Nodulisporic Acid Opens Insect Glutamate-Gated Chloride Channels: Identification of a New High Affinity Modulator

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• 作者：McHardy M. Smith ; Vivien A. Warren ; Brande S. Thomas ; Richard M. Brochu ; Eric A. Ertel ; Susan Rohrer ; James Schaeffer ; Dennis Schmatz ; Brian R. Petuch ; Yui Sing Tang ; Peter T. Meinke ; Gregory J. Kaczo
• 刊名：Biochemistry
• 出版年：2000
• 出版时间：May 9, 2000
• 年：2000
• 卷：39
• 期：18
• 页码：5543 - 5554
• 全文大小：172K
• 年卷期：v.39,no.18(May 9, 2000)
• ISSN：1520-4995

文摘

Nodulisporic acid (NA) is an indole diterpene fungal product with insecticidal activity. NAactivates a glutamate-gated chloride channel (GluCl) in grasshopper neurons and potentiates channel openingby glutamate. The endectocide ivermectin (IVM) induces a similar, but larger current than NA. UsingDrosophila melanogaster head membranes, a high affinity binding site for NA was identified. Equilibriumbinding studies show that an amide analogue, N-(2-hydroxyethyl-2,2-³H)nodulisporamide ([³H]NAmide),binds to a single population of sites in head membranes with a K_D of 12 pM and a B_max of 1.4 pmol/mgof protein. A similar K_D is determined from the kinetics of ligand binding and dissociation. Four lines ofevidence indicate that the binding site is a GluCl. First, NA potentiates opening of a glutamate-gatedchloride current in grasshopper neurons. Second, glutamate inhibits the binding of [³H]NAmide byincreasing the rate of dissociation 3-fold. Third, IVM potently inhibits the binding of [³H]NAmide andIVM binds to GluCls. Finally, the binding of [³H]IVM is inhibited by NA. The B_max of [³H]IVM is twicethat of [³H]NAmide, and about half of the [³H]IVM binding sites are inhibited by NA with high affinity(K_I = 25 pM). In contrast, [³H]IVM binding to Caenorhabditis elegans membranes is not inhibited byNA at 100 nM, and there are no high affinity binding sites for NA on these membranes. Thus, half of theDrosophila IVM receptors and all of the NA receptors are associated with GluCl. NA distinguishes betweennematode and insect GluCls and identifies subpopulations of IVM binding sites.