title: Sliding of a 43S ribosomal complex from the recognized AUG codon triggered by a delay in eIF2-bound GTP hydrolysis.
abstract: During eukaryotic translation initiation, 43S ribosomal complex scans mRNA leader unless an AUG codon in an appropriate context is found. Establishing the stable codon-anticodon base-pairing traps the ribosome on the initiator codon and triggers structural rearrangements, which lead to Pi release from the eIF2-bound GTP. It is generally accepted that AUG recognition by the scanning 43S complex sets the final point in the process of start codon selection, while latter stages do not contribute to this process. Here we use translation reconstitution approach and kinetic toe-printing assay to show that after the 48S complex is formed on an AUG codon, in case GTP hydrolysis is impaired, the ribosomal subunit is capable to resume scanning and slides downstream to the next AUG. In contrast to leaky scanning, this sliding is not limited to AUGs in poor nucleotide contexts and occurs after a relatively long pause at the recognized AUG. Thus, recognition of an AUG per se does not inevitably lead to this codon being selected for initiation of protein synthesis. Instead, it is eIF5-induced GTP hydrolysis and Pi release that irreversibly trap the 48S complex, and this complex is further stabilized by eIF5B and 60S joining.
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow 119991, Russia.
authors.forename: Ilya M
authors.initials: IM
authors.lastname: Terenin
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia School of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow 119234, Russia.
authors.forename: Kseniya A
authors.initials: KA
authors.lastname: Akulich
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia.
authors.forename: Dmitry E
authors.initials: DE
authors.lastname: Andreev
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia Department of Molecular Biology, Biological Faculty, Lomonosov Moscow State University, Moscow 119234, Russia.
authors.forename: Sofya A
authors.initials: SA
authors.lastname: Polyanskaya
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia shatsky@genebee.msu.ru.
authors.forename: Ivan N
authors.initials: IN
authors.lastname: Shatsky
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119234, Russia Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow 119991, Russia Department of Biochemistry, Biological Faculty, Lomonosov Moscow State University, Moscow 119234, Russia dmitriev_sergey@genebee.msu.ru.
authors.forename: Sergey E
authors.identifier: http://orcid.org/0000-0002-1774-8475
authors.initials: SE
authors.lastname: Dmitriev
mesh_terms: D000926:Anticodon / Q000235:genetics; D003062:Codon / Q000235:genetics; D004926:Escherichia coli; D015852:Eukaryotic Initiation Factor-2 / Q000235:genetics* / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D010448:Peptide Initiation Factors / Q000235:genetics* / Q000378:metabolism; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics*; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics*; D000097574:Eukaryotic Translation Initiation Factor 5A
chemical_list: D000926:Anticodon; D003062:Codon; D015852:Eukaryotic Initiation Factor-2; D006160:Guanosine Triphosphate; D046912:Multiprotein Complexes; D010448:Peptide Initiation Factors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D000097574:Eukaryotic Translation Initiation Factor 5A