Department of Applied Biological Chemistry, Faculty of Agriculture, Kindai University, Nara, Japan.
Centre for Respiratory Biology, UCL Respiratory, University College London, London, UK.
Br J Pharmacol. 2018 Jun;175(11):1999-2012. doi: 10.1111/bph.13914. Epub 2017 Aug 24.
Neonicotinoid insecticides interact with the orthosteric site formed at subunit interfaces of insect nicotinic ACh (nACh) receptors. However, their interactions with the orthosteric sites at α-non α and α-α subunit interfaces remain poorly understood. The aim of this study was to elucidate the mechanism of neonicotinoid actions using the Drosophila Dα1-chicken β2 hybrid nACh receptor.
Computer models of the (Dα1) (β2) nACh receptor in complex with imidacloprid and thiacloprid were generated. Amino acids in the Dα1 subunit were mutated to corresponding amino acids in the human α4 subunit to examine their effects on the agonist actions of neonicotinoids on (Dα1) (β2) and (Dα1) (β2) nACh receptors expressed in Xenopus laevis oocytes using voltage-clamp electrophysiology.
The (Dα1) (β2) nACh receptor models indicated that amino acids in loops D, E and G probably determine the effects of neonicotinoids. The amino acid mutations tested had minimal effects on the EC for ACh. However, the R57S mutation in loop G, although having minimal effect on imidacloprid's actions, reduced the affinity of thiacloprid for the (Dα1) (β2) nACh receptor, while scarcely affecting thiacloprid's action on the (Dα1) (β2) nACh receptor. Both the K140T and the combined R57S;K140T mutations reduced neonicotinoid efficacy but only for the (Dα1) (β2) nACh receptor. Combining the E78K mutation with the R57S;K140T mutations resulted in a selective reduction of thiacloprid's affinity for the (Dα1) (β2) nACh receptor.
These findings suggest that a triangle of residues from loops D, E and G contribute to the selective actions of neonicotinoids on insect-vertebrate hybrid nACh receptors.
This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc.
新烟碱类杀虫剂与昆虫烟碱乙酰胆碱（nACh）受体亚基界面形成的正位部位相互作用。然而，它们与α-非α和α-α亚基界面的正位部位的相互作用仍知之甚少。本研究旨在使用果蝇 Dα1-鸡β2 杂种 nACh 受体阐明新烟碱类作用的机制。
生成与吡虫啉和噻虫啉复合的（Dα1）（β2）nACh 受体的计算机模型。将 Dα1 亚基中的氨基酸突变为人类α4 亚基中的相应氨基酸，以使用电压钳电生理学检查它们对新烟碱类对在非洲爪蟾卵母细胞中表达的（Dα1）（β2）和（Dα1）（β2）nACh 受体的激动剂作用的影响。
（Dα1）（β2）nACh 受体模型表明，环 D、E 和 G 中的氨基酸可能决定新烟碱类的作用。测试的氨基酸突变对 ACh 的 EC 影响最小。然而，尽管 G 环中的 R57S 突变对吡虫啉的作用影响最小，但它降低了噻虫啉与（Dα1）（β2）nACh 受体的亲和力，而对噻虫啉对（Dα1）（β2）nACh 受体的作用几乎没有影响。K140T 和 R57S;K140T 突变均降低了新烟碱类的效力，但仅对（Dα1）（β2）nACh 受体有效。将 E78K 突变与 R57S;K140T 突变相结合导致噻虫啉对（Dα1）（β2）nACh 受体的亲和力选择性降低。
这些发现表明，来自环 D、E 和 G 的残基三角形有助于新烟碱类对昆虫-脊椎动物杂种 nACh 受体的选择性作用。
本文是关于烟碱乙酰胆碱受体的专题部分的一部分。要查看该部分中的其他文章，请访问 http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc。
