Antagonist conformations within the beta(2)-adrenergic receptor ligand binding pocket

The interactions between beta-adrenergic receptor (beta AR) antagonists and the beta(2)AR were studied with the use of photoaffinity labels. A proteolytic map of the receptor was made and confirmed through amino-terminal amino acid sequencing by locating sites of derivatization. [I-125]Iodoazidothiophenylalprenolol (IAPTA) is a photoaffinity derivative of the PAR antagonist alprenolol with a photoactivatable group on the aryloxy end of the molecule. IAPTA exclusively derivatizes a peptide consisting of transmembrane domains (TMs) 6 and 7 of the hamster lung beta(2)AR supporting the contention that TMs 6 and 7 interact with the aryloxy portion of the PAR antagonist pharmacophore. The beta AR antagonist photoaffinity labels [I-125]iodoazidobenzylpindolol (IABP), [(125)]iodoazidophenyl CGP-12177A (IAPCGP), and [I-125]iodocyanopindololdiazarene (ICYPdz) are similar in that their photoactive moieties are attached to the amino end of the antagonist pharmacophore, IABP derivatized TMs 5-7 and a peptide containing TM 1 to approximately equal extents. IAPCGP derivatized TMs 6 and 7 much greater than TM 5 = TM 4 = TMs 2 and 3 = TM 1. ICYPdz derivatized TM 1 much greater than TMs 6 and 7 > TM 4. We conclude that the aryloxy end of the beta AR antagonist pharmacophore is highly constrained within TMs 6 and 7, whereas the amino terminus is much less constrained and able to assume multiple conformations. Molecular dynamics simulations predict that IABP, IAPCGP, and ICYPdz favor a folded conformation, with both ends close together. Derivatization of TMs 6 and 7 by IABP, IAPCGP, and ICYPdz suggests the folded conformation of these compounds in the ligand binding pocket.