LYSINE(200) LOCATED IN THE 5TH TRANSMEMBRANE DOMAIN OF THE HISTAMINE H-1 RECEPTOR INTERACTS WITH HISTAMINE BUT NOT WITH ALL H-1 AGONISTS

Previously, we have shown that asparagine(207) in the fifth transmembr ane domain of the histamine H-1 receptor is crucial for the binding of the N-tau-nitrogen of the imidazole ring of histamine (Leurs et al., Biochem. Biophys. Res. Commun., 201, 295, 1994). In view of the potent ial interaction of the imidazole ring of histamine histamine with a bi nding sire, formed by asparagine(207) and lysine(200), we mutated lysi ne(200) in the fifth transmembrane domain of the histamine H-1 recepto r to a nonfunctional alanine residue. This mutation did nor affect the binding of the tested H-1 receptor antagonists bur resulted in a 5-fo ld lower affinity for histamine. The binding of other H-1 receptor ago nists was not affected. In stably transfected CHO cells histamine was 55-fold less effective in activating the H-1 Lys(200)Ala receptor (EC( 50) = 66 mu M) compared to the wild type H-1 receptor (EC(50) = 1.2 mu M). Receptor activation by the 2-methyl and the 2-(3-bromophenyl)-ana logues however was hardly affected by the mutation, indicating that th e 2-substituent probably prevents the interaction with the lysine(200) residue. Finally, the Lys(200)Ala mutation reduced the production of [H-3]inositol phosphates, stimulated by the non-imidazole H-1 receptor agonist 2-pyridylethylamine. These data indicate that lysine(200) int eracts with the N-pi-nitrogen of histamine and is important for the ac tivation of the H-1 receptor by histamine and the non-imidazole agonis t 2-pyridylethylamine. (C) 1995 Academic Press. Inc.