title: Role of basic amino acids of the human angiotensin type 1 receptor in the binding of the non-peptide antagonist candesartan.
abstract: To explain the insurmountable/long-lasting binding of biphenyltetrazole-containing AT1-receptor antagonists such as candesartan, to the human angiotensin II type 1-receptor, a model is proposed in which the basic amino acids Lys199 and Arg 167 of the receptor interact respectively with the carboxylate and the tetrazole group of the antagonists. To validate this model, we have investigated the impact of substitution of Lys199 by Ala or Gln and of Arg167 by Ala on the binding properties of [3H]candesartan and on competition binding by candesartan, EXP3174, irbesartan, losartan, angiotensin II (Ang II) and [Sar1-Ile8]angiotensin. Our results indicate that both amino acids play an important role in the AT1-receptor ligand binding. Whereas the negative charge of Lys 199 is involved in an ionic bond with the end-standing carboxylate group of the peptide ligands, its polarity also contributes to the non-peptide antagonist binding. Substitution of Arg167 by Ala completely abolished [3H]Ang II, as well as [3H] candesartan, binding. Whereas these results are in line with the proposed model, it cannot be excluded that both amino acid residues are important for the structural integrity of the AT1-receptor with respect to its ligand binding properties.
authors.affiliation: Department of Molecular and Biochemical Pharmacology, Institute for Molecular Biology and Biotechnology, Free University of Brussels (VUB), B-1640 Sint-Genesius Rode, Belgium, gvauquel@ vub.ac.be.
authors.forename: Georges
authors.initials: G
authors.lastname: Vauquelin
authors.affiliation: Department of Molecular and Biochemical Pharmacology, Institute for Molecular Biology and Biotechnology, Free University of Brussels (VUB), B-1640 Sint-Genesius Rode, Belgium.
authors.forename: Frederik Lp
authors.initials: FL
authors.lastname: Fierens
authors.affiliation: Department of Physiology, Semmelweis University Medical School, H-1444 Budapest, PO Box 259, Hungary.
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Gáborik
authors.affiliation: Department of Molecular and Biochemical Pharmacology, Institute for Molecular Biology and Biotechnology, Free University of Brussels (VUB), B-1640 Sint-Genesius Rode, Belgium.
authors.forename: Tam
authors.initials: T
authors.lastname: Le Minh
authors.affiliation: Department of Molecular and Biochemical Pharmacology, Institute for Molecular Biology and Biotechnology, Free University of Brussels (VUB), B-1640 Sint-Genesius Rode, Belgium.
authors.forename: Jean-Paul
authors.initials: JP
authors.lastname: De Backer
authors.affiliation: Department of Physiology, Semmelweis University Medical School, H-1444 Budapest, PO Box 259, Hungary.
authors.forename: László
authors.initials: L
authors.lastname: Hunyady
authors.affiliation: Department of Molecular and Biochemical Pharmacology, Institute for Molecular Biology and Biotechnology, Free University of Brussels (VUB), B-1640 Sint-Genesius Rode, Belgium.
authors.forename: Patrick Ml
authors.initials: PM
authors.lastname: Vanderheyden
keywords: CHO cells; angiotensin II; candesartan; human AT1-receptor; insurmountable; mutation; non-peptide antagonist