Competitive Binding Assay for the G-Protein-Coupled Receptor 30 (GPR30) or G-Protein-Coupled Estrogen Receptor (GPER)
The role of 2-methoxyestradiol is becoming a major area of investigation because of its therapeutic utility, though its mechanism is not fully explored. Recent studies have identified the G-protein-coupled receptor 30 (GPR30, GPER) as a high-affinity membrane receptor for 2-methoxyestradiol. However, studies aimed at establishing the binding affinities of steroid compounds for specific targets are difficult, as the tracers are highly lipophilic and often result in nonspecific binding in lipid-rich membrane preparations with low-level target receptor expression. 2-Methoxyestradiol binding studies are essential to elucidate the underlying effects of this novel estrogen metabolite and to validate its targets; therefore, this competitive receptor-binding assay protocol was developed in order to assess the membrane receptor binding and affinity of 2-methyoxyestradiol.
Key words2-Methoxyestradiol GPR30 G-protein-coupled receptor 30 GPER G-protein-coupled estrogen receptor Endoplasmic reticulum membrane Radioligand binding Competitive binding
The authors would like to acknowledge the laboratory personnel responsible for optimization of this protocol; in particular, the authors wish to acknowledge the graduate student Siva Koganti and fellow Upendra Gumaste. Studies were funded by NIH Grant #DK072140.
- 6.Sakamoto H, Matsuda K, Hosokawa K, Nishi M, Morris JF, Prossnitz ER, Kawata M (2007) Expression of G protein-coupled receptor-30, a G protein-coupled membrane estrogen receptor, in oxytocin neurons of the rat paraventricular and supraoptic nuclei. Endocrinology 148(12):5842–5850CrossRefGoogle Scholar