Journal: Nature Communications

Article Title: Structural basis for lipid-mediated activation of G protein-coupled receptor GPR55

doi: 10.1038/s41467-025-57204-y

Figure Lengend Snippet: a Cryo-EM map (consensus, EMDB-51285) of the GPR55-Gα 13 β 1 γ 2 -ScFv16-LPI complex at two different contour levels. The enlarged cryo-EM map for LPI (yellow sticks) is shown in blue mesh. b Full model corresponding to the signaling complex of ( a ) (shown as cartoon representation). c Chemical structure of LPI. d Overview of ligand binding pocket position of LPI. e Cryo-EM map (consensus, EMDB-51281) of the GPR55-Gα 13 β 1 γ 2 -ScFv16-ML184 complex at two different contour levels. The enlarged cryo-EM map for ML184 (salmon sicks) is shown in blue mesh. f Full model corresponding to the signaling complex of ( d ). g Overview of ligand binding pocket position of ML184. The yellow circle highlights the position of the polar head group of LPI. h Chemical structure of ML184.

Article Snippet: The stable GPR55-Gα 13 β 1 γ 2 complex was formed by the addition of 25 mU mL −1 apyrase (NEB, cat. #M0398) and agonists (10 µM LPI or 20 µM ML184), followed by incubation for 1.5 h at room temperature.

Techniques: Cryo-EM Sample Prep, Ligand Binding Assay

Journal: Nature Communications

Article Title: Structural basis for lipid-mediated activation of G protein-coupled receptor GPR55

doi: 10.1038/s41467-025-57204-y

Figure Lengend Snippet: a Ligand binding pocket of LPI (yellow sticks). The amino acid side chain and backbone that show interactions with LPI are shown as cyan sticks. Hydrogen bonds are indicated as black dashed lines. b Effect of GPR55 mutants on LPI potency as determined by G protein dissociation assays with Gα 13 . Data represent means ± 95% confidence interval (CI) from 3–7 independent experiments as indicated in Supplementary Table . c ligand binding pocket of ML184. d Effect of GPR55 mutants on ML184 potency as determined by G protein dissociation assays with Gα 13 . Data represent means ± 95% CI from 3–7 independent experiments as indicated in Supplementary Table . e Magnified view of the ML184 (salmon sticks) binding pocket (as in a ). The binding pocket surface was displayed in yellow with the ECL2 surface hidden for better visibility of the binding pocket. f Efficacy and constitutive activity at different GPR55 mutants. The degree of activation was calculated by normalization of BRET² ratios (BRET² ratio of 30 µM ligand for efficacy or basal BRET² ratios of 1.5% DMSO) to the respective BRET² ratios for the wt GPR55 plus G 13 -biosensor at 30 µM ML184 (100% activation) and for a mock-transfection plus G 13 -biosensor (0% activation). The pEC 50 means of all seven mutations were compared with the pEC 50 mean of the wt GPR55 to evaluate statistically significant differences using ordinary one-way ANOVA with Dunnett’s post-hoc test (adjusted P -values: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, for absolute P -values see Supplementary Table ).

Article Snippet: The stable GPR55-Gα 13 β 1 γ 2 complex was formed by the addition of 25 mU mL −1 apyrase (NEB, cat. #M0398) and agonists (10 µM LPI or 20 µM ML184), followed by incubation for 1.5 h at room temperature.

Techniques: Ligand Binding Assay, Binding Assay, Activity Assay, Activation Assay, Transfection

Journal: Nature Communications

Article Title: Structural basis for lipid-mediated activation of G protein-coupled receptor GPR55

doi: 10.1038/s41467-025-57204-y

Figure Lengend Snippet: a Side view of the GPR55-LPI complex (cyan surface representation and yellow spheres, respectively) shows membrane gate into the hydrophobic channel of the orthosteric binding pocket between helices IV and V. Membrane lipids were not resolved in this structure. b Side view of the GPR55-ML184 (light cyan surface representation) complex resolved a CLR molecule (green spheres) bound in a similar cleft between helices IV and V. c Superimposition of the GPR55-LPI and GPR55-ML184 structures (receptor in cartoon representation, with side chains that contact CLR as sticks) show the CLR (green sticks) binding pocket with direct contacts to ML184 (salmon sticks). Subtle sidechain rearrangements in the LPI structure within the CLR interface are observed. The dotted yellow line represents the distance measurement (in Å) between K180 5.37 and CLR.

Article Snippet: The stable GPR55-Gα 13 β 1 γ 2 complex was formed by the addition of 25 mU mL −1 apyrase (NEB, cat. #M0398) and agonists (10 µM LPI or 20 µM ML184), followed by incubation for 1.5 h at room temperature.

Techniques: Membrane, Binding Assay

Journal: Nature Communications

Article Title: Structural basis for lipid-mediated activation of G protein-coupled receptor GPR55

doi: 10.1038/s41467-025-57204-y

Figure Lengend Snippet: a Overview of the GPR55-G 13 -ML184 signaling complex. ML184 and CLR are shown as salmon and green spheres, respectively. Colored rectangles highlight specific G protein interaction sections as described in panels ( e , f ), respectively. The map quality of one water molecule deviates significantly from four well-resolved water molecules. The modeling of this water molecule was guided by its coordination with R119 3.50 , helix VI, and helix VII. b , c G protein dissociation assays of ( b ) GPR55 and ( c ) TBXA2R with Gα 12 and Gα 13 . Assays were performed in HEK293H cells transiently transfected with wt Gβ 3 , Gγ 9 -GFP, and Gα 12 -Rluc8 or Gα 13 -Rluc8. Data represents means ± SEM from 3–7 independent experiments as specified in Supplementary Table . The negative control data for U-46619 at GPR55 and ML184 or LPI at the TBXA2R was analyzed from three independent experiments. d Sequence alignment of different G protein segments of Gα 13 involved in GPR55 binding with the respective residues of Gα 12 . Black arrows highlight amino acid differences between Gα 12 and Gα 13 within 4 Å of GPR55. Residues R360 and Q338 are predominantly solvent-exposed and not shown in the following panels. e Protein-protein interface between Gα 13 (green cartoon and sticks) and GPR55 (cyan cartoon and sticks) with a focus on the C -terminal α5-helix of Gα 13 . The cryo-EM composite map (EMDB-51284) for five water molecules is shown with orange mesh. f Protein-protein interactions of residues between the α4-helix and β6-sheet of Gα 13 as well as of the initial α5-helix residues with GPR55.

Article Snippet: The stable GPR55-Gα 13 β 1 γ 2 complex was formed by the addition of 25 mU mL −1 apyrase (NEB, cat. #M0398) and agonists (10 µM LPI or 20 µM ML184), followed by incubation for 1.5 h at room temperature.

Techniques: Transfection, Negative Control, Sequencing, Binding Assay, Solvent, Cryo-EM Sample Prep