title: Photoaffinity labeling of avermectin binding sites from Caenorhabditis elegans and Drosophila melanogaster.
abstract: An azido-avermectin analog [4'' alpha-(4-azidosalicylamido-epsilon-caproylamido-beta-alan ylamido)-4''-deoxyavermectin B1a; azido-AVM] was synthesized and used to photoaffinity label avermectin binding sites present in the membranes of Caenorhabditis elegans and Drosophila melanogaster. Azido-AVM was biologically active and behaved like a competitive inhibitor of [3H]ivermectin binding to C. elegans membranes (Ki = 0.2 nM). Radiolabeled azido-AVM bound specifically and with high affinity to C. elegans membranes (Kd = 0.14 nM) and, upon photoactivation, became covalently linked to three C. elegans polypeptides of 53, 47, and 8 kDa. Photoaffinity labeling of a membrane preparation from D. melanogaster heads resulted in labeling of a single major polypeptide of approximately 47 kDa. The proteins that were covalently tagged in these experiments are believed to be associated with avermectin-sensitive chloride channels present in the neuromuscular systems of C. elegans and D. melanogaster. Azido-AVM did not bind to rat brain membranes and therefore was selective for the nematode and insect receptors.
authors.affiliation: Department of Biochemical Parasitology, Merck Sharp and Dohme Research Laboratories, Rahway, NJ 07065.
authors.forename: S P
authors.initials: SP
authors.lastname: Rohrer
authors.forename: P T
authors.initials: PT
authors.lastname: Meinke
authors.forename: E C
authors.initials: EC
authors.lastname: Hayes
authors.forename: H
authors.initials: H
authors.lastname: Mrozik
authors.forename: J M
authors.initials: JM
authors.lastname: Schaeffer
mesh_terms: D000345:Affinity Labels; D000818:Animals; D001665:Binding Sites; D002107:Caenorhabditis / Q000737:chemistry*; D018118:Chloride Channels; D004331:Drosophila melanogaster / Q000737:chemistry*; D007559:Ivermectin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000302:isolation & purification*; D010777:Photochemistry; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D000345:Affinity Labels; D018118:Chloride Channels; D008565:Membrane Proteins; D011506:Proteins; D007559:Ivermectin; C019264:avermectin