Stoichiometric Analysis of Oligomerization of Membrane Proteins on Living Cells Using Coiled-Coil Labeling and Spectral Imaging

Many membrane proteins are proposed to work as oligomers; however, the conclusion is sometimes controversial, as for β₂-adorenergic receptor (β₂AR), which is one of the best-studied family A G-protein-coupled receptors. This is due to the lack of methods for easy and precise detection of the oligomeric state of membrane proteins on living cells. Here, we show that a combination of the coiled-coil tag–probe labeling method and spectral imaging enable a stoichiometric analysis of the oligomeric state of membrane proteins on living cells using monomeric, dimeric, and tetrameric standard membrane proteins. Using this method, we found that β₂ARs do not form constitutive homooligomers, while they exhibit their functions such as the cyclic adenosine 5'-monophosphate (cAMP) signaling and internalization upon agonist stimulation