Yeast-Based Biosensors As a Low-Cost Healthcare Diagnostic

This talk will introduce Yeast-based biosensors (YBBs), a low cost healthcare diagnostic, based on directed evolution of G-protein coupled receptors.  Access to low-cost, point-of-care (POC) diagnostics enables screening, diagnosis and treatment monitoring for a number of critical diseases. The use of POC diagnostics reduces hospital stays, increases adherence to treatment, and has wider economic benefits compared to laboratory testing. To date, POC diagnostics, such as lateral flow sandwich immunoassays, nucleic acid amplification tests, and colorimetric chemical reactions, are available for proteins, DNA/RNA, and small molecule disease biomarkers. However, there is a technology gap for assaying peptides and small proteins biomarkers less than 15 kDa, which include many biomarkers that are degradation products of a larger protein. These smaller biomarkers are very appealing as they often appear in easy-to-access specimens (i.e. saliva, urine, feces, and blood) suitable for POC diagnostics

Yeast based biosensors (YBBs) could fill this gap, as active-dry yeast is cheap and does not require a cold-chain and the G-protein coupled receptor (GPCR) Ste2p naturally detects a unique peptide a-factor. We have used a directed evolution strategy to retarget Ste2p from detecting a-factor to detecting a biomarker for renal failure.  Key to accomplishing this was employing a substrate walking approach using intermediate chimeric peptides with increasing similarity to the target sequence.  The permissible step size or peptide ligand dissimilarity was determined by screening Ste2p mutants against peptides of varying dissimilarity.  Using this step size, we designed and evolved receptors for a series of peptides walking toward a biomarker for chronic kidney disease.  We have  demonstrated the clinical utility of YBBs by showing specific activation in human urine by the cystatin C peptide. YBBs could serve as a useful diagnostic in resource-limited settings, as well as have useful applications in bioterrorism detection, drug discovery, and environmental monitoring.