The C-Terminal Domain of Eukaryotic Initiation Factor 5 Promotes Start Codon Recognition by Its Dynamic Interplay with eIF1 and eIF2β
Rafael E. Luna,
Haribabu Arthanari,
Hiroyuki Hiraishi,
Jagpreet Nanda,
Pilar Martin-Marcos,
Michelle A. Markus,
Barak Akabayov,
Alexander G. Milbradt,
Lunet E. Luna,
Hee-Chan Seo,
Sven G. Hyberts,
Amr Fahmy,
Mikhail Reibarkh,
David Miles,
Patrick R. Hagner,
Elizabeth M. O'Day,
Tingfang Yi,
Assen Marintchev,
Alan G. Hinnebusch,
Jon R. Lorsch,
Katsura Asano,
Gerhard Wagner
Affiliations
Rafael E. Luna
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Haribabu Arthanari
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Hiroyuki Hiraishi
Molecular, Cellular and Developmental Biology Program, Division of Biology, Kansas State University, Manhattan, KS 66506, USA
Jagpreet Nanda
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
Pilar Martin-Marcos
Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA
Michelle A. Markus
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Barak Akabayov
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Alexander G. Milbradt
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Lunet E. Luna
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Hee-Chan Seo
Department of Molecular Biology, University of Bergen, Bergen 5020, Norway
Sven G. Hyberts
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Amr Fahmy
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Mikhail Reibarkh
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
David Miles
Molecular, Cellular and Developmental Biology Program, Division of Biology, Kansas State University, Manhattan, KS 66506, USA
Patrick R. Hagner
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Elizabeth M. O'Day
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Tingfang Yi
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Assen Marintchev
Department of Physiology and Biophysics, Boston University School of Medicine, Boston, MA 02118, USA
Alan G. Hinnebusch
Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA
Jon R. Lorsch
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
Katsura Asano
Molecular, Cellular and Developmental Biology Program, Division of Biology, Kansas State University, Manhattan, KS 66506, USA
Gerhard Wagner
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA
Recognition of the proper start codon on mRNAs is essential for protein synthesis, which requires scanning and involves eukaryotic initiation factors (eIFs) eIF1, eIF1A, eIF2, and eIF5. The carboxyl terminal domain (CTD) of eIF5 stimulates 43S preinitiation complex (PIC) assembly; however, its precise role in scanning and start codon selection has remained unknown. Using nuclear magnetic resonance (NMR) spectroscopy, we identified the binding sites of eIF1 and eIF2β on eIF5-CTD and found that they partially overlapped. Mutating select eIF5 residues in the common interface specifically disrupts interaction with both factors. Genetic and biochemical evidence indicates that these eIF5-CTD mutations impair start codon recognition and impede eIF1 release from the PIC by abrogating eIF5-CTD binding to eIF2β. This study provides mechanistic insight into the role of eIF5-CTD's dynamic interplay with eIF1 and eIF2β in switching PICs from an open to a closed state at start codons.
