Trends in Pharmacological Sciences
ReviewGeneric GPCR residue numbers – aligning topology maps while minding the gaps
Section snippets
New era in GPCR research
G protein-coupled receptors (GPCRs) constitute the largest family of human cell surface receptors [1]. They respond endogenously to ions, neurotransmitters, lipids, carbohydrates, nucleotides, amino acids, peptides and proteins; and also sense light, pain, tastes, and odors [2]. Their abundance in human physiological systems, as well as their accessibility and druggability, have made them a major drug target family – ∼30% of the marketed drugs act on GPCRs [3]. The GPCRs are typically
Generic residue numbers - maps to navigate GPCR topology
All GPCRs share a structural core of seven transmembrane (7TM) helices, making up the machinery for signal transduction across the cell membrane. The 7TM domain contains or is part of the binding site of class A and B1 receptor ligands, and serves as a site for allosteric modulation of class B2, C, and F GPCRs 9, 10. So far, 109 GPCRs have been drugged [11], the vast majority with ligands binding within the TM region [12]. The conserved 7TM scaffold allows for the alignment of sequences or
Class A GPCR residue numbering
The Ballesteros–Weinstein numbering scheme [14] is based on the presence of highly conserved residues in each of the seven transmembrane (TM) helices. It consists of two numbers where the first denotes the helix, 1–7, and the second the residue position relative to the most-conserved residue, defined as number 50. For example, 5.42 denotes a residue located in TM5, eight residues before the most-conserved residue, P5.50. The residue numbers can be counted directly within the receptor protein
Class B, C, and F GPCR residue numbering
Class B, C, and F schemes have been established using the same procedure as the class A Ballesteros-Weinstein system, but use unique reference positions (X.50) such that the residue numbers can be counted directly within the receptor protein sequence (alignment). The class B GPCR Wootten [21] scheme is based on the B1/secretin subclass, but the reference residues are the most conserved also for five of the B2/adhesion receptor helices and the remaining two, TM3–4, still display high
Cross-class GPCR residue numbering
The low sequence conservation between the GPCR classes has hitherto hindered (correct) sequence alignments, although some inter-class receptor modeling studies correctly aligned the majority of the seven helices (e.g., 28, 29, 30). The structural conservation is higher and the recent crystallographic data have opened up for structure-based sequence alignments from class A to B 22, 23, 27, C 25, 31, and F 32, 33. Some helices display large inter-class lateral deviations or different bending but,
Extracellular loop 2 residue numbering
Sequence analysis shows that there is a large diversity in the lengths and compositions of the N- and C-termini as well as in the extracellular and intracellular loops connecting the TM helices of GPCRs [46]. Nevertheless, for the extracellular loop 2 (EL2) a similar residue-numbering scheme has been applied 41, 47 in which EL2 residues are labeled 45.X, indicating the location between TM4 and TM5 (“45”) [47]. The reference position (X.50) is a conserved cysteine forming a disulfide bridge with
Mind the gap – introducing a novel complementary crystal structure-based GPCR residue numbering
Class A GPCRs have long been known to contain helix kinks in TM5–7 that are induced by highly conserved proline residues, which are unable to form backbone hydrogen bonds [51]. Upon receptor activation, they act as hinges that allow the helices to tilt inwards to tighten the ligand cavity and outwards to widen the G protein-binding pocket [7]. This does not affect the generic residue numbers because the best corresponding alignment of residues is obtained knowing that receptor structures are
Numbering made easy – residue-numbering web tools at GPCRDB
GPCRDB has been a major community resource for more than 20 years 15, 71, 72, 73. It contains reference data including the largest available collection of receptor mutants, crystal structures, 3D structure models in the inactive and active states, and sequence alignments for all species in UniProt. Recently, GPCRDB was equipped with a new suite of interactive web browser tools and diagrams; these include phylogenetic trees, sequence motif search (e.g., for a binding site), and receptor sequence
Potential exceptions and future directions
The sequence- and structure-based numbering systems share some limitations. Generic residues can only be assigned to receptor regions with a conserved structural fold, in other words the 7TM domain, whereas the termini and loops can only be compared, in the best case, within receptor subfamilies. Both systems also depend on sequence homology to produce sequence alignments, although this dependency decreases for the GPCRDB numbering as new crystal structure templates become available. In fact,
Concluding remarks
The first crystal structures of the class B, C, and F GPCRs have opened up the field for receptor function studies and drug design. In addition, cross-class sequence alignments can now be constructed, enabling us to uncover the common cogs and cranks within the 7TM machinery. We have described the schemes for generic numbering of such residue hotspots for chemical structure-activity relationships, pharmacological effects of receptor mutants, and structural mechanisms. Furthermore, the
Acknowledgments
The Lundbeck Foundation provided financial support to V.I. and D.E.G., who was also funded by the Carlsberg Foundation. V.C., V.K., and R.C.S are supported by NIH grants U54 GM094618 and P01 DA035764. G.V. acknowledges TIPharma for financial supports. This work was also supported by COST Action CM1207 (GLISTEN). Kasper Harpsøe is acknowledged for discussions on structure superposition and residue numbering.
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