Elongation factor G initiates translocation through a power stroke

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Prokaryotic protein synthesis involves elongation factor G (EF-G) and GTP hydrolysis for ribosome translocation.
• A long-standing debate questions whether EF-G's GTPase energy drives tRNA/mRNA movement or EF-G dissociation.
• Previous studies visualized EF-G on the ribosome using inhibitors, but dynamics during normal translocation remained unclear.

Purpose of the Study:

• To investigate the structural dynamics and rotational motions of EF-G domains during normal, uninhibited ribosomal translocation.
• To elucidate the role of EF-G's GTPase activity in ribosome unlocking and dissociation.

Main Methods:

• Single-molecule polarized total internal reflection fluorescence (polTIRF) microscopy was employed.
• Rotational motions of EF-G domains were detected during translocation in real-time.

Main Results:

• EF-G undergoes a small global rotation (~10°) relative to the ribosome post-GTP hydrolysis, unlocking the ribosome.
• A larger rotational motion occurs within EF-G's domain III prior to its dissociation from the ribosome.
• These rotations are crucial for facilitating tRNA and mRNA movement.

Conclusions:

• EF-G's GTPase energy drives ribosome unlocking through specific domain rotations.
• The study reveals the dynamic mechanism of EF-G during normal translocation, clarifying its role in protein synthesis.