Methanothermobacter thermautotrophicus tRNAGln Confines the Amidotransferase GatCAB to Asparaginyl-tRNAAsn Formation

Sheppard, Kelly; Sherrer, R. Lynn; Söll, Dieter

doi:10.1016/j.jmb.2008.01.064

Cited by 18 publications

(43 citation statements)

References 44 publications

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“…In contrast, archaeal GatCAB does not use the first base pair of the tRNA acceptor stem to distinguish Asp-tRNA Asn from Asp-tRNA Asp , recognizing aa-tRNA species with either a U1–A72 or a G1–C72 base pair (10,13,14). However, the process of distinguishing a U1–A72 base pair from a G1–C72 base pair is not known.…”

Section: Resultsmentioning

confidence: 99%

Two distinct regions in Staphylococcus aureus GatCAB guarantee accurate tRNA recognition

Nakamura¹,

Sheppard²,

Yamane³

et al. 2009

Nucleic Acids Research

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In many prokaryotes the biosynthesis of the amide aminoacyl-tRNAs, Gln-tRNAGln and Asn-tRNAAsn, proceeds by an indirect route in which mischarged Glu-tRNAGln or Asp-tRNAAsn is amidated to the correct aminoacyl-tRNA catalyzed by a tRNA-dependent amidotransferase (AdT). Two types of AdTs exist: bacteria, archaea and organelles possess heterotrimeric GatCAB, while heterodimeric GatDE occurs exclusively in archaea. Bacterial GatCAB and GatDE recognize the first base pair of the acceptor stem and the D-loop of their tRNA substrates, while archaeal GatCAB recognizes the tertiary core of the tRNA, but not the first base pair. Here, we present the crystal structure of the full-length Staphylococcus aureus GatCAB. Its GatB tail domain possesses a conserved Lys rich motif that is situated close to the variable loop in a GatCAB:tRNAGln docking model. This motif is also conserved in the tail domain of archaeal GatCAB, suggesting this basic region may recognize the tRNA variable loop to discriminate Asp-tRNAAsn from Asp-tRNAAsp in archaea. Furthermore, we identified a 310 turn in GatB that permits the bacterial GatCAB to distinguish a U1–A72 base pair from a G1–C72 pair; the absence of this element in archaeal GatCAB enables the latter enzyme to recognize aminoacyl-tRNAs with G1–C72 base pairs.

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Section: Resultsmentioning

confidence: 99%

Two distinct regions in Staphylococcus aureus GatCAB guarantee accurate tRNA recognition

Nakamura¹,

Sheppard²,

Yamane³

et al. 2009

Nucleic Acids Research

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“…E. coli total tRNA containing about 20 % M. catarrhalis tRNA Asn (see above) was 32 Plabelled on its 39-terminal phosphodiester link using the E. coli CCAadding enzyme and [a-32 P]ATP as previously described (Sheppard et al, 2007(Sheppard et al, , 2008, with some modifications as follows: briefly, 172 mM tRNA Asn in 50 mM Tris/HCl (pH 8.0), 20 mM MgCl 2 , 5 mM DTT and 0.02 mM NaPPi was incubated for 35 min at room temperature in the presence of 2 mg ml 21 (43 nM) pure CCA-adding enzyme from E. coli (Cudny & Deutscher, 1986) and 1 mCi ml 21 (37 kBq) [a-32 P]ATP (PerkinElmer and Analytical Sciences). Protein in 50 ml samples were phenol (Tris-buffered pH 7.9)/chloroform extracted, and the aqueous phase was filtered through Bio-Spin 30 columns to remove excess [a-32 P]ATP.…”

Section: Figmentioning

confidence: 99%

“…We used Gln, reported to be a better amide donor than Asn for bacterial GatCAB (Nakamura et al, 2006;Sheppard et al, 2007Sheppard et al, , 2008. M. catarrhalis tRNA Asn overproduced in E. coli (see Methods) was used to prepare Asp-tRNA Asn with P. aeruginosa AspRS as a substrate to test the efficiency of M. catarrhalis GatCAB variants in amidation of Asp-tRNA Asn to Asn-tRNA Asn .…”

Section: Overproduction and Purification Of M Catarrhalis Gatcabsmentioning

confidence: 99%

“…However, many bacterial genomes lack genes encoding an asparaginyl-tRNA synthetase (AsnRS) and/or a glutaminyl-tRNA synthetase (GlnRS), and form glutaminyl-tRNA Gln (GlntRNA Gln ) and asparaginyl-tRNA Asn (Asn-tRNA Asn ) via an indirect pathway by the GatCAB-mediated transamidation of glutamyl-tRNA Gln (Glu-tRNA Gln ) or aspartyl-tRNA Asn (AsptRNA Asn ) formed by a non-discriminating glutamyl-tRNA synthetase (ND-GluRS) or a non-discriminating aspartyltRNA synthetase (ND-AspRS), respectively (Becker & Kern, 1998;Curnow et al, 1996;Lapointe et al, 1986;Wilcox & Nirenberg, 1968). In nature, there is one type of bacterial amidotransferase (AdT), corresponding to GatCAB, with a dual specificity when both substrates Asp-tRNA Asn and GlutRNA Gln exist in vivo (Becker et al, 2000;Curnow et al, 1997;Raczniak et al, 2001;Tumbula et al, 2000), and two types of archaeal AdT, namely GatDE and GatCAB, which act only as a Glu-AdT or Asp-AdT, respectively (Sheppard et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

