The following is a summary of “Glycoprotein-glycoprotein Receptor Binding Detection Using Bioluminescence Resonance Energy Transfer,” published in the June 2024 issue of Endocrinology by Adamczuk, et al.

Glycoprotein receptors, members of the expansive G protein-coupled receptor family, play a critical role in binding glycoprotein hormones through their large extracellular domains. Traditional methods for studying hormone-receptor interactions, such as ligand-binding assays using radiolabeling or thermal shift assays, have limitations in capturing receptor localization and activity comprehensively. For a study, researchers sought to explore bioluminescence resonance energy transfer (BRET) as a method to analyze these interactions in live cells, focusing on the Gaussia luciferase (Gluc)-luteinizing hormone/chorionic gonadotropin receptor (LHCGR) fusion and its ligands, human chorionic gonadotropin (hCG) or LH, fused with enhanced green fluorescent protein (eGFP).

They expressed Gluc-LHCGR and other Gluc-tagged G protein-coupled receptors, including somatostatin and C-X-C motif chemokine receptors, on the plasma membrane of living cells. They utilized BRET to monitor luminescent activity, correlating it with membrane receptor expression to confirm full receptor functionality. The chimeric eGFP-ligands were secreted from cells and evaluated for their ability to bind and activate both wild-type LHCGR and Gluc-LHCGR variants.

The findings demonstrated that Gluc-LHCGR and other Gluc-tagged receptors are effectively expressed on the plasma membrane, where their luminescent activity mirrors receptor expression levels. Chimeric eGFP-ligands were efficiently secreted and demonstrated robust binding and activation of both wild-type LHCGR and Gluc-LHCGR. Notably, BRET facilitated the study of interactions involving clinically relevant mutations in hormones and LHCGR variants, showcasing its efficiency as a rapid, safe, and cost-effective bioassay.

The study underscored the utility of BRET as a valuable tool for investigating hormone-receptor interactions in real time within living cells. By overcoming the limitations of traditional assays, BRET enabled comprehensive insights into receptor function and response to mutations, thereby advancing the understanding of glycoprotein receptor biology. Future applications of the method may further elucidate complex signaling mechanisms and aid in developing targeted therapeutic strategies.

Reference: academic.oup.com/endo/article-abstract/165/6/bqae052/7659435