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Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System
Published on: August 1, 2016
Structural basis for anticodon recognition by discriminating glutamyl-tRNA synthetase
S Sekine1, O Nureki, A Shimada
1Cellular Signaling Laboratory, RIKEN Harima Institute at SPring-8, 1-1-1 Kouto, Mikazuki-cho, Sayo, Hyogo 679-5148, Japan.
Nature Structural Biology
|February 27, 2001
Summary
Researchers elucidated how discriminating glutamyl-tRNA synthetases (GluRS) evolved. A single arginine residue (Arg 358) in Thermus thermophilus GluRS dictates specificity, distinguishing between glutamyl-tRNA (tRNAGlu) and glutaminyl-tRNA (tRNAGln).
Area of Science:
- Molecular Biology
- Structural Biology
- Biochemistry
Background:
- Glutamyl-tRNA synthetases (GluRSs) exist in two forms: 'non-discriminating' and 'discriminating', depending on the presence of glutaminyl-tRNA synthetase (GlnRS) in the translation system.
- Non-discriminating GluRSs aminoacylate both tRNAGlu and tRNAGln, while discriminating GluRSs selectively aminoacylate only tRNAGlu.
Purpose of the Study:
- To determine the structural basis for anticodon discrimination by a 'discriminating' GluRS.
- To elucidate the molecular mechanism underlying the evolution of GluRS specificity.
Main Methods:
- X-ray crystallography of a 'discriminating' GluRS.tRNAGlu complex from Thermus thermophilus at 2.4 A resolution.
- Site-directed mutagenesis to investigate the role of specific residues in anticodon recognition.
Main Results:
- The crystal structure reveals that GluRS recognizes the tRNAGlu anticodon via two alpha-helical domains that maintain base stacking.
- A single arginine residue (Arg 358) was identified as the key determinant for discriminating between the glutamyl-tRNA (tRNAGlu) and glutaminyl-tRNA (tRNAGln) anticodons.
- Mutation of Arg 358 to glutamine abolished the discrimination, resulting in a GluRS that acts like a 'non-discriminating' enzyme.
Conclusions:
- The study reveals the precise molecular mechanism by which discriminating GluRS evolved to selectively aminoacylate tRNAGlu.
- The findings provide insights into the evolutionary trajectory of aminoacyl-tRNA synthetases and the genetic code.
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