0000000000326672
AUTHOR
Kurt Jarnagin
Na+ ions binding to the bradykinin B2 receptor suppress agonist-independent receptor activation.
Control of the balance between receptor activation and inactivation is a prerequisite for seven transmembrane domain (7TM) receptor function. We asked for a mechanism to stabilize the inactive receptor conformation which prevents agonist-independent receptor activation. Na+ ions have reciprocal effects on agonist versus antagonist interaction with various 7TM receptors. To investigate the Na+ dependence of receptor activation we chose the bradykinin B2 receptor as a prototypic 7TM receptor. Decrease of the intracellular Na+ content from 40 mM to 10 mM of COS-1 cells transiently expressing rat B2 receptors activated the B2 receptor in the absence of agonist as shown by a 3-fold increase in t…
The N-terminal Amino Group of [Tyr8]Bradykinin Is Bound Adjacent to Analogous Amino Acids of the Human and Rat B2 Receptor
To obtain data of the bradykinin B2 receptor's agonist binding site, we used a combined approach of affinity labeling and "immunoidentification" of receptor fragments generated by cyanogen bromide cleavage. Domain-specific antibodies to the various extracellular receptor domains were applied to detect receptor fragments with covalently attached [125I-Tyr8]bradykinin. As a cross-linker we used the homobifunctional reagent disuccinimidyl tartarate (DST), which reacts preferentially with primary amines. With this technique a [125I-Tyr8]bradykinin-labeled receptor fragment derived from the third extracellular domain was identified. The epsilon-amino group of lysine (Lys172) of the human B2 rece…
Extracellular Domains of the Bradykinin B2 Receptor Involved in Ligand Binding and Agonist Sensing Defined by Anti-peptide Antibodies
Many of the physiological functions of bradykinin are mediated via the B2 receptor. Little is known about binding sites for bradykinin on the receptor. Therefore, antisera against peptides derived from the putative extracellular domains of the B2 receptor were raised. The antibodies strongly reacted with their corresponding antigens and cross-reacted both with the denatured and the native B2 receptor. Affinity-purified antibodies to the various extracellular domains were used to probe the contact sites between the receptor and its agonist, bradykinin or its antagonist HOE140. Antibodies to extracellular domain 3 (second loop) efficiently interfered, in a concentration-dependent manner, with…