A Multiplex GPCR-Mediated Peptide Tagging System for a Growing Yeast Synthetic Biology Toolbox

Straightforward methods for specifically detecting and quantifying proteins are essential for both basic and applied research and notably in synthetic biology. Previously we demonstrated that the yeast mating pathway could be hijacked to detect species-specific fungal peptide pheromones using their corresponding mating GPCRs. Here we asked if our yeast biosensor could detect proteins in addition to peptides - a question not previously resolved in the literature. As such, we repurposed the Saccharomyces cerevisiae fungal mating pheromone alpha-factor as a peptide tag and fused it terminally and internally to the protein Smt3. Our biosensor was able to detect the tagged protein in the nanomolar range using fluorescence as a read-out. We extended the assay to four additional orthogonal peptide pheromone tags, demonstrating a cheap, non-labor-intensive, and high-throughput assay compatible with multiplexing for protein detection. With its ability to detect proteins our living yeast biosensor could be useful for the optimization of protein producing cell-factories, for building logic gates and myriad other applications in synthetic biology. image

Automated summary

This study developed a straightforward method for detecting proteins using a yeast biosensor. By repurposing fungal mating pheromones as peptide tags, the biosensor was able to detect proteins in a cost-effective and high-throughput manner. The biosensor has potential applications in protein-producing cell factories and synthetic biology.