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 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 stoichiometirc analysis of 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 cAMP signaling and internalization upon agonist stimulation.