A GPCR-based yeast biosensor for biomedical, biotechnological, and point-of-use cannabinoid determination

Abstract

Eukaryotic cells use G-protein coupled receptors to sense diverse signals, ranging from chemical compounds to light. Here, we exploit the remarkable sensing capacity of G-protein coupled receptors to construct yeast-based biosensors for real-life applications. To establish proof-of-concept, we focus on cannabinoids because of their neuromodulatory and immunomodulatory activities. We construct a CB2 receptor-based biosensor, optimize it to achieve high sensitivity and dynamic range, and prove its effectiveness in three applications of increasing difficulty. First, we screen a compound library to discover agonists and antagonists. Second, we analyze 54 plants to discover a new phytocannabinoid, dugesialactone. Finally, we develop a robust portable device, analyze body-fluid samples, and confidently detect designer drugs like JWH-018. These examples demonstrate the potential of yeast-based biosensors to enable diverse applications that can be implemented by non-specialists. Taking advantage of the extensive sensing repertoire of G-protein coupled receptors, this technology can be extended to detect numerous compounds.

Delta‐8 tetrahydrocannabinol: a scoping review and commentary

“…Chemically determining if THC is delta-8 or delta-9 is difficult, costly and time-consuming [2,5,17,27,45,47,[95][96][97][98][99][100][101][102][103]. Although in the preliminary stages of research, chromatography methods have been developed to distinguish between different manufacturing methods [89].…”

Olfactory Receptors as an Emerging Chemical Sensing Scaffold

“…A second fertile area has been the use of GPCR-based sensors in yeast for the development of point-of-care diagnostics, such as the detection of fungal pathogens, 37 and cannabinoids in artificial bodily fluids. 36 Finally, GPCR-based sensors have been used for metabolic engineering applications, including the quantification of microbially produced serotonin 38 and melatonin. 39 Recent technical improvements to GPCR-based sensors in yeast encompass the evaluation of different promoters and vector copy numbers to modulate the sensor response, 39 the biosynthesis of cholesterol to improve human GPCR trafficking, 40 and the introduction of CRISPR landing pads in the genome for rapid sensor construction.…”

“…Recently identified ligands using GPCR-based sensors in yeast. Information obtained from refs ( 33 − 36 ).…”

“…Recent examples include the discovery that antimicrobials activate serotonin receptor 4 in colon cells, leading to colon cell motility; the identification of kynurenic acid, a neuroprotection agent, as a ligand for the orphan GPCR HCAR3; the identification of a new histamine receptor 2 blocker scaffold, 8-hydroxyquinoline, that does not decompose into the carcinogen N -nitrosodimethylamine for the development of improved antiacids; and the discovery of new ligands for the cannabinoid receptor (Figure B). A second fertile area has been the use of GPCR-based sensors in yeast for the development of point-of-care diagnostics, such as the detection of fungal pathogens, and cannabinoids in artificial bodily fluids . Finally, GPCR-based sensors have been used for metabolic engineering applications, including the quantification of microbially produced serotonin and melatonin .…”

Automation of yeast spot assays using an affordable liquid handling robot

“…The Opentrons OT-2 (OT-2; Opentrons Labworks Inc., New York, USA) is an automated pipetting machine that is a low-cost and open-source hardware that is easy to customize by end-users [16]. The OT-2 is used for automating experimental procedures in yeast research, such as high-throughput screening [17], micro-cultivation experiments [18], DNA cloning [19], bioreactor assays [20], and proteomics [21]. If it is possible to automate yeast spot assays using OT-2, it is expected to be a solution with a low cost and few technical barriers that can be used in various laboratories, especially in academia.…”