Exploring the folding landscape of leptin: Insights into threading pathways.

Area of Science:

• Protein folding and biophysics
• Computational biology and structural bioinformatics

Background:

• Protein topology, including entanglements like knots and piercings, significantly impacts protein folding, stability, and function.
• Understanding the initial protein contacts driving folding in entangled topologies is crucial but remains challenging.
• Pierced Lasso Topology (PLT) proteins feature a covalent loop through which the backbone threads, offering a simpler model than true knots.

Purpose of the Study:

• To visualize and differentiate folding pathways, specifically slipknotting versus plugging, in the PLT protein leptin.
• To identify the critical native contacts that dictate the threading mechanism in entangled proteins.
• To provide insights into protein folding dynamics that are not observable through in vitro experiments.

Main Methods:

• Utilized the Energy Landscape Visualization Method (ELViM), a multidimensional projection technique, for in silico analysis.
• Visualized early threaded conformations of leptin to distinguish folding pathways.
• Identified key residue contacts governing the slipknotting and plugging transition pathways.

Main Results:

• ELViM successfully visualized and differentiated slipknotting and plugging pathways in leptin.
• Critical contacts were identified, revealing that hairpin loop formation in the unfolded state can inhibit the dominant slipknotting pathway.
• Distinct folding pathways associated with slipknotting and plugging were elucidated.

Conclusions:

• The study provides novel insights into the folding mechanisms of entangled proteins, specifically PLTs.
• ELViM serves as a powerful tool for studying in silico protein threading events with residue-specific detail.
• Findings can inform de novo protein design and guide future in vitro experiments.