PT  - JOURNAL ARTICLE
AU  - Dana B. Fathy
AU  - Donald J. Kyle
AU  - L. M. Fredrik Leeb-Lundberg
TI  - High-Affinity Binding of Peptide Agonists to the Human B1 Bradykinin Receptor Depends on Interaction between  the Peptide N-Terminal <span class="sc">l</span>-Lysine and the Fourth Extracellular  Domain of the Receptor
DP  - 2000 Jan 01
TA  - Molecular Pharmacology
PG  - 171--179
VI  - 57
IP  - 1
4099  - http://molpharm.aspetjournals.org/content/57/1/171.short
4100  - http://molpharm.aspetjournals.org/content/57/1/171.full
SO  - Mol Pharmacol2000 Jan 01; 57
AB  - The aim of this study was to identify the location of the N terminus of peptide agonist ligands when bound to the human B1 bradykinin (BK) receptor. To reach this aim, we exploited the fact that high-affinity binding of kinin peptides to the human B1 receptor subtype requires a peptide N-terminal l-Lys, whereas high-affinity binding to the B2 receptor subtype does not require this residue. This was done by comparing the affinities of BK, a B2 receptor-selective peptide, and kallidin or Lys-BK, a less receptor-selective peptide, for chimeric proteins in which each B1 receptor domain had been substituted in the human B2 receptor and expressed in HEK293 cells. Individual substitution of transmembrane domains 1–7 (TM-I–VII) and extracellular domains 1–4 (EC-I–IV) of the B1 receptor in the B2 receptor influenced the affinities of BK and Lys-BK approximately equally. In contrast, substitution of B1 EC-IV dramatically reduced the affinity and potency of BK, whereas these parameters for Lys-BK were essentially unaltered. Substitution of either the N- or C-terminal half of B1 EC-IV in the B2 receptor only had a limited effect on the peptide binding constants, indicating the involvement of multiple residues throughout this domain. Complementary mutations of the N-terminal residue in Lys-BK revealed that both the positive charge and the proper spatial orientation of this residue were required for interaction with B1 EC-IV. Thus, the N-terminal residue of peptide agonists when bound to the human B1 receptor is positioned extracellularly and interacts with EC-IV. The American Society for Pharmacology and Experimental Therapeutics