PMID:15845537
| Citation |
Uter, NT, Gruic-Sovulj, I and Perona, JJ (2005) Amino acid-dependent transfer RNA affinity in a class I aminoacyl-tRNA synthetase. J. Biol. Chem. 280:23966-77 |
|---|---|
| Abstract |
Steady-state and transient kinetic analyses of glutaminyl-tRNA synthetase (GlnRS) reveal that the enzyme discriminates against noncognate glutamate at multiple steps during the overall aminoacylation reaction. A major portion of the selectivity arises in the amino acid activation portion of the reaction, whereas the discrimination in the overall two-step reaction arises from very weak binding of noncognate glutamate. Further transient kinetics experiments showed that tRNA(Gln) binds to GlnRS approximately 60-fold weaker when noncognate glutamate is present and that glutamate reduces the association rate of tRNA with the enzyme by 100-fold. These findings demonstrate that amino acid and tRNA binding are interdependent and reveal an important additional source of specificity in the aminoacylation reaction. Crystal structures of the GlnRS x tRNA complex bound to either amino acid have previously shown that glutamine and glutamate bind in distinct positions in the active site, providing a structural basis for the amino acid-dependent modulation of tRNA affinity. Together with other crystallographic data showing that ligand binding is essential to assembly of the GlnRS active site, these findings suggest a model for specificity generation in which required induced-fit rearrangements are significantly modulated by the identities of the bound substrates. |
| Links |
PubMed Online version:10.1074/jbc.M414259200 |
| Keywords |
Amino Acids/metabolism; Amino Acyl-tRNA Synthetases/chemistry; Amino Acyl-tRNA Synthetases/metabolism; Binding Sites; Crystallography, X-Ray; Escherichia coli/enzymology; Kinetics; Models, Molecular; Protein Binding; RNA, Transfer/metabolism |
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