G protein-coupled receptors (GPCR)
There are different families of cellular receptors, and the largest one is the family of G protein-coupled receptors (GPCR). GPCRs are involved in many cellular functions and diseases, such as cancer or neurological disorders. Signal transduction by a GPCR begins with an inactive G protein coupled to the receptor. Once the GPCR recognizes a ligand, the conformation of the receptor changes to activate the G proteins, causing them to dissociate from the other receptor and to associate with effector proteins. The interaction of G proteins with the receptor or with effector proteins can be monitored using BRET. A commercial assay which improves the sensitivity and reliability of BRET is NanoBRET™.
GPCRs activate 2 main signal transduction pathways: the cAMP pathway and the phosphatidylinositol pathway. A valuable tool for the study of each pathway is the quantification of substances produced by the pathway. HTRF® assays are available for the quantification of cAMP, the second messenger of the cAMP pathway, and inositol monophosphate, a stable metabolite of inositol triphosphate produced by the phosphatidylinositol pathway.
Reporter gene assays
One important cellular response is the regulation of gene expression, but the expression of many genes is difficult to measure because they don’t have a visible effect that can be quantified. An excellent tool to study the regulation of gene expression is the use of reporter genes. The Dual-Luciferase Reporter® system (DLR™) is a very popular reporter gene system which can be optimally measured in any of our microplate readers.
Protein kinase activity
As mentioned above, other important cellular responses are the regulation of protein-protein interactions and of enzymatic activity. Both can be regulated by phosphorylation of proteins, which is performed by protein kinases. The activity of protein kinases can be monitored by measuring the concentration of ADP in a solution containing a protein kinase of interest. The Transcreener® ADP2 FI assay offers a very convenient, fast, and reliable way to quantify ADP.
Dual-Luciferase Reporter™, DLR® and NanoBRET™ are trademarks or Promega Corporation.
HTRF® is a registered trademark of Cisbio Bioassays.
Transcreener® is a registered trademark of Bellbrook Labs.