Human eukaryotic initiation factor 2 (eIF2)-GTP-Met-tRNAi ternary complex and eIF3 stabilize the 43 S preinitiation...

Area of Science:

• Molecular Biology
• Protein Interactions
• Translation Initiation

Background:

• Successful mRNA recruitment requires a stable 43S preinitiation complex (PIC).
• The precise roles of eIF1, eIF1A, eIF3, and the ternary complex (TC) in stabilizing the 43S PIC are not fully understood.

Purpose of the Study:

• To elucidate the complex network of interactions governing the stability of the human 43S PIC.
• To quantify the affinities of eIF1, eIF1A, and eIF3j within the 43S PIC using reconstituted components.

Main Methods:

• Reconstitution of the human 43S PIC in vitro.
• Systematic measurement of protein affinities using fluorescence anisotropy.

Main Results:

• All 43S PIC components exhibit high-affinity binding to the 40S ribosomal subunit.
• Human eIF1 and eIF1A demonstrate cooperative binding to the 40S subunit, an evolutionarily conserved interaction.
• Negative cooperativity was observed between eIF3j binding and the binding of eIF1, eIF1A, and TC; however, eIF3 significantly enhances eIF1 and eIF3j affinity, while TC indirectly boosts eIF1A affinity.

Conclusions:

• A comprehensive thermodynamic framework for the human 43S PIC has been established.
• Key interactions between 43S PIC components are identified, highlighting their importance in maintaining complex stability for efficient translation initiation.