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PMID: 8692925 Published · ppublish English Comparative Study Journal Article Research Support, U.S. Gov't, P.H.S.

Interactions between tRNA identity nucleotides and their recognition sites in glutaminyl-tRNA synthetase determine the cognate amino acid affinity of the enzyme.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 93 ·No. 14 ·1996-07-09 ·Pages 6953-8

Ibba M, Hong KW, Sherman JM, Sever S, Söll D

Abstract

Sequence-specific interactions between aminoacyl-tRNA synthetases and their cognate tRNAs both ensure accurate RNA recognition and prevent the binding of noncognate substrates. Here we show for Escherichia coli glutaminyl-tRNA synthetase (GlnRS; EC 6.1.1.18) that the accuracy of tRNA recognition also determines the efficiency of cognate amino acid recognition. Steady-state kinetics revealed that interactions between tRNA identity nucleotides and their recognition sites in the enzyme modulate the amino acid affinity of GlnRS. Perturbation of any of the protein-RNA interactions through mutation of either component led to considerable changes in glutamine affinity with the most marked effects seen at the discriminator base, the 10:25 base pair, and the anticodon. Reexamination of the identity set of tRNA(Gln) in the light of these results indicates that its constituents can be differentiated based upon biochemical function and their contribution to the apparent Gibbs' free energy of tRNA binding. Interactions with the acceptor stem act as strong determinants of tRNA specificity, with the discriminator base positioning the 3' end. The 10:25 base pair and U35 are apparently the major binding sites to GlnRS, with G36 contributing both to binding and recognition. Furthermore, we show that E. coli tryptophanyl-tRNA synthetase also displays tRNA-dependent changes in tryptophan affinity when charging a noncognate tRNA. The ability of tRNA to optimize amino acid recognition reveals a novel mechanism for maintaining translational fidelity and also provides a strong basis for the coevolution of tRNAs and their cognate synthetases.

MeSH Terms
Amino Acyl-tRNA Synthetases/chemistry,metabolism Animals Base Sequence Binding Sites Calorimetry Cloning, Molecular Consensus Sequence Escherichia coli Humans Kinetics Models, Structural Molecular Sequence Data Nucleic Acid Conformation Protein Folding RNA, Transfer, Gln/chemistry,genetics Recombinant Proteins/chemistry,metabolism Sequence Homology, Nucleic Acid
Chemicals
RNA, Transfer, Gln Recombinant Proteins Amino Acyl-tRNA Synthetases glutaminyl-tRNA synthetase
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Ibba M
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520-8114, USA.
Hong K W
Sherman J M
Sever S
Söll D
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.93.14.6953
Published
1996-07-09
Pages
6953-8
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC38915
Subset
IM
Databases
GENBANK
K00181, K00182, M15679, M16251, M35400, M35401, M35402, U32863, X02445, X15813, X16590, X60715, X68534, X78550, X78552, X79104, Z67879
External Links
PubMed source DOI: 10.1073/pnas.93.14.6953 PMC Full Text View journal details
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