title: Screening of randomly mutagenized glucagon-like peptide-1 library by using an integrated yeast-mammalian assay system.
abstract: Glucagon-like peptide-1 (GLP1) is a 30-amino acid peptide hormone activating the GLP1 receptor (GLP1R), a class B G-protein coupled receptor (GPCR), and is considered to be effective for treating diabetes and other metabolic diseases. Phage display is the first innovative technology in order to prepare and screen a large polypeptide library including GLP1R agonists, but this methodology is not as effective in discovering functional peptides such as activators for GPCRs. Here, we report a novel functional screening system for GPCR-acting peptides, which integrates a yeast peptide secretion system into a biological detection system with GPCR-producing mammalian cells. Using this screening system, we found attractive GLP1R agonists with several substitutions from a random mutant GLP1 library which was secreted by yeast, Saccharomyces cerevisiae. This system established here not only enables peptides to be analyzed in the soluble form but also needs no chemical synthesis, purification, and condensation of peptides of interests, and therefore, can be widely applied to the discovery of novel bioactive peptides acting on GPCRs.
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Shigemori
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
authors.forename: Kouichi
authors.initials: K
authors.lastname: Kuroda
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan. Electronic address: miueda@kais.kyoto-u.ac.jp.
authors.forename: Mitsuyoshi
authors.initials: M
authors.lastname: Ueda
mesh_terms: D000818:Animals; D015723:Gene Library; D052216:Glucagon-Like Peptide 1 / Q000235:genetics*; D057166:High-Throughput Screening Assays / Q000379:methods; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D000097789:Glucagon-Like Peptide-1 Receptor Agonists
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D052216:Glucagon-Like Peptide 1; D000097789:Glucagon-Like Peptide-1 Receptor Agonists
keywords: Bioactive peptides; CRE; DMEM; Dulbecco’s modified Eagle Medium; Ex4; Functional screening; G-protein coupled receptor; G-protein coupled receptors; GLP1; GPCR; Glucagon-like peptide-1; Mammalian cells; Yeast; cyclic AMP-responsive element; exendin-4; glucagon-like peptide-1