

TypeⅢprotein secretion systems (T3SS) is a molecular syringe, which specifically inject bacterial proteins called effectors directly into host cells of many pathogens of plants and animals. Pseudomonas syringae pv. Tomato DC3000 delivers its effector protein HopUl into Arabidopsis thaliana and tomato by T3SS to suppress plant immune responses. HopU1 is the first identified mono-ADP-ribosyltransferase in plant pathogen and its substrate GRP7, a glycine-rich RNA-binding protein is also new for all known ADP-ribosyltransferases. HopU1 displays its strong impact on plant physiological activity through transferring single ADP-ribose to 47and 49 Arginine residue of GRP7, which is essential for plant innate immunity. Although ADP-ribosyltransferases have been well characterized in animals, no plant pathogen ADP-ribosyltransferase structure has been reported. Here, we report the crystal structure of HopU1 at resolution of 2.7 A. The overall structure of HopU1 adopts a typical "fourβ-stranded core" fold with conserved ARTT and PN loops, which are similar to other ADP-ribosyltransferase structures. Intriguingly, HopUl has two unique loops (L1 and L4), which are different from other ADP-ribosyltransferases. Systematic structural and biochemical studies demonstrated that L1, L4 and ARTT loops are essential for substrate recognition and corresponding enzymatic activity. Basing on these analyses, we propose a structure model for HopU1-(3NAD-GRP7 complex. In summary, the results reveal the reaction mechanism and substrate recognition of HopU1, the first identified ADP-ribosyltransferase in plant pathogen.