Initiation factor 2 stabilizes the ribosome in a semirotated conformation

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• The initiation phase of protein synthesis is critical for gene expression control.
• Conformational changes of the ribosome during initiation are poorly understood compared to elongation.
• Bacterial initiation factors (IFs) 1, 2, and 3 are essential for forming the initiation complex.

Purpose of the Study:

• To investigate ribosome conformational rearrangements during subunit joining in translation initiation.
• To monitor intersubunit rotation and L1 stalk movement using single-molecule Förster Resonance Energy Transfer (smFRET).
• To elucidate the role of initiation factors and GTP hydrolysis in these conformational changes.

Main Methods:

• Single-molecule Förster Resonance Energy Transfer (smFRET) was employed.
• Monitoring of intersubunit rotation and L1 stalk movement in real-time.
• Analysis of ribosome conformations during subunit association.

Main Results:

• Ribosome subunit association results in a semirotated conformation with a half-closed L1 stalk.
• Formation of this intermediate requires fMet-tRNA(fMet) and GTP-bound IF2.
• GTP hydrolysis by IF2 leads to L1 stalk opening and a nonrotated conformation.

Conclusions:

• A semirotated intermediate facilitates ribosome subunit association during initiation.
• L1 stalk movement is coupled to intersubunit rotation and/or IF2 binding.
• This study provides insights into the dynamic mechanism of translation initiation.