Abstract
Considerable evidence has accumulated in recent years suggesting that G protein-coupled receptors (GPCRs) associate in the plasma membrane to form homo- and/or heteromers. Nevertheless, the stoichiometry, fraction and lifetime of such receptor complexes in living cells remain topics of intense debate. Motivated by experimental data suggesting differing stabilities for homomers of the cognate human β1- and β2-adrenergic receptors, we have carried out approximately 160 microseconds of biased molecular dynamics simulations to calculate the dimerization free energy of crystal structure-based models of these receptors, interacting at two interfaces that have often been implicated in GPCR association under physiological conditions. Specifically, results are presented for simulations of coarse-grained (MARTINI-based) and atomistic representations of each receptor, in homodimeric configurations with either transmembrane helices TM1/H8 or TM4/3 at the interface, in an explicit lipid bilayer. Our results support a definite contribution to the relative stability of GPCR dimers from both interface sequence and configuration. We conclude that β1- and β2-adrenergic receptor homodimers with TM1/H8 at the interface are more stable than those involving TM4/3, and that this might be reconciled with experimental studies by considering a model of oligomerization in which more stable TM1 homodimers diffuse through the membrane, transiently interacting with other protomers at interfaces involving other TM helices.
Highlights
G Protein-Coupled Receptors (GPCRs) have been reported to associate in the cell membrane to form dimers/oligomers
G Protein-Coupled Receptors (GPCRs) are the largest family of membrane proteins targeted by drugs in clinical practice
Substantial evidence exists in support of their association in cell membranes, it is unclear how general and/or long-lasting this phenomenon is and whether it plays a significant role in GPCR function
Summary
G Protein-Coupled Receptors (GPCRs) have been reported to associate in the cell membrane to form dimers/oligomers. Very few GPCR oligomerization studies have been able to provide any information about the fraction of receptors that are interacting at a given time or the corresponding dynamics of the interactions, rendering it impossible to determine, with any certainty, which molecular species (i.e. individual protomers, dimers, or higher-order oligomers) signal through interaction with intracellular proteins. Single-molecule total internal reflection fluorescence microscopy (TIR-FM) was recently used to track the position of individual molecules of the M1 muscarinic acetylcholine receptor (M1R) labeled with fluorescent M1R antagonists in living cells [10] Both single- and dual-color imaging experiments suggested a transient (,0.5 seconds) formation of M1R dimers and a dimeric fraction of only ,30% dimers at any given time. It remains to be determined whether or not the features highlighted in these studies are the same for all GPCRs, or just specific subtypes
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