Nucleic Acids Research, 2012, Vol. 40, No. 11 4973 Scheme 1. Formation of Asn-tRNAAsn in H. pylori. (A) The major pathway used in H. pylori (the main ‘Asn-transamidosome pathway’) starts with (1) the formation of the tRNPC, linking GatCAB and tRNAAsn, onto which (2) ND-AspRS can then dock to form the Asn-transamidosome. In this ribonucleoprotein, tRNAAsn gets aspartylated more efﬁciently (3), and the aspartylated CCA-end of tRNAAsn shifts from the ND-AspRS active site to the GatCAB phosphorylation/amidation active site. Asn-tRNAAsn biosynthesis proceeds thanks to amide donor hydrolysis (here free Gln) to provide the ammonia moiety necessary for transamidation (4) (28). When Asn-tRNAAsn is formed, it is released together with ND-AspRS, and used in protein synthesis (5). (B) When GatCAB is not in sufﬁcient amounts to produce enough tRNPC, free tRNAAsn binds to ND-AspRS (6) and gets aminoacylated less efﬁciently (7). The dissociation of Asp-tRNAAsn is rate-limiting, but if Asp-tRNAAsn is released (8), GatCAB can still recognize it (9) and transamidate it (10) (28) into Asn-tRNAAsn, which can be released to fuel translation (11). (C) Since the release of Asp-tRNAAsn from ND-AspRS is rate-limiting, Asp-tRNAAsn remains within the ND-AspRS active site for longer times, enabling free GatCAB to bind to the ND-AspRS/Asp-tRNAAsn complex (12) to form an Asn-transamidosome, in which Asn-tRNAAsn can be synthesized through transamidation (4) rejoining the main Asn-transamidosome cycle. This latter mechanism can be qualiﬁed as a ‘rescue mechanism’. The table summarizes the different pathway characteristics. The aspartylation and transamidation steps are highlighted to stress the fact that the Asn-transamidosome pathway couples both reactions, while they are sequential when GatCAB is absent or when the tRNPC is not involved.