Drug discovery targeting human 5-HT(2C) receptors: residues S3.36 and Y7.43 impact ligand-binding pocket structure via hydrogen bond formation.
Specific activation of serotonin (5-HT) 5-HT(2C) G protein-coupled receptors may be therapeutic for obesity and neuropsychiatric disorders. Mutagenesis coupled with computational and molecular modeling experiments based on the human β₂ adrenergic receptor structure was employed to delineate the interactions of different ligands at human 5-HT(2C) residues D3.32, S3.36 and Y7.43. No binding of the tertiary amine radioligand ([³H]-mesulergine) could be detected when the 5-HT(2C) D3.32 residue was mutated to alanine (D3.32A). The S3.36A point-mutation greatly reduced affinity of primary amine ligands, modestly reduced affinity of a secondary amine, and except for the 5-HT(2C)-specific agonist N(CH₃)₂-PAT, affinity of tertiary amines was unaffected. Molecular modeling results indicated that the primary amines form hydrogen bonds with the S3.36 residue, whereas, with the exception of N(CH₃)₂-PAT, tertiary amines do not interact considerably with this residue. The Y7.43A point-mutation greatly reduced affinity of 5-HT, yet reduced to a lesser extent the affinity of tryptamine that lacks the 5-hydroxy moiety present in 5-HT; modeling results indicated that the 5-HT 5-hydroxy moiety hydrogen bonds with Y7.43 at the 5-HT(2C) receptor. Additional modeling results showed that 5-HT induced a hydrogen bond between Y7.43 and D3.32. Finally, modeling results revealed two low-energy binding modes for 5-HT in the 5-HT(2C) binding pocket, supporting the concept that multiple agonist binding modes may stabilize different receptor active conformations to influence signaling. Ligand potencies for modulating WT and point-mutated 5-HT(2C) receptor-mediated phospholipase C activity were in accordance with the affinity data. Ligand efficacies, however, were altered considerably by the S3.36A mutation only.
Department of Medicinal Chemistry, University of Florida, P.O. Box 100485, 1600 SW Archer Road, Gainesville, FL 32610-0485, USA.
Clinton E
CE
Canal
Tania C
TC
Cordova-Sintjago
Nancy Y
NY
Villa
Li-Juan
LJ
Fang
Raymond G
RG
Booth
D000588:Amines / Q000737:chemistry; D016503:Drug Delivery Systems; D015195:Drug Design; D006801:Humans; D006860:Hydrogen Bonding; D008024:Ligands; D001523:Mental Disorders / Q000188:drug therapy / Q000503:physiopathology; D008958:Models, Molecular; D009422:Nervous System Diseases / Q000188:drug therapy / Q000503:physiopathology; D009765:Obesity / Q000188:drug therapy / Q000503:physiopathology; D017354:Point Mutation; D044405:Receptor, Serotonin, 5-HT2C / Q000187:drug effects* / Q000378:metabolism; D058826:Serotonin 5-HT2 Receptor Agonists / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000378:metabolism
D000588:Amines; D008024:Ligands; D044405:Receptor, Serotonin, 5-HT2C; D058826:Serotonin 5-HT2 Receptor Agonists; D010738:Type C Phospholipases