The basic cycle of polypeptide elongation shown in outline in Figure 6–64 has an additional feature that makes translation especially efficient and accurate. Two elongation factors enter and leave the ribosome during each cycle, each hydrolyzing GTP to GDP and undergoing conformational changes in the process. These factors are called EF-Tu and EF-G in bacteria, and EF1 and EF2 in eukaryotes. Under some conditions in vitro, ribosomes can be forced to synthesize proteins
Figure 6–64 Translating an mRNA molecule. Each amino acid added to the growing end of a polypeptide chain is selected by complementary base-pairing between the anticodon on its attached tRNA molecule and the next codon on the mRNA chain. Because only one of the many types of tRNA molecules in a cell can base-pair with each codon, the codon determines the specific amino acid to be added to the growing polypeptide chain. The four-step cycle shown is repeated over and over during the synthesis of a protein. In step 1, an aminoacyl-tRNA molecule binds to a vacant A site on the ribosome. In step 2, a new peptide bond is formed. In step 3, the large subunit translocates relative to the small subunit, leaving the two tRNAs in hybrid sites: P on the large subunit and A on the small, for one; E on the large subunit and P on the small, for the other. In step 4, the small subunit translocates carrying its mRNA a distance of three nucleotides through the ribosome. This “resets” the ribosome with a fully empty A site, ready for the next aminoacyl-tRNA molecule to bind. As indicated, the mRNA is translated in the 5ʹ-to-3ʹ direction, and the N-terminal end of a protein is made first, with each cycle adding one amino acid to the C-terminus of the polypeptide chain (Movie 6.7 and Movie 6.8).
Figure 6–63 The path of mRNA (blue) through the small ribosomal subunit. The orientation is the same as that in the right-hand panel of Figure 6–62B. (Courtesy of Harry F. Noller, based on data in G.Z. Yusupova et al., Cell 106:233–241, 2001. With permission from Elsevier.)
Figure 6–65 Detailed view of the translation cycle. The outline of translation presented in Figure 6–64 has been expanded to show the roles of the two elongation factors EF-Tu and EF-G, which drive translation in the forward direction. As explained in the text, EF-Tu provides opportunities for proofreading of the codon–anticodon match. In this way, incorrectly paired tRNAs are selectively rejected, and the accuracy of translation is improved. The binding of a molecule of EF-G to the ribosome and the subsequent hydrolysis of GTP lead to a rearrangement of the ribosome structure, moving the mRNA being decoded exactly three nucleotides through it (Movie 6.9).
without the aid of these elongation factors and GTP hydrolysis, but this synthesis is very slow, inefficient, and inaccurate. Coupling the GTP hydrolysis-driven changes in the elongation factors to transitions between different states of the ribosome speeds up protein synthesis enormously. The cycles of elongation factor association, GTP hydrolysis, and dissociation also ensure that all such changes occur in the “forward” direction, helping translation to proceed efficiently (Figure 6–65).
In addition to moving translation forward, EF-Tu increases its accuracy. As we discussed in Chapter 3, EF-Tu can simultaneously bind GTP and aminoacyl-tR-NAs (see Figures 3–72 and 3–73), and it is in this form that the initial codon–anticodon interaction occurs in the A site of the ribosome. Because of the free-energy change associated with base-pair formation, a correct codon–anticodon match will bind more tightly than an incorrect interaction. However, this difference in affinity is relatively modest and cannot by itself account for the high accuracy of translation.