Fig. 5.
Proofreading factors, F, for codon selection by ternary complex increase linearly with increasing aa-tRNA affinity to EF-Tu at unaltered accuracy (Anf) of EF-Tu−free codon selection. F and Anf from different tRNA mutants are plotted against the tRNA affinities to EF-Tu (KA). Measurements are shown for Glu-tRNAGlu mutants misreading (A) GAU, (B) GGA, and (C) GAC and Phe-tRNAPhe mutants misreading (D) CUC, (E) UCC, and (F) UUA by their ternary complexes (F, ▪) or aa-tRNAs only (Anf, ○). In each case, linear regression was used to obtain maximum likelihood estimates of the y axis intercepts F2 and Anf as (A) 8.2 ± 5.0 and 12 ± 0.8, (B) 23 ± 8 and 25 ± 2, (C) 12 ± 5 and 16 ± 1, (D) 34 ± 4 and 39 ± 2, (E) 14 ± 2 and 12 ± 0.3, and (F) 27 ± 6 and 14 ± 0.4. Errors of F2 and Anf estimates represent SD from data fitting procedure (see SI Materials and Methods). Data in the plots represent weighted averages from at least two independent experiments ± propagated SD in both dimensions. Measurements were performed in polymix buffer containing 2.3 mM free Mg2+.