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Identifying Receptor‐Ligand Interactions That Drive Gβγ‐Dependent Signaling
Author(s) -
Cameron Robert Bruce,
Beeson Craig Bruce,
Schnellmann Rick
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.664.12
Subject(s) - formoterol , clenbuterol , chemistry , formoterol fumarate , agonist , receptor , pharmacology , ligand (biochemistry) , medicine , endocrinology , biochemistry , biology , chromatography , budesonide , corticosteroid
Understanding differences in ligand bias enable more targeted application of pharmacotherapy to various disease states. Our laboratory has previously demonstrated that in renal proximal tubule cells (RPTC), the beta‐2 adrenergic receptor agonist formoterol induces mitochondrial biogenesis, while the beta‐2 adrenergic receptor agonist clenbuterol does not. We hypothesized that formoterol and clenbuterol engage distinct signaling programs by engaging in different interaction profiles with the beta‐2 adrenergic receptor. We examined Gα s ‐ and Gβγ‐dependent signaling in RPTC following stimulation with 30 nM formoterol and 30 nM clenbuterol and found that both ligands caused significant accumulation of cAMP but that only formoterol caused a significant increase in Akt phosphorylation. To understand the structural basis of these differences in signaling, formoterol and clenbuterol were docked to the orthosteric binding site of both inactive (3NYA, 3NY8, 5D5B) and active (4DLE, 4LDL, 4LDO) crystal structures of the beta‐2 adrenergic receptor. For each pose, receptor‐ligand interactions were tabulated at each structural feature. Each interaction was assigned a value of +1 for formoterol and −1 for clenbuterol, and interactions were summed for all six crystal structures. The methoxyphenyl group of formoterol showed a greater likelihood of aromatic interactions with W109 and V114 and contact interactions with F193. Interestingly, the formamide and hydroxyl groups of formoterol exhibited a greater interaction frequency with the backbone of C191, but the amino group of clenbuterol was more likely to engage in strong interactions at that site. Similarly, the secondary amino groups of both formoterol and clenbuterol interacted with D113, but formoterol was more likely to do so with strong hydrogen bond or ionic interactions. Overall, the longer and larger functional groups of formoterol allowed it to stretch across the binding pocket of the beta‐2 adrenergic receptor to simultaneously interact with TM3, TM5, and ECL2 in a manner distinct from clenbuterol and may drive the differences in signaling between formoterol and clenbuterol. These interaction profiles provide key structural data for the development of beta‐2 adrenergic receptor agonists that selectively activate specific signaling programs. Support or Funding Information R.B.C. is supported by F30 DK104550 and T32 GM008716 (National Institutes of Health). C.C.B. is supported by P20 GM103542 (National Institutes of Health). R.G.S. is supported by R01 GM084147 (National Institutes of Health) and 1BX000851 (Department of Veterans Affairs).