Structure analysis of multi-factor complex of eukaryotic translation initiation factor

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For all living organisms, the regulation of gene expression is an important control mechanism in their biological processes such as environmental adaptation, development, and differentiation. The translation is the final step in the flow of the gene expression, and it can be divided into three distinct phases : initiation, elongation and termination. Initiation is most complex of these phases and involves greatest number of regulatory processes that all performed by more than 30 factors (eIFs) and ribosome. The regulation at this phase allows for immediate and rapid response to cellular stress (starvation, heat shock, virus infection) or apoptosis, and controls the cellular activity level in their biological processes such as memory, development, and differentiation. The fundamental processes in translation initiation are the assembling components of eIF1, eIF2-GTP, eIF3, eIF5, and Met-tRNA^i to a multi-factor complex (MFC), binding MFC to ribosomal small subunit (43S) and then binding … More 5'end of mRNA to 43S, and scanning mRNA to recognize the start codon on the resulting 48S complex. In this study, the functional and structural analysis of MFC mainly on eIF2 was focused. Especially, conformation changes and interaction of the components during assemble were analyzed for understanding the reaction mechanism and regulation of translation initiation. Last year, we have solved the structures of a/eIF2βγ and a/eIF2βγ-GDP complex. Based on the obtained structures the functional analysis of a/eIF2βγ was carried out this year.Considering the structural and functional homology between a/eIF2γ and translation elongation factor EF-Tu, switch1 motif of a/eIF2γ may play an important role in binding and releasing Met-tRNA^i similarly to that of EF-Tu. In this study, the obtained structure of a/eIF2βγ-GDP complex showed that a/eIF2β was interacted with switch1 motif of a/eIF2βγ and consequently switch1 motif was positioned in the distant from GDP-binding site. Moreover, conformation changes of switch1 region between the structures of a/eIF2βγ and a/eIF2βγ-GDP, and a/eIF2βγ and a/eIF2γ were found. Therefore, we mutated conserved residues of switch1 motif, and inspected the effect of these mutations on Met-tRNA^i binding. The paper was in submitted. Less