The binding of propranolol at 5-hydroxytryptamine1D beta T355N mutant receptors may involve formation of two hydrogen bonds to asparagine.
Although the beta-adrenergic receptor antagonist (-)-propranolol binds with relatively low affinity at human 5-hydroxytryptamine1D beta receptors (Ki = 10,200 nM), it displays significantly higher affinity (Ki = 17 nM) at its species homolog, 5-HT1B receptors, and at a mutant 5-HT1D beta receptor (Ki = 16 nM), where the threonine residue at position 355 (T355) is replaced with an asparagine residue (i.e., a T355N mutant). Propranolol contains two oxygen atoms, an ether oxygen atom and a hydroxyl oxygen atom, and it has been speculated that the enhanced affinity of propranolol for the T355N mutant receptor is related to the ability of the asparagine residue to hydrogen bond with the ether oxygen atom. However, the specific involvement of the propranolol oxygen atoms in binding to the wild-type and T355N mutant 5-HT1D beta receptors has never been addressed experimentally. A modification of a previously described 5-HT1D beta receptor graphic model was mutated by replacement of T355 with asparagine. Propranolol was docked with the wild-type and T355N mutant 5-HT1D beta receptor models in an attempt to understand the difference in affinity of the ligand for the receptors. The binding models suggest that the asparagine residue of the mutant receptor can form hydrogen bonds with both oxygen atoms of propranolol, whereas the threonine moiety of the wild-type receptor can hydrogen-bond only to one oxygen atom. To test this hypothesis, we prepared and examined several analogues of propranolol that lacked either one or both oxygen atoms. The results of radioligand binding experiments are consistent with the hypothesis that both oxygen atoms of propranolol could participate in binding to the mutant receptor, whereas only the ether oxygen atom participates in binding to the wild-type receptor. As such, this is the first investigation of serotonin receptors that combines the use of molecular modeling, mutant receptors generated by site-directed mutagenesis, and synthesis to investigate structure/affinity relationships.
Glennon
R A
RA
Department of Medicinal Chemistry, School of Pharmacy, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0540, USA.
Dukat
M
M
Westkaemper
R B
RB
Ismaiel
A M
AM
Izzarelli
D G
DG
Parker
E M
EM
D000319:Adrenergic beta-Antagonists / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D003198:Computer Simulation; D006801:Humans; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D009154:Mutation; D011433:Propranolol / Q000378:metabolism*; D011869:Radioligand Assay; D011985:Receptors, Serotonin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D000319:Adrenergic beta-Antagonists; D011985:Receptors, Serotonin; D001216:Asparagine; D011433:Propranolol