A single amino acid change of translation termination factor eRF1 switches between bipotent and omnipotent stop-codon specificity.
In eukaryotes a single class-1 translation termination factor eRF1 decodes the three stop codons: UAA, UAG and UGA. Some ciliates, like Euplotes, have a variant code, and here eRF1s exhibit UAR-only specificity, whereas UGA is reassigned as a sense codon. Since eukaryote eRF1 stop-codon recognition is associated with its N-terminal domain, structural features should exist in the N domain of ciliate eRF1s that restrict their stop-codon specificity. Using an in vitro reconstituted eukaryotic translation system we demonstrate here that a chimeric eRF1 composed of the N domain of Euplotes aediculatus eRF1 fused to the MC domains of human eRF1 exhibits UAR-only specificity. Functional analysis of eRF1 chimeras constructed by swapping Euplotes N domain sequences with the cognate regions from human eRF1 as well as site-directed mutagenesis of human eRF1 highlighted the crucial role of the alanine residue in position 70 of E. aediculatus eRF1 in restricting UGA decoding. Switching the UAR-only specificity of E. aediculatus eRF1 to omnipotent mode is due to a single point mutation. Furthermore, we examined the influence of eRF3 on the ability of chimeric and mutant eRF1s to induce peptide release in response to different stop codons.
Eliseev
Boris
B
Engelhardt Institute of Molecular Biology, the Russian Academy of Sciences, Moscow, Russia.
Kryuchkova
Polina
P
Alkalaeva
Elena
E
Frolova
Ludmila
L
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D018388:Codon, Terminator; D016812:Euplotes / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D010443:Peptide Chain Termination, Translational; D010454:Peptide Termination Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment
D018388:Codon, Terminator; C489002:ETF1 protein, human; D010454:Peptide Termination Factors; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C087971:peptide-chain-release factor 3