“…Similarly, most bacteria and all archaea lack GlnRS and use an ND-GluRS and GatCAB or GatDE as a Glu-tRNA AdT (Glu-AdT) to form Gln-tRNA Gln (Curnow et al, 1997;Horiuchi et al, 2001;Sheppard et al, 2007Sheppard et al, , 2008; reviewed by Huot et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

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Natural insertion of the bro-1 β-lactamase gene into the gatCAB operon affects Moraxella catarrhalis aspartyl-tRNAAsn amidotransferase activity

2012

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Only about half of bacterial species use an asparaginyl-tRNA synthetase (AsnRS) to attach Asn to its cognate tRNA Asn . Other bacteria, including the human pathogen Moraxella catarrhalis, a causative agent of otitis media, lack a gene encoding AsnRS, and form Asn-tRNA Asn by an indirect pathway catalysed by two enzymes: first, a non-discriminating aspartyl-tRNA synthetase (ND-AspRS) catalyses the formation of aspartyl-tRNA Asn (Asp-tRNA Asn ); then, a tRNAdependent amidotransferase (GatCAB) transamidates this 'incorrect' product into Asn-tRNA Asn . As M. catarrhalis has a Gln-tRNA synthetase, its GatCAB functions as an Asp-tRNA Asn amidotransferase. This pathogen rapidly evolved to about 90 % ampicillin resistance worldwide by insertion of a bro-1 b-lactamase gene within the gatCAB operon. Comparison of the GatCAB subunits from bro-1 b-lactamase-positive and bro-negative strains showed that the laterally transferred bro-1 gene, inserted into the gatCAB operon, affected the C-terminal sequence of GatA. The identity between the C-terminal sequences of GatA wt (residues 479-491) and of GatA BRO-1 (residues 479-492) was about 36 %, whereas the rest of the GatA sequence was relatively conserved. The characterization of these two distinct GatCABs as well as the hybrid GatCAB containing GatA(1-478) wt (479-492) BRO-1 and truncated GatCAB enzymes of M. catarrhalis showed that the substitution in GatA wt of residues 479-492 of GatA BRO-1 causes increased specificity for glutamine, and decreased specificity for Asp-tRNA Asn in the transamidation reaction. We conclude that the bro gene insertion has altered the kinetic parameters of Asp-tRNA Asn amidotransferase, and we propose a model for gatA evolution after the insertion of bro-1 at the carboxyl end of gatA.

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Evolution and variation in amide aminoacyl‐tRNA synthesis

Lewis,

Fallon,

Dittemore

et al. 2024

IUBMB Life

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The amide proteogenic amino acids, asparagine and glutamine, are two of the twenty amino acids used in translation by all known life. The aminoacyl‐tRNA synthetases for asparagine and glutamine, asparaginyl‐tRNA synthetase and glutaminyl tRNA synthetase, evolved after the split in the last universal common ancestor of modern organisms. Before that split, life used two‐step indirect pathways to synthesize asparagine and glutamine on their cognate tRNAs to form the aminoacyl‐tRNA used in translation. These two‐step pathways were retained throughout much of the bacterial and archaeal domains of life and eukaryotic organelles. The indirect routes use non‐discriminating aminoacyl‐tRNA synthetases (non‐discriminating aspartyl‐tRNA synthetase and non‐discriminating glutamyl‐tRNA synthetase) to misaminoacylate the tRNA. The misaminoacylated tRNA formed is then transamidated into the amide aminoacyl‐tRNA used in protein synthesis by tRNA‐dependent amidotransferases (GatCAB and GatDE). The enzymes and tRNAs involved assemble into complexes known as transamidosomes to help maintain translational fidelity. These pathways have evolved to meet the varied cellular needs across a diverse set of organisms, leading to significant variation. In certain bacteria, the indirect pathways may provide a means to adapt to cellular stress by reducing the fidelity of protein synthesis. The retention of these indirect pathways versus acquisition of asparaginyl‐tRNA synthetase and glutaminyl tRNA synthetase in lineages likely involves a complex interplay of the competing uses of glutamine and asparagine beyond translation, energetic costs, co‐evolution between enzymes and tRNA, and involvement in stress response that await further investigation.

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Methanothermobacter thermautotrophicus tRNAGln Confines the Amidotransferase GatCAB to Asparaginyl-tRNAAsn Formation

Cited by 18 publications

References 44 publications

Two distinct regions in Staphylococcus aureus GatCAB guarantee accurate tRNA recognition

Two distinct regions in Staphylococcus aureus GatCAB guarantee accurate tRNA recognition

Natural insertion of the bro-1 β-lactamase gene into the gatCAB operon affects Moraxella catarrhalis aspartyl-tRNAAsn amidotransferase activity

Evolution and variation in amide aminoacyl‐tRNA synthesis

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