title: The asparagine-transamidosome from Helicobacter pylori: a dual-kinetic mode in non-discriminating aspartyl-tRNA synthetase safeguards the genetic code.
abstract: Helicobacter pylori catalyzes Asn-tRNA(Asn) formation by use of the indirect pathway that involves charging of Asp onto tRNA(Asn) by a non-discriminating aspartyl-tRNA synthetase (ND-AspRS), followed by conversion of the mischarged Asp into Asn by the GatCAB amidotransferase. We show that the partners of asparaginylation assemble into a dynamic Asn-transamidosome, which uses a different strategy than the Gln-transamidosome to prevent the release of the mischarged aminoacyl-tRNA intermediate. The complex is described by gel-filtration, dynamic light scattering and kinetic measurements. Two strategies for asparaginylation are shown: (i) tRNA(Asn) binds GatCAB first, allowing aminoacylation and immediate transamidation once ND-AspRS joins the complex; (ii) tRNA(Asn) is bound by ND-AspRS which releases the Asp-tRNA(Asn) product much slower than the cognate Asp-tRNA(Asp); this kinetic peculiarity allows GatCAB to bind and transamidate Asp-tRNA(Asn) before its release by the ND-AspRS. These results are discussed in the context of the interrelation between the Asn and Gln-transamidosomes which use the same GatCAB in H. pylori, and shed light on a kinetic mechanism that ensures faithful codon reassignment for Asn.
authors.affiliation: Institut de Biologie Moléculaire et Cellulaire, UPR 9002 du CNRS, Architecture et Réactivité de l'ARN, Université de Strasbourg, 15 rue René Descartes, 67084 Strasbourg Cedex, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Fischer
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Huot
authors.forename: Bernard
authors.initials: B
authors.lastname: Lorber
authors.forename: Guillaume
authors.initials: G
authors.lastname: Diss
authors.forename: Tamara L
authors.initials: TL
authors.lastname: Hendrickson
authors.forename: Hubert D
authors.initials: HD
authors.lastname: Becker
authors.forename: Jacques
authors.initials: J
authors.lastname: Lapointe
authors.forename: Daniel
authors.initials: D
authors.lastname: Kern
mesh_terms: D001216:Asparagine / Q000378:metabolism; D001226:Aspartate-tRNA Ligase / Q000378:metabolism*; D001224:Aspartic Acid / Q000378:metabolism; D005815:Genetic Code; D016480:Helicobacter pylori / Q000201:enzymology*; D007700:Kinetics; D019884:Nitrogenous Group Transferases / Q000378:metabolism*; D012348:RNA, Transfer, Asn / Q000378:metabolism*; D012349:RNA, Transfer, Asp / Q000378:metabolism; D046249:Transfer RNA Aminoacylation
chemical_list: D012348:RNA, Transfer, Asn; D012349:RNA, Transfer, Asp; D001224:Aspartic Acid; D001216:Asparagine; C525150:GatCAB amidotransferase, Helicobacter pylori; D019884:Nitrogenous Group Transferases; D001226:Aspartate-tRNA Ligase