Site-directed mutagenesis of the human 5-HT1B receptor

• Published in: European journal of pharmacology Vol. 349; no. 2-3; pp. 367 - 375
• Main Authors: GRANÄS, C, NORDVALL, G, LARHAMMAR, D
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier 22.05.1998
• Subjects: Amino Acid Substitution; Animals; Biological and medical sciences; CHO Cells; Cricetinae; Cyclic AMP - biosynthesis; General pharmacology; Humans; Medical sciences; Mutagenesis, Site-Directed; Pharmacokinetics. Pharmacogenetics. Drug-receptor interactions; Pharmacology. Drug treatments; Receptor, Serotonin, 5-HT1B; Receptors, Serotonin - chemistry; Receptors, Serotonin - drug effects; Receptors, Serotonin - metabolism; Serotonin - metabolism; Serotonin - pharmacology; Serotonin Receptor Agonists - metabolism; Serotonin Receptor Agonists - pharmacology; Sumatriptan; Rodentia; Site directed mutagenesis; Molecular interaction; Ergot derivatives; 5-HT1 Serotonine receptor; In vitro; Ovary; Vertebrata; Mammalia; Structure activity relation; Animal; Established cell line; Hamster; Methysergide
• ISSN: 0014-2999; 1879-0712
• DOI: 10.1016/S0014-2999(98)00213-1

Site-directed mutagenesis was used to investigate the molecular interactions involved in ligand binding to the human 5-HT1B receptor. Six mutants were constructed at four positions and expressed in Chinese hamster ovary cells. Substitution of the amino acid F185 in transmembrane region IV by an alanine increased the affinities of sumatriptan, methysergide and 8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT) 3-4-fold and substitution by a methionine increased the affinities of methysergide and methiothepin 2- and 3-fold, respectively. Substitution of amino acid S334 in transmembrane region VI by an alanine increased the affinity of 8-OH-DPAT 5-fold. In accordance with this, the EC50 value of 8-OH-DPAT was decreased 7-fold. This suggests that the serine at position 334 causes steric hindrance for 8-OH-DPAT binding that is lost in the S334A mutant. Mutation of F354 in transmembrane region VII, which differs between receptor subtypes, increased the affinity of methiothepin 2-3-fold but the affinities of the other compounds tested were essentially unchanged.