Translation Initiation Factor 2γ Mutant Alters Start Codon Selection Independent of Met-tRNA Binding

Abstract

ABSTRACT Selection of the AUG start codon for translation in eukaryotes is governed by codon-anticodon interactions between the initiator Met-tRNA i Met and the mRNA. Translation initiation factor 2 (eIF2) binds Met-tRNA i Met to the 40S ribosomal subunit, and previous studies identified Sui − mutations in eIF2 that enhanced initiation from a noncanonical UUG codon, presumably by impairing Met-tRNA i Met binding. Consistently, an eIF2γ-N135D GTP-binding domain mutation impairs Met-tRNA i Met binding and causes a Sui − phenotype. Intragenic A208V and A382V suppressor mutations restore Met-tRNA i Met binding affinity and cell growth; however, only A208V suppresses the Sui − phenotype associated with the eIF2γ-N135D mutation. An eIF2γ-A219T mutation impairs Met-tRNA i Met binding but unexpectedly enhances the fidelity of initiation, suppressing the Sui − phenotype associated with the eIF2γ-N135D,A382V mutant. Overexpression of eIF1, which is thought to monitor codon-anticodon interactions during translation initiation, likewise suppresses the Sui − phenotype of the eIF2γ mutants. We propose that structural alterations in eIF2γ subtly alter the conformation of Met-tRNA i Met on the 40S subunit and thereby affect the fidelity of start codon recognition independent of Met-tRNA i Met binding affinity.