The bacterial translocon SecYEG opens upon ribosome binding

Area of Science:

• Molecular biology
• Protein translocation
• Membrane protein complexes

Background:

• Co-translational translocation involves ribosome and SecYEG complex alignment.
• A signal is needed to displace the SecYEG sealing plug for nascent chain entry.
• The mechanism of SecYEG channel opening remains unidentified.

Purpose of the Study:

• To investigate the trigger for SecYEG channel opening.
• To characterize the functional properties of the SecYEG channel upon ribosome binding.
• To understand the conformational states of SecYEG during protein translocation.

Main Methods:

• Purification and reconstitution of the SecYEG channel.
• Planar lipid bilayer reconstitution assays.
• Ion conductivity measurements.
• Fluorescence correlation spectroscopy (FCS) for proteoliposome analysis.

Main Results:

• Ribosome binding induces a conformational transition in the SecYEG channel.
• The SecYEG channel opens to form a large ion-conducting pore upon ribosome interaction.
• The ion conductivity matches that of a plug deletion mutant.
• The open probability of the SecYEG channel is close to unity when bound to a ribosome.
• A closed post-translational conformation is proposed to prevent proton leak.

Conclusions:

• Ribosome binding is the trigger for SecYEG channel opening during co-translational translocation.
• The SecYEG channel exists in a highly conductive open state when interacting with ribosomes.
• A closed, post-translational conformation is critical for cellular viability by preventing proton leakage.