Single fluorescent-molecule video imaging and tracking in living cells are revolutionizing our understanding of molecular interactions in the plasma membrane and intracellular membrane systems. They have revealed that molecular interactions occur surprisingly dynamically on much shorter time scales (≪1s) than those expected from the results by conventional techniques, such as pull-down assays (minutes to hours). Single-molecule imaging has unequivocally showed that G-protein-coupled receptors (GPCRs) undergo dynamic equilibrium between monomers and dimers, by enabling the determination of the 2D monomer-dimer equilibrium constant, the dimer dissociation rate constant (typically ∼10s(-1)), and the formation rate constant. Within one second, GPCRs typically undergo several cycles of monomer and homo-dimer formation with different partners.
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