title: Conserved discrimination against misacylated tRNAs by two mesophilic elongation factor Tu orthologs.
abstract: Elongation factor Tu (EF-Tu) binds and loads elongating aminoacyl-tRNAs (aa-tRNAs) onto the ribosome for protein biosynthesis. Many bacteria biosynthesize Gln-tRNA (Gln) and Asn-tRNA (Asn) by an indirect, two-step pathway that relies on the misacylated tRNAs Glu-tRNA (Gln) and Asp-tRNA (Asn) as intermediates. Previous thermodynamic and experimental analyses have demonstrated that Thermus thermophilus EF-Tu does not bind Asp-tRNA (Asn) and predicted a similar discriminatory response against Glu-tRNA (Gln) [Asahara, H., and Uhlenbeck, O. (2005) Biochemistry 46, 6194-6200; Roy, H., et al. (2007) Nucleic Acids Res. 35, 3420-3430]. By discriminating against these misacylated tRNAS, EF-Tu plays a direct role in preventing misincorporation of aspartate and glutamate into proteins at asparagine and glutamine codons. Here we report the characterization of two different mesophilic EF-Tu orthologs, one from Escherichia coli, a bacterium that does not utilize either Glu-tRNA (Gln) or Asp-tRNA (Asn), and the second from Helicobacter pylori, an organism in which both misacylated tRNAs are essential. Both EF-Tu orthologs discriminate against these misacylated tRNAs, confirming the prediction that Glu-tRNA (Gln), like Asp-tRNA (Asn), will not form a complex with EF-Tu. These results also demonstrate that the capacity of EF-Tu to discriminate against both of these aminoacyl-tRNAs is conserved even in bacteria like E. coli that do not generate either misacylated tRNA.
authors.affiliation: Department of Chemistry, Johns Hopkins University, 3400 North Charles Street, Baltimore, Maryland 21218, USA.
authors.forename: Terry J T
authors.initials: TJ
authors.lastname: Cathopoulis
authors.forename: Pitak
authors.initials: P
authors.lastname: Chuawong
authors.forename: Tamara L
authors.initials: TL
authors.lastname: Hendrickson
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D010444:Peptide Elongation Factor Tu / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012348:RNA, Transfer, Asn / Q000737:chemistry / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D012346:RNA, Transfer, Amino Acyl; D012348:RNA, Transfer, Asn; D010444:Peptide Elongation Factor Tu