Elongation factor Tu (EF-Tu) binds all elongator aminoacyl-transfer RNAs (a
a-tRNAs) for delivery to the ribosome during protein synthesis. Here, we sh
ow that EF-Tu binds misacylated tRNAs over a much wider range of affinities
than it binds the corresponding correctly acylated tRNAs, suggesting that
the protein exhibits considerable specificity for both the amino acid side
chain and the tRNA body. The thermodynamic contributions of the amino acid
and the tRNA body to the overall binding affinity are independent of each o
ther and compensate for one another when the tRNAs are correctly acylated.
Because certain misacylated tRNAs bind EF-Tu significantly more strongly or
weakly than cognate aa-tRNAs, EF-Tu may contribute to translational accura
cy.