title: A carbene-generating photoaffinity probe for beta-adrenergic receptors.
abstract: A new radioiodinated (2.2 Ci/mumol) iodocyanopindolol derivative carrying a 4-(3-trifluoromethyldiazirino)benzoyl residue has been synthesized. The long-wavelength absorption of the diazirine permits formation of the carbene by photolysis under very mild conditions. [125I]ICYP-diazirine binds with high affinity (Kd = 60 pM) to beta-receptors from turkey erythrocyte membranes. Upon irradiation, [125I]ICYP-diazirine is covalently incorporated in a Mr 40 000 protein. Stereoselective inhibition of photolabeling by the (-)enantiomers of alprenolol and isoproterenol indicated that the Mr 40 000 protein contains a beta-adrenergic binding site. The yield of specific labeling was up to 8.2% of total beta-receptor binding sites. The Mr 40 000 protein photolabeled in the membrane could be solubilized at comparable yield with either digitonin or Triton X-100. Irradiation of digitonin-solubilized turkey erythrocyte membranes with [125I]ICYP-diazirine resulted in specific labeling of two proteins with Mr 40 000 and 50 000. In guinea-pig lung membranes, at least five proteins were photolabeled, of which one (with approximately Mr 67 000) was labeled specifically.
mesh_terms: D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D001389:Azirines / Q000378:metabolism*; D004072:Digitonin; D004910:Erythrocyte Membrane / Q000378:metabolism; D006168:Guinea Pigs; D008168:Lung / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D010782:Photolysis; D011941:Receptors, Adrenergic / Q000032:analysis*; D011943:Receptors, Adrenergic, beta / Q000032:analysis* / Q000378:metabolism; D012995:Solubility; D014422:Turkeys
chemical_list: D000345:Affinity Labels; D001389:Azirines; D008565:Membrane Proteins; D011941:Receptors, Adrenergic; D011943:Receptors, Adrenergic, beta; C037848:1-(2-(3-trifluoromethyldiazirino)benzoyl)-3,3-dimethyl-6-hydroxy-7-(2-cyano-3-iodoindol-4-yloxy)-1,4-diazaheptane; D004072:Digitonin