CorrectionsBIOCHEMISTRY. For the article ''RNA degradosomes exist in vivo in Escherichia coli as multicomponent complexes associated with the cytoplasmic membrane via the N-terminal region of ribonuclease E'' by Gunn-Guang Liou, Wann-Neng Jane, Stanley N.

G protein-coupled receptors (GPCRs) are embedded in phospholipids that strongly influence drug-stimulated signaling. Anionic lipids are particularly important for GPCR signaling complex formation, but a mechanism for this role is not understood. Using NMR spectroscopy, we explore the impact of anionic lipids on the function-related conformational equilibria of the human A2A adenosine receptor (A2AAR) in bilayers containing defined mixtures of zwitterionic and anionic phospholipids. Anionic lipids prime the receptor to form complexes with G proteins through a conformational selection process. Without anionic lipids, signaling complex formation proceeds through a less favorable induced fit mechanism. In computational models, anionic lipids mimic interactions between a G protein and positively charged residues in A2AAR at the receptor intracellular surface, stabilizing a pre-activated receptor conformation. Replacing these residues strikingly alters the receptor response to anionic lipids in experiments. High sequence conservation of the same residues among all GPCRs supports a general role for lipid-receptor charge complementarity in signaling.

The 826 human G protein‐coupled receptors (GPCRs) are involved in nearly every physiological process and comprise the largest class of “druggable” proteins. For over the past decade, crystal and cryo‐EM structures have focused on revealing molecular details of GPCR interactions with small molecules and intracellular partner proteins, which are well‐established regulators of GPCR function. More recently, literature data from biological and biophysical studies have demonstrated that phospholipids found in the endogenous membrane environment also control GPCR function and act as allosteric modulators. In particular, phospholipids with charged headgroups have been reported to be especially important regulators of GPCR function and facilitate the formation of signaling complexes. Using nuclear magnetic resonance (NMR) spectroscopy in aqueous solutions, we investigated the structural basis for the regulation of GPCR activity by phospholipids in nanodiscs containing the human A2A adenosine receptor (A2AAR), a representative class A GPCR. Lipid nanodiscs containing mixtures of uncharged and charged phospholipids were produced, yielding homogeneous preparations over a wide range of lipid compositions. NMR data of uniformly stable‐isotope labeled A2AAR showed the receptor was properly folded in lipid nanodiscs and shared a similar global conformation to detergent micelle preparations of A2AAR. Using 19F NMR with conformationally‐sensitive probes located at the A2AAR intracellular surface, we systematically investigated the influence of lipid composition over a wide range of mixtures of charged and uncharged phospholipids. A2AAR complexes with antagonists showed almost no response to variation in lipid composition. In striking contrast to this, A2AAR complexes with agonists showed a clear response to changes in the relative proportion of charged phospholipids. NMR data of agonist complexes in nanodiscs containing lower proportions of charged lipids were highly similar to data of antagonist complexes, showing a higher proportion of neutral lipids negated the efficacy of bound agonists. As the proportion of charged phospholipids increased, the relative population of an A2AAR active conformation also increased. The dependence of the receptor response to bound agonists on lipid composition suggested specific interactions between lipid headgroups and charged amino acids at the A2AAR intracellular surface underlie the regulation of activity. Results from this study indicate that lipid composition must be carefully selected in studies of GPCRs in nanodiscs. Our data also provide a potential molecular mechanism for how changes in membrane composition, due to disease for example, can directly impact the efficacy of drugs.

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