Abstract
Lipids are becoming known as essential allosteric modulators of G protein-coupled receptor (GPCRs). However, how they exert their effects on GPCR conformation at the atomic level is still unclear. In light of recent experimental data, we have performed several long-timescale molecular dynamics (MD) simulations, totalling 24 μs, to rigorously map allosteric modulation and conformational changes in the β2 adrenergic receptor (β2AR) that occur as a result of interactions with three different phospholipids. In particular, we identify different sequential mechanisms behind receptor activation and deactivation, respectively, mediated by specific lipid interactions with key receptor regions. We show that net negatively charged lipids stabilize an active-like state of β2AR that is able to dock Gsα protein. Clustering of anionic lipids around the receptor with local distortion of membrane thickness is also apparent. On the other hand, net-neutral zwitterionic lipids inactivate the receptor, generating either fully inactive or intermediate states, with kinetics depending on lipid headgroup charge distribution and hydrophobicity. These chemical differences alter membrane thickness and density, which differentially destabilize the β2AR active state through lateral compression effects.
Highlights
G protein-coupled receptors (GPCRs) are implicated in the regulation of many physiological and pathological processes[1]
We focus on the allosteric effects of dioleoyl-sn-glycero-3-p hosphoglycerol (DOPG), DOPE, and DOPC phospholipids on the conformational changes of the active state of β2 adrenergic receptor (β2AR), including modelled intracellular loop 3 (ICL3), in unbiased atomistic long-timescale Molecular dynamics (MD) simulations
In light of observations of relevant β2AR crystal structures, proposed ICL3 functionality[23], and experimental data that shows β2AR is strongly allosterically modulated by different phospholipids[44], we have investigated the sum of these effects by employing long-timescale MD simulations of the active state of apo β2AR (PDB entry: 3SN6)[14] in three different homogenous membranes consisting of lipids: DOPC, DOPE and DOPG, respectively
Summary
Β2AR is one of the most studied and frequently crystallized Class A GPCRs. the structure of intracellular loop 3 (ICL3) that connects TM5 and TM6 is unknown because it is missing in all β2AR crystal structures. This is supported by the overall RMSD of the receptor remaining at 2.0–3.4 Å compared to the initial active crystal structure and finishing at 2.3 Å (SI Fig. 10) Taken together, these results indicate that DOPG lipids are able to stabilize an active-like state of β2AR, even without a bound G protein or bound agonist. In order to probe further, we calculated g(r) a second time, for just positively charged sidechains located at the intracellular end of TM6 (four lysine residues: K2636.25, K2676.29, K2706.32, K2736.35, see SI Fig. 12) This analysis reveals peaks in g(r) at 4.5–5.0 Å in all three membrane types (DOPG > DOPC > DOPE), which likely reflects electrostatic interactions between positive charges on TM6 and negative charges on lipid phosphate groups. The more hydrophobic headgroup of DOPC lipids facilitates moderate interaction with TM6, and an absence of inter-lipid H-bonds contributes to lower membrane density
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