Binding of eIF3 in complex with eIF5 and eIF1 to the 40S ribosomal subunit is accompanied by dramatic structural...

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Eukaryotic initiation factor 3 (eIF3) is a large, essential multiprotein complex crucial for translation initiation.
• Its high flexibility has prevented high-resolution structural determination of free eIF3.
• Previous work established a 2D interaction map of yeast eIF3 subunits.

Purpose of the Study:

• To determine the 3D structure of yeast eIF3.
• To elucidate the conformational changes of eIF3 upon binding to the 40S ribosomal subunit.
• To map the position of eIF5 within the translation initiation complex.

Main Methods:

• In vitro reconstitution of yeast eIF3.
• Chemical cross-linking and trypsin digestion.
• Advanced mass spectrometry for 3D structure determination.
• Biochemical and genetic analyses.

Main Results:

• A high-resolution 3D structure of free yeast eIF3 was determined, revealing a tightly packed conformation.
• Cross-linking data supported the 2D interaction map and showed exposed WD40 and RRM domains.
• eIF3's compact structure is further stabilized by eIF1 and eIF5 binding.
• A model was proposed where eIF3 initially contacts the 40S subunit's solvent-exposed side, then opens to embrace it.
• The position of eIF5 was mapped to the region below the P- and E-sites of the 40S subunit.

Conclusions:

• Yeast eIF3 adopts a compact structure in its free state, with exposed functional domains.
• The binding of eIF3 to the 40S subunit likely involves a conformational transition from a compact to an open state.
• This conformational flexibility is critical for eIF3's role in coordinating translation initiation.
• The study provides new insights into the structural dynamics of translation initiation factors.