Binding of human Cdc123 to eIF2γ.

Eukaryotic initiation factor 2 (eIF2) plays a key role in protein synthesis and in its regulation. The assembly of this heterotrimeric factor is facilitated by Cdc123, a member of the ATP grasp family that binds the γ subunit of eIF2. Notably, some mutations related to MEHMO syndrome, an X-linked intellectual disability, affect Cdc123-mediated eIF2 assembly. The mechanism of action of Cdc123 is unclear and structural information for the human protein is awaited. Here, the crystallographic structure of human Cdc123 (Hs-Cdc123) bound to domain 3 of human eIF2γ (Hs-eIF2γD3) was determined. The structure shows that the domain 3 of eIF2γ is bound to domain 1 of Cdc123. In addition, the long C-terminal region of Hs-Cdc123 provides a link between the ATP and Hs-eIF2γD3 binding sites. A thermal shift assay shows that ATP is tightly bound to Cdc123 whereas the affinity of ADP is much smaller. Yeast cell viability experiments, western blot analysis and two-hybrid assays show that ATP is important for the function of Hs-Cdc123 in eIF2 assembly. These data and recent findings allow us to propose a refined model to explain the mechanism of action of Cdc123 in eIF2 assembly.