Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:6.1.1.10 (methionyl-tRNA synthetase)
 387 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Photoinduced covalent cross-linking has been used to identify a common surface of four methionine-accepting tRNAs which interact specifically with the Escherichia coli methionine:tRNA ligase (EC 6.1.1.10). tRNA--ligase mixtures were irradiated, and the covalently linked complexes were isolated and digested with T1 RNase (Schimmel & Budzik, 1977). The fragments lost from the elution profile of the T1 RNase digest were considered to have been cross-linked to the protein and therefore in intimate contact with the enzyme. Only specific cognate tRNA--ligase pairs produce covalently linked complexes. The four substrate tRNAs used in this study have substantially different sequences, but all showed a common cross-linking pattern, supporting the view that the sites cross-linked to the enzyme reflect the functionally common contact surface rather than particularly photoreactivity regions of tRNA. The cross-linked contact surface is comprised of three regions: (1) the narrow groove of the anticodon stem and its extension into the anticodon loop; (2) the 3' terminal residues; and (3) the 3' side of the "T arm". Unlike previous studies with other tRNAs, the D arm is not involved and significant radiation damage is suffered by the tRNA which must be taken into account in the analysis. The results are consistent with and complement chemical modification studies [Schulman, L. H., & Pelka, H. (1977) Biochemistry 16, 4256].
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PMID:Photocross-linking analysis of the contact surface of tRNA Met in complexes with Escherichia coli methionine:tRNA ligase. 36 5

Maturation of precursor transfer RNA (pre-tRNA) includes excision of the 5' leader and 3' trailer sequences, removal of introns and addition of the CCA terminus. Nucleotide modifications are incorporated at different stages of tRNA processing, after the RNA molecule adopts the proper conformation. In bacteria, tRNA(Ile2) lysidine synthetase (TilS) modifies cytidine into lysidine (L; 2-lysyl-cytidine) at the first anticodon of tRNA(Ile2) (refs 4-9). This modification switches tRNA(Ile2) from a methionine-specific to an isoleucine-specific tRNA. However, the aminoacylation of tRNA(Ile2) by methionyl-tRNA synthetase (MetRS), before the modification by TilS, might lead to the misincorporation of methionine in response to isoleucine codons. The mechanism used by bacteria to avoid this pitfall is unknown. Here we show that the TilS enzyme specifically recognizes and modifies tRNA(Ile2) in its precursor form, thereby avoiding translation errors. We identified the lysidine modification in pre-tRNA(Ile2) isolated from RNase-E-deficient Escherichia coli and did not detect mature tRNA(Ile2) lacking this modification. Our kinetic analyses revealed that TilS can modify both types of RNA molecule with comparable efficiencies. X-ray crystallography and mutational analyses revealed that TilS specifically recognizes the entire L-shape structure in pre-tRNA(Ile2) through extensive interactions coupled with sequential domain movements. Our results demonstrate how TilS prevents the recognition of tRNA(Ile2) by MetRS and achieves high specificity for its substrate. These two key points form the basis for maintaining the fidelity of isoleucine codon translation in bacteria. Our findings also provide a rationale for the necessity of incorporating specific modifications at the precursor level during tRNA biogenesis.
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PMID:Structural basis for translational fidelity ensured by transfer RNA lysidine synthetase. 1984 69