Evidence for dimerization in the β 2 ‐adrenergic receptor from the evolutionary trace method

The evolutionary trace method (Lichtarge et al., Proc Natl Acad Sci USA 1996, 93, 7507) was applied to the adrenergic‐receptor sequences. The conserved and “conserved‐in‐class” residues were determined for successive splits along the phylogenetic tree. These residues were then plotted on the internal and external faces of a model of the β 2 ‐adrenergic receptor. The adrenergic‐receptor model was constructed using knowledge of the helix–helix packing angles in the cryoelectron microscopy structure of rhodopsin and the ideal ridges‐in‐grooves helix packing patterns known to reproduce these angles. Two clusters were observed on the external (lipid‐facing) surface of the receptor model: a major one on helices 5 and 6 and a minor one on helices 2 and 3. The importance of some of the residues on helices 5 and 6 was confirmed by site‐direceted mutagenesis. In contrast, very few residues were plotted on the external face of helices 1, 4, or 7. The major cluster is consistent with the dimerization interface in G‐protein‐coupled receptor domain‐swapped dimers, which is proposed to occur between helices 5 and 6. The minor cluster is of unknown function. The clusters on the internal faces contain the known ligand‐binding sites, as determined by site‐directed mutagenesis. In particular, there is a line of conserved residues on helices 2–7 at a depth of about 14 Å. On helices 2 and 3, and on 6 and 7, the cluster extends considerably deeper than the known binding site. These deeper clusters contain the conserved DRY and NPXXY motifs on helices 3 and 7, respectively, and so are probably related to receptor activation. ©1999 John Wiley & Sons, Inc. Int J Quant Chem 74: 371–379, 1999