Screening of protein-based inhibitors for the intracellular domain of epidermal growth factor receptor by directed evolution using the yeast Gγ recruitment system.

Protein-based therapeutics, including antibodies and antibody-like-proteins, have increasingly attracted attention due to their high specificity compared to small-molecular drugs. The Gγ recruitment system, one of the in vivo yeast two-hybrid systems for detecting protein-protein interactions, has been previously developed using yeast signal transduction machinery. In this study, we modified the Gγ recruitment system to screen the protein mutants that efficiently bind to the intracellular domain of the epidermal growth factor receptor L858R mutant (cytoEGFR).

Droplet-based microfluidic platform for detecting agonistic peptides that are self-secreted by yeast expressing a G-protein-coupled receptor.

Background: Single-cell droplet microfluidics is an important platform for high-throughput analyses and screening because it provides an independent and compartmentalized microenvironment for reaction or cultivation by coencapsulating individual cells with various molecules in monodisperse microdroplets. In combination with microbial biosensors, this technology becomes a potent tool for the screening of mutant strains. In this study, we demonstrated that a genetically engineered yeast strain that can fluorescently sense agonist ligands via the heterologous expression of a human G-protein-coupled receptor (GPCR) and concurrently secrete candidate peptides is highly compatible with single-cell droplet microfluidic technology for the high-throughput screening of new agonistically active peptides.

Synthetic production of prenylated naringenins in yeast using promiscuous microbial prenyltransferases.

Reconstitution of prenylflavonoids using the flavonoid biosynthetic pathway and prenyltransferases (PTs) in microbes can be a promising attractive alternative to plant-based production or chemical synthesis. Here, we demonstrate that promiscuous microbial PTs can be a substitute for regiospecific but mostly unidentified botanical PTs. To test the prenylations of naringenin, we constructed a yeast strain capable of producing naringenin from l-phenylalanine by genomic integration of six exogenous genes encoding components of the naringenin biosynthetic pathway.

Comparative analyses of site-directed mutagenesis of human melatonin MTNR1A and MTNR1B receptors using a yeast fluorescent biosensor.

Melatonin is an indoleamine neurohormone made by the pineal gland. Its receptors, MTNR1A and MTNR1B, are members of the G-protein-coupled receptor (GPCR) family and are involved in sleep, circadian rhythm, and mood disorders, and in the inhibition of cancer growth. These receptors, therefore, represent significant molecular targets for insomnia, circadian sleep disorders, and cancer.