title: Two-step model of stop codon recognition by eukaryotic release factor eRF1.
abstract: Release factor eRF1 plays a key role in the termination of protein synthesis in eukaryotes. The eRF1 consists of three domains (N, M and C) that perform unique roles in termination. Previous studies of eRF1 point mutants and standard/variant code eRF1 chimeras unequivocally demonstrated a direct involvement of the highly conserved N-domain motifs (NIKS, YxCxxxF and GTx) in stop codon recognition. In the current study, we extend this work by investigating the role of the 41 invariant and conserved N-domain residues in stop codon decoding by human eRF1. Using a combination of the conservative and non-conservative amino acid substitutions, we measured the functional activity of >80 mutant eRF1s in an in vitro reconstituted eukaryotic translation system and selected 15 amino acid residues essential for recognition of different stop codon nucleotides. Furthermore, toe-print analyses provide evidence of a conformational rearrangement of ribosomal complexes that occurs during binding of eRF1 to messenger RNA and reflects stop codon decoding activity of eRF1. Based on our experimental data and molecular modelling of the N-domain at the ribosomal A site, we propose a two-step model of stop codon decoding in the eukaryotic ribosome.
authors: Engelhardt Institute of Molecular Biology, the Russian Academy of Sciences, 119991 Moscow, Russia.
authors: Polina
authors: P
authors: Kryuchkova
authors: Alexander
authors: A
authors: Grishin
authors: Boris
authors: B
authors: Eliseev
authors: Anna
authors: A
authors: Karyagina
authors: Ludmila
authors: L
authors: Frolova
authors: Elena
authors: E
authors: Alkalaeva
mesh_terms: D000595:Amino Acid Sequence; D018388:Codon, Terminator; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010443:Peptide Chain Termination, Translational; D010454:Peptide Termination Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012270:Ribosomes / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D018388:Codon, Terminator; C489002:ETF1 protein, human; D010454:Peptide Termination Factors; D010455:Peptides