PANCS-spec-Binders: A system for rapidly discovering isoform- or epitope-specific binders

Area of Science:

• Molecular Biology
• Protein Engineering
• Biotechnology

Background:

• Proteins that bind target proteins, known as binders, are crucial for biological research and therapeutics.
• Current methods for discovering binders often lack the specificity required, leading to promiscuous binding with related proteins or undesired interactions.
• Existing binder discovery platforms, like phage-assisted non-continuous selection of binders (PANCS-Binders), primarily enrich for affinity, making it difficult to identify binders with desired specificity profiles.

Purpose of the Study:

• To develop a novel method for discovering protein binders with controlled specificity.
• To enable the rapid identification of binders that exhibit high selectivity between related proteins (isoforms) or target specific epitopes.
• To address the limitations of existing methods that often require extensive secondary screening and frequently result in failure to achieve desired specificity.

Main Methods:

• Development of PANCS-spec-Binders, a platform incorporating simultaneous selection and counterselection for controlled binder specificity.
• Application of PANCS-spec-Binders for the discovery of isoform-selective binders targeting HRAS over KRAS.
• Utilizing PANCS-spec-Binders to identify epitope-specific binders for LC3B, including those that avoid a specific interaction region.

Main Results:

• PANCS-spec-Binders successfully enabled the discovery of isoform-selective binders with >100-fold higher affinity for HRAS compared to KRAS.
• The platform facilitated the identification of epitope-specific binders, demonstrating control over binding to specific regions of a target protein.
• The method proved effective in identifying binders that specifically avoid a defined epitope, showcasing precise control over binding interactions.

Conclusions:

• PANCS-spec-Binders represents a significant advancement in binder discovery, enabling rapid identification of reagents with defined specificity.
• The platform addresses critical needs in both selectivity (isoform-specific) and epitope-specific binding, crucial for research and therapeutic applications.
• This method overcomes the limitations of traditional approaches, offering a high-fidelity solution for discovering specific binders within days.