2013 Rustbelt RNA Meeting
Many microorganisms, including Helicobacter pylori, rely on an indirect pathway for the synthesis of Gln-tRNAGln and/or Asn-tRNAAsn. In these cases, Asn-tRNAAsn production proceeds via misacylation of tRNAAsn (catalyzed by a non-discriminating aspartyl-tRNA synthetase, ND-AspRS) to generate Asp-tRNAAsn, which is subsequently converted to Asn-tRNAAsn by an amidotransferase (AdT). Gln-tRNAGln is produced via an analogous process, relying on a misacylating glutamyl-tRNA synthetase (either ND-GluRS or GluRS2) and AdT. However, efficient delivery of both misacylated tRNAs from the two misacylating enzymes to AdT is required to ensure the stability of the aminoacyl ester bond and to minimize translational errors. Some bacteria, like T. thermophilus, utilize a tRNA-dependent complex called the transamidosome, which contains AdT, tRNAAsn, and ND-AspRS. This Asn-transamidosome traps misacylated Asp-tRNAAsn until it is converted to Asn-tRNAAsn. Recent studies have shown that the H. pylori Asn-transamidosome is more dynamic, suggesting a requirement for an alternative mechanism to facilitate assembly of a stable complex. We have identified a novel ATPase called Hp0100 that is a component of a stable, tRNA-independent H. pylori transamidosome. Hp0100 contains two distinct ATPase active sites, which are activated by misacylated tRNAs. Hp0100 dramatically enhances the capacity of AdT to convert Asp-tRNAAsn into Asn-tRNAAsn but has minimal effects on ND-AspRS function. Our results highlight the importance of the novel ATPase Hp0100, for the stable assembly and function of H. pylori transamidosome.
1. Silva, G. N., Fatma, S., Floyd, A. M., Fischer, F., Chuawong, P., Cruz, A. N., Simari, R. M., Joshi, N., Kern, D., and Hendrickson, T. L. (2013) A tRNA-independent mechanism for transamidosome assembly promotes aminoacyl-tRNA transamidation. J. biol. chem. 288, 3816-3822
2. Fischer F., Huot J. L., Lorber B., Diss G., Hendrickson T. L., Becker H. D., Lapointe J., Kern D. (2012) The asparagine-transamidosome from Helicobacter pylori: a dual-kinetic mode in non-discriminating aspartyl-tRNA synthetase safeguards the genetic code. Nucleic Acids Res. 40, 4965–4976
Keywords: Transamidosome, Non-discriminating Aspartyl-tRNA Synthetase, Hp0100