Abstract
The universally conserved His-66 of elongation factor Tu (EF-Tu) stacks on the side chain of the esterified Phe of Phe-tRNA(Phe). The affinities of eight aminoacyl-tRNAs were differentially destabilized by the introduction of the H66A mutation into Escherichia coli EF-Tu, whereas Ala-tRNA(Ala) and Gly-tRNA(Gly) were unaffected. The H66F and H66W proteins each show a different pattern of binding of 10 different aminoacyl-tRNAs, clearly showing that this position is critical in establishing the specificity of EF-Tu for different esterified amino acids. However, the H66A mutation does not greatly affect the ability of the ternary complex to bind ribosomes, hydrolyze GTP, or form dipeptide, suggesting that this residue does not directly participate in ribosomal decoding. Selective mutation of His-66 may improve the ability of certain unnatural amino acids to be incorporated by the ribosome.
Highlights
The cognate-esterified amino acid is critical for optimal delivery of aminoacyl-tRNAs to the ribosome by elongation factor Tu (EF-Tu)
One goal of this paper is to evaluate the contribution of His-66 in E. coli EF-Tu to the specificity of the protein for binding different esterified amino acids
The H66A Mutation Only Destabilizes Binding to Certain aa-tRNAs—As shown in Fig. 1, the imidazole ring of His-66 is stacked between the esterified phenylalanine and the side chain of Val-79 and is positioned 4 – 6 Å from three potential hydrogen bond partners Gln-97, Asn-273, and Glu-215
Summary
The cognate-esterified amino acid is critical for optimal delivery of aminoacyl-tRNAs to the ribosome by EF-Tu. Results: Mutation of His-66 in EF-Tu alters the specific binding of many, but not all aminoacyl-tRNAs, but does not affect decoding. Conclusion: His-66 is critical for the specificity of EF-Tu for the esterified amino acid. Significance: Selective mutation of His-66 could improve the incorporation efficiency of unnatural amino acids into proteins. The H66F and H66W proteins each show a different pattern of binding of 10 different aminoacyltRNAs, clearly showing that this position is critical in establishing the specificity of EF-Tu for different esterified amino acids. Selective mutation of His-66 may improve the ability of certain unnatural amino acids to be incorporated by the ribosome
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
