Key Laboratory of Natural Pesticide & Chemical Biology, Ministry of Education; College of Agriculture, South China Agricultural University, Guangzhou 510642, China.
Toxins (Basel). 2019 Jun 18;11(6):349. doi: 10.3390/toxins11060349.
Destruxin A (DA), a major secondary metabolite of , has anti-immunity to insects. However, the detailed mechanism and its interactions with target proteins are elusive. Previously, three immunophilins, peptidyl-prolyl cis-trans isomerase (BmPