Two Acidic Residues of Escherichia coli Methionyl-tRNA Synthetase Act as Negative Discriminants Towards the Binding of Non-cognate tRNA Anticodons

Schmitt, Emmanuelle; Meinnel, Thierry; Panvert, Michel; Mechulam, Yves; Blanquet, Sylvain

doi:10.1006/jmbi.1993.1540

Cited by 53 publications

(35 citation statements)

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“…This finding is consistent with the structural data of Taq and E. coli MutS-mismatch complexes that predict electrostatic repulsion of Glu with the phosphate backbone of a normal DNA duplex (6,7). Supporting evidence for the role of acidic residues in enhancing substrate selectivity stems from studies of Glu and Asp residues in the tRNA and DNA binding sites of E. coli methionyl-tRNA synthetase and E. coli RuvA, respectively (25,26). Alteration of the charge by amino acid substitution resulted in increased binding of noncognate tRNAs by methionyltRNA synthetase and duplex DNA by RuvA.…”

Section: Effect Of a Glu To Ala Mutation In The Mismatch Binding Sitesupporting

confidence: 85%

The Phe-X-Glu DNA Binding Motif of MutS

Schofield

Brownewell²,

Nayak

et al. 2001

Journal of Biological Chemistry

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The crystal structures of MutS protein from Thermus aquaticus and Escherichia coli in a complex with a mismatch-containing DNA duplex reveal that the Glu residue in a conserved Phe-X-Glu motif participates in a hydrogen-bonded contact with either an unpaired thymidine or the thymidine of a G-T base-base mismatch. Here, the role of hydrogen bonding in mismatch recognition by MutS is assessed. The relative affinities of MutS for DNA duplexes containing nonpolar shape mimics of A and T, 4-methylbenzimidazole (Z), and difluorotoluene (F), respectively, that lack hydrogen bonding donors and acceptors, are determined in gel mobility shift assays. The results provide support for an induced fit mode of mismatch binding in which duplexes destabilized by mismatches are preferred substrates for kinking by MutS. Hydrogen bonding between the O⑀2 group of Glu and the mismatched base contributes only marginally to mismatch recognition and is significantly less important than the aromatic ring stack with the conserved Phe residue. A MutS protein in which Ala is substituted for Glu 38 is shown to be defective for mismatch repair in vivo. DNA binding studies reveal a novel role for the conserved Glu residue in the establishment of mismatch discrimination by MutS.

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Section: Effect Of a Glu To Ala Mutation In The Mismatch Binding Sitesupporting

confidence: 85%

The Phe-X-Glu DNA Binding Motif of MutS

Schofield

Brownewell²,

Nayak

et al. 2001

Journal of Biological Chemistry

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“…These changes render the tRNAs aminoacylatable with valine and isoleucine, respectively (7,22). As previously reported (7), valyl-tRNA f Met (GAC) can be formylated by FMTec, although the efficiency of the reaction is reduced by a factor of 3000 as compared with that measured with the natural methionylated substrate (Table I).…”

Section: Specificity Toward the Aminoacyl Group Esterified Tosupporting

confidence: 67%

Recognition of tRNAs by Methionyl-tRNA Transformylase from Mammalian Mitochondria

Takeuchi

Vial

Panvert

et al. 2001

Journal of Biological Chemistry

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Protein synthesis involves two methionine-isoaccepting tRNAs, an initiator and an elongator. In eubacteria, mitochondria, and chloroplasts, the addition of a formyl group gives its full functional identity to initiator MettRNA Met . In Escherichia coli, it has been shown that the specific action of methionyl-tRNA transformylase on Met-tRNA f Met mainly involves a set of nucleotides in the acceptor stem, particularly a C 1 A 72 mismatch. In animal mitochondria, only one tRNA Met species has yet been described. It is admitted that this species can engage itself either in initiation or elongation of translation, depending on the presence or absence of a formyl group. In the present study, we searched for the identity elements of tRNA Met that govern its formylation by bovine mitochondrial transformylase. The main conclusion is that the mitochondrial formylase preferentially recognizes the methionyl moiety of its tRNA substrate. Moreover, the relatively small importance of the tRNA acceptor stem in the recognition process accounts for the protection against formylation of the mitochondrial tRNAs that share with tRNA Met an A 1 U 72 motif.

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“…Substitution of highly conserved residues Trp-461, Asn-452, or Arg-395 shows their involvement in the binding of MetRS to anticodon site of tRNA (8)(9)(10). Two negatively charged residues, Asp-449 and Asp-456, also contribute to the overall specificity of the anticodon recognition, although they do not make direct contact with tRNA (11). Because of their acidic nature, these residues are able to reject all other noncognate tRNAs from binding to the anticodon sites of MetRS.…”

mentioning

confidence: 99%

A study of communication pathways in methionyl- tRNA synthetase by molecular dynamics simulations and structure network analysis

Ghosh

Vishveshwara

2007

Proc. Natl. Acad. Sci. U.S.A.

229

288

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The enzymes of the family of tRNA synthetases perform their functions with high precision by synchronously recognizing the anticodon region and the aminoacylation region, which are separated by Ϸ70 Å in space. This precision in function is brought about by establishing good communication paths between the two regions. We have modeled the structure of the complex consisting of Escherichia coli methionyl-tRNA synthetase (MetRS), tRNA, and the activated methionine. Molecular dynamics simulations have been performed on the modeled structure to obtain the equilibrated structure of the complex and the cross-correlations between the residues in MetRS have been evaluated. Furthermore, the network analysis on these simulated structures has been carried out to elucidate the paths of communication between the activation site and the anticodon recognition site. This study has provided the detailed paths of communication, which are consistent with experimental results. Similar studies also have been carried out on the complexes (MetRS ؉ activated methonine) and (MetRS ؉ tRNA) along with ligand-free native enzyme. A comparison of the paths derived from the four simulations clearly has shown that the communication path is strongly correlated and unique to the enzyme complex, which is bound to both the tRNA and the activated methionine. The details of the method of our investigation and the biological implications of the results are presented in this article. The method developed here also could be used to investigate any protein system where the function takes place through longdistance communication.dynamic cross-correlations ͉ methionyl-AMP ͉ protein structure network ͉ shortest pathways of communication ͉ stacking A crucial step in the translation of the genetic code is the aminoacylation of tRNA, which involves the molecular recognition between the aminoacyl-tRNA synthetases (aaRS) and their cognate tRNA. Each synthetase consists of the catalytic domain and the anticodon domain that are separated by Ϸ70 Å. Each tRNA connects these two regions with its anticodon and the acceptor stems. The mechanism of differentiations between cognate and noncognate tRNAs depends on contacts of anticodon domain of synthetase and anticodon stem of tRNA. The efficiency of the selection mechanism controls the overall accuracy of protein synthesis (1, 2). Recognition of the protein (aaRS) and the tRNA is explained by using the induced-fit mechanism, which suggests conformational changes in protein, tRNA, or both, leading to the final bound complex (3). However, the details of communication between the anticodon region and the aminoacylation region are less understood.In all living cells, protein synthesis starts with methionine.

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Two Acidic Residues of Escherichia coli Methionyl-tRNA Synthetase Act as Negative Discriminants Towards the Binding of Non-cognate tRNA Anticodons

Cited by 53 publications

References 0 publications

The Phe-X-Glu DNA Binding Motif of MutS

The Phe-X-Glu DNA Binding Motif of MutS

Recognition of tRNAs by Methionyl-tRNA Transformylase from Mammalian Mitochondria

A study of communication pathways in methionyl- tRNA synthetase by molecular dynamics simulations and structure network analysis

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