The synthetic approach to prostaglandins via bicyclo-[2.2.1]heptane derivatives, which has previously been 10

SUMMARYThe selective a 2 -adrenergic receptor antagonist rauwolscine was structurally modified to yield a series of arylamine carboxamide derivatives, which were investigated as potential molecular probes for the localization and structural characterization of a 2 -adrenergic receptors. The arylamine carboxamides differ in the number of carbon atoms separating the reactive phenyl moiety from the fused ring structure of the parent compound, rauwolscine carboxylate. Competitive inhibition studies with

No abstract

No abstract

A functionalized derivative of the a2-selective antagonist rauwolscine formed the basis for a photoaffinity adduct that has allowed identification of the hormone-binding subunit of the brain a2-adrenergic receptor protein. Rauwolscine carboxylate underwent reaction with 4-N-t-butyloxycarbonylaminoaniline, leading to the synthesis of rauwolscine 4-aminophenyl carboxamide (Rau-AmPC). Rau-AmPC was radioiodinated and converted to the arylazide derivative, 17a-hydroxy-ide (12'I-Rau-AzPC), via a diazonium salt intermediate. The characterization of '2sI-Rau-AzPC as a photolabile probe employed a2-adrenergic receptors, which were first solubilized from porcine brain membranes and partially purified by affinity chromatography utilizing a yohimbine-agarose affinity matrix. In the partially purified receptor preparation incubated with 125I-Rau-AzPC, photolysis resulted in covalent labeling of a major (Mr, 62,000) peptide as determined by NaDodSO4/PAGE and autoradiography. Labeling of this peptide was inhibited by the a2-selective antagonist, yohimbine, and the non-subtype-selective a-antagonist, phentolamine, but not by the a,-antagonist, prazosin, or the fl-receptor antagonist, (-)-alprenolol. The aadrenergic agonist epinephrine also inhibited labeling in a stereoselective manner. These data indicate that the photolabeled Mr 62,000 peptide is the hormone-binding subunit of the a2-adrenergic receptor protein. The availability of this radioiodinated photoaffinity probe for the a2-adrenergic receptor should facilitate further structural and biophysical characterization of the receptor protein.a-Adrenergic receptors have been subtyped as a, and a2 based on their relative affinities for a variety of synthetic compounds, as assessed in both functional and radioligand binding studies (1, 2). a2-Adrenergic receptors are located at both pre-and postjunctional sites in the central nervous system and peripheral organs, as well as in noninnervated tissues (2, 3). Although a2-adrenergic receptors are coupled in an inhibitory manner to adenylate cylase in many tissues, other mechanisms of signal transduction may be involved as well (4,5). In addition, species and tissue differences in a2-adrenergic receptors exist with respect to ligand recognition properties, which may reflect differences in the molecular structure of their hormone-binding subunits (6, 7). To understand the mechanisms involved in signal transduction and the factors that are responsible for discriminating among various agonist/antagonist ligands requires purification and structural characterization of the receptor itself. Although progress in this regard has been made with the human platelet a2-adrenergic receptor (8-14), characterization of this receptor in innervated tissues has not progressed as rapidly. Photoaffinity labels provide a tool for identifying the hormone-binding subunit of receptor proteins and have proven invaluable in studies concerned with the structural characterization of a,-and P-adrenergic receptors (15)(16)(17)(18)(19). This report descr...