High-throughput determination of protein affinities using unmodified peptide libraries in nanomolar scale

High-throughput determination of protein affinities using unmodified peptide libraries in nanomolar scale

Blog Article

Summary: Protein-protein interactions (PPIs) are of fundamental importance for our understanding of physiology and pathology.PPIs involving short, linear motifs play truvisionhealthftp.com a major role in immunological recognition, signaling, and regulation and provide attractive starting points for pharmaceutical intervention.Yet, state-of-the-art protein-peptide affinity determination approaches exhibit limited throughput and sensitivity, often resulting from ligand immobilization, labeling, or synthesis.

Here, we introduce a high-throughput method for in-solution analysis of protein-peptide interactions using a phenomenon called temperature related intensity change (TRIC).We use TRIC for the identification and fine-mapping of low- and high-affinity protein interaction sites and the definition of sequence binding requirements.Validation is achieved by microarray-based studies using wild-type and mutated recombinant protein and the native protein within tissue lysates.

On-chip neutralization and strong correlation with structural data establish TRIC here as a quasi-label-free method to determine binding affinities of unmodified peptide libraries with large dynamic range.