Arrest Peptide Profiling resolves co-translational folding pathways and chaperone interactions in vivo

Area of Science:

• Molecular biology
• Biochemistry
• Cell biology

Background:

• Cytosolic proteins initiate folding co-translationally as they emerge from ribosomes.
• This early folding is critical for protein structure and function, guided by molecular chaperones.
• Detecting and understanding co-translational folding in real-time remains a significant challenge.

Purpose of the Study:

• To quantitatively analyze co-translational protein folding dynamics in live cells.
• To elucidate how protein topology influences folding pathways.
• To investigate the role of nascent chain-binding chaperones in protein folding.

Main Methods:

• Development of a high-throughput method: Arrest Peptide Profiling (AP Profiling).
• Integration of AP Profiling with single-molecule experiments.
• Analysis of GTPase domains to understand folding pathway determinants.

Main Results:

• AP Profiling successfully defined co-translational folding for GTPase domains.
• Protein topology was shown to shape folding pathways.
• Ablation of specific chaperones revealed distinct, localized folding changes, explaining chaperone functional redundancy.

Conclusions:

• This study provides unprecedented insight into cellular folding pathways of complex proteins.
• AP Profiling enables systematic studies of nascent protein folding with high resolution and throughput.
• The findings advance our understanding of protein biogenesis and quality control in cells.