A High-Throughput Method for Screening Peptide Activators of G-Protein-Coupled Receptors

Area of Science:

• Biochemistry and Molecular Biology
• Pharmacology
• Cell Biology

Background:

• G-protein-coupled receptors (GPCRs) are crucial drug targets, but identifying novel peptide activators remains challenging.
• Existing screening methods often lack efficiency and specificity for discovering peptide modulators.
• Protein-protein interaction (PPI) based approaches offer a promising avenue for GPCR drug discovery.

Purpose of the Study:

• To develop and validate an innovative, high-throughput screening method for identifying peptide activators of GPCRs.
• To discover novel peptide activators targeting the glucagon-like peptide-1 receptor (GLP-1R).
• To explore the potential of these identified peptides in structure-function and drug discovery research.

Main Methods:

• Designed a large library (92,918) of peptides fused with transmembrane domains of glycosylphosphatidylinositol-anchored proteins (GPI-APs).
• Utilized a pooled lentiviral system for efficient cellular membrane expression of the GPI-AP-peptide fusions.
• Employed fluorescence-activated cell sorting (FACS) for high-throughput screening and identification of GPCR-activating peptides.

Main Results:

• Successfully screened the GPI-AP-peptide library to identify novel activators of the glucagon-like peptide-1 receptor (GLP-1R).
• Discovered a peptide, PepA3, derived from the Frizzled-like (FZ) domain of human Carboxypeptidase Z (CPZ).
• PepA3 and its variants (PepA, PepA2) demonstrated comparable efficacy but lower potency to GLP-1 in activating GLP-1R, suggesting distinct binding mechanisms.

Conclusions:

• The developed PPI-based screening technology is efficient for identifying novel GPCR-activating peptides.
• The identified peptides, such as PepA3, represent potential leads for GLP-1R targeted therapeutics.
• This platform facilitates GPCR research, enabling structure-function studies and drug discovery efforts.