Mutations in domain IV of elongation factor EF-G confer-1 frameshifting

A recent crystal structure of a ribosome complex undergoing partial translocation in the absence of elongation factor EF-G showed disruption of codon-anticodon pairing and slippage of the reading frame by -1, directly implicating EF-G in preservation of the translational reading frame. Among mutations identified in a random screen for dominant-lethal mutations of EF-G were a cluster of six that map to the tip of domain IV, which has been shown to contact the codon-anticodon duplex in trapped translocation intermediates. In vitro synthesis of a full-length protein using these mutant EF-Gs revealed dramatically increased -1 frameshifting, providing new evidence for a role for domain IV of EF-G in maintaining the reading frame. These mutations also caused decreased rates of mRNA translocation and rotational movement of the head and body domains of the 30S ribosomal subunit during translocation. Our results are in general agreement with recent findings from Rodnina and coworkers based on in vitro translation of an oligopeptide using EF-Gs containing mutations at two positions in domain IV, who found an inverse correlation between the degree of frameshifting and rates of translocation. Four of our six mutations are substitutions at positions that interact with the translocating tRNA, in each case contacting the RNA backbone of the anticodon loop. We suggest that EF-G helps to preserve the translational reading frame by preventing uncoupled movement of the tRNA through these contacts; a further possibility is that these interactions may stabilize a conformation of the anticodon that favors base-pairing with its codon.

Automated summary

Recent studies have shown that the ribosome complex undergoes partial translocation without elongation factor EF-G can disrupt codon-anticodon pairing, leading to frame slippage, implicating EF-G in maintaining translational reading frame. Mutations in EF-G, particularly in domain IV, were found to increase -1 frameshifting during protein synthesis, slow down mRNA translocation rates, and inhibit rotational movement of the 30S ribosomal subunit. These interactions with the translocating tRNA backbone may help EF-G prevent uncoupled tRNA movement and stabilize the anticodon conformation for better base-pairing with the codon.