Fig 1.
TM domains of the available crystal structures.
Top: Two views of the 24 inactive crystal structures from classes A, B, C, and F (aligned to β2) show the general GPCR fold of the transmembrane (TM) bundle. Class A in green, class B in blue (CRF1, GLR), class C in orange (MGLU1, MGLU5), class F in magenta (SMO). Bottom: Same views for only the 19 inactive class A structures showing the highly conserved class A TM fold. A detailed view of the conserved hydrogen bonding networks is shown in S1 Fig.
Table 1.
Number of GPCR sequences by class.
The total number of candidate human GPCR sequences that were considered are listed. The full list of Uniprot ACs is in S2 Table.
Fig 2.
Conserved inter-helical contacts.
Top left: Diagram of 40 conserved inter-helical contacts (CHICOs) present in at least 23 out of 24 studied class A structures. The contacts common to all classes are shown in purple, and contacts present only in class A in orange. Top right: List of these contacts in Ballesteros-Weinstein numbering scheme. Bottom: Extracellular view of the same contacts in the β2 crystal structure. The contacts in the inner and outer half of the membrane are shows on the left and right respectively.
Table 2.
Selection of the alignment between class A and classes B, C, and F.
This table shows the selection process for assigning BW.50 residues to non class A proteins. Shifting BW.50 residue on each helix renumbers the relative BW numbers, effectively changing the labels of contacts observed in these proteins. Subsequently, the number of common contacts each structure shares with the class A structures changes for different BW residue assignments. The second rightmost column shows the cumulative number of contact occurrences among the 24 class A structures (including active conformations). The BW assignment with the highest number of contacts is selected (except for MGLU5, see text). The selected alignment is in bold.
Fig 3.
Testing the robustness of the alignment of the Vomeronasal receptors with the other groups.
The table shows similarity between TMs averaged over all pairs of sequences formed from the two groups (red denotes high similarity, blue low similarity). For most TMs the optimal choices agree with the optimal alignment to Aα (full table in S5 Fig); all combinations are shown only for TM5. The same table but using the GPCRtm substitution matrix [74] instead of BLOSUM62 is shown in S7 Fig. GPCRtm was developed in particular for GPCR proteins, but in this case both matrices result in the same alignment.
Fig 4.
Testing the robustness of the alignment of the Taste2 receptors with the other groups.
The table shows similarity between TMs averaged over all pairs of sequences formed from the two groups (red denotes high similarity, blue low similarity). For most TMs the optimal choices agree with the optimal alignment to Aα (full table in S6 Fig) only TM6 shows a second possible alignment at offset +4. The same table but using the GPCRtm substitution matrix instead of BLOSUM62 is in S8 Fig. Again, both matrices result in the same alignment.
Fig 5.
TM 3 sequence alignment for the 25 crystal structures.
Other TMs are shown in Fig 6. The sequences are taken from the selected PDB files. The TM helix residues are colored in the Zappos scheme, which captures the chemical nature of each residue (e.g. helix breakers, proline and glycine, are shown in purple). The loop residues are shown in grey. The BW n.50 residue (numbering displayed below the sequences) is the most conserved within the class A. The consensus sequence is most similar to class A, because most sequences are from this class. The largest differences are for the last 5 sequences, which belong to the classes B, C, and F. The figure was prepared using Jalview.
Fig 6.
Sequence alignments for TMs 1,2,4–7 for the 25 crystal structures.
Same caption as Fig 5, where TM3 is shown.
Fig 7.
The phylogenetic tree based only on TM similarity using the GRoSS alignment (loops were ignored).
Color coding denotes the GPCR class. Proteins with known crystal structure are emphasized with a dot. The full resolution version of this figure is in S4 Fig.
Fig 8.
Native activation “hot-spot” residues (NACHOs), which are contacts that change upon receptor activation.
The width of the green lines is proportional to the number of contacts common to all six structures (RHO, β2AR, M2, and their active structures). Blue shows the contacts present only in inactive structures, and not in inactive structures; while red shows the opposite. The upper diagrams show contacts in the extracellular half of the membrane. We see that there is no systematic change common to the class A receptors in the conformation of the extracellular half of the TMs. This is not obvious, because there are conformational changes accompanying ligand binding. All the systematic changes, which enable G protein binding, occur in the intracellular half of the TMs. The list only contains 15 different residues in 15 different contacts. Thus many of the residues switch partners upon activation.
Table 3.
Examples of natural variants and mutations that are associated with functional change or disease and which coincide with the NACHO residues.
Table 4.
Summary of SNPs annotated on Uniprot. The complete list is in S3 Table.
Fig 9.
Magnitude of the rigid body moves of the helices necessary to map one structure to another.
All TMs 1–7 from all available structure pairs were compared and each symbol denotes which TM is the data point from. The coordinate system is defined in the text. The maximal observed deviation is approximately proportional to the sequence dissimilarity of the two compared TMs, and it follows the same trend within class A (blue symbols) and across the GPCR superfamily (green symbols). The red symbols, which correspond to the active-inactive structure pairs, show rigid body moves caused by receptor activation. S10 Fig has an analogous plot of residual RMSD vs. similarity for each helix after the best rigid body transformation. RMSD shows a similar trend as the plots in this figure.