Luna, Rafael E.Arthanari, HaribabuHiraishi, HiroyukiAkabayov, BarakTang, LeimingCox, ChristianMarkus, Michelle A.Luna, Lunet E.Ikeda, YukaWatanabe, RyosukeBedoya, EdwardYu, CathyAlikhan, ShumsWagner, GerhardAsano, Katsura2014-05-062014-05-062014-05-06http://hdl.handle.net/2097/17685Scanning of the mRNA transcript by the preinitiation complex (PIC) requires a panel of eukaryotic initiation factors including eIF1 and eIF1A, the main transducers of stringent AUG selection. eIF1A plays an important role in start codon recognition; however, its molecular contacts with eIF5 are unknown. Using NMR, we unveil eIF1A’s binding surface on the carboxyl-terminal domain of eIF5 (eIF5-CTD). We validated this interaction by observing that eIF1A does not bind to an eIF5-CTD mutant, altering the revealed eIF1A-interaction site. We also found that the interaction between eIF1A:eIF5-CTD is conserved between human and yeast. Using GST pull down assays of purified proteins, we showed that the N-terminal tail (NTT) of eIF1A mediates the interaction with eIF5-CTD and eIF1. Genetic evidence indicates that overexpressing eIF1 or eIF5 suppresses the slow growth phenotype of eIF1A-NTT mutants. These results suggest that the eIF1A:eIF5-CTD interaction during scanning PICs contributes to the maintenance of eIF1 within the open PIC.en-USPermission to archive granted by American Chemical Society, March 20, 2014. This document is the unedited Author's version of a Submitted Work that was subsequently accepted for publication in Biochemistry, copyright © American Chemical Society after peer review. To access the final edited and published work see http://pubs.acs.org/doi/full/10.1021/bi4009775Preinitiation complexEukaryotic initiation factorseIF1eIF1AeIF5Article (author version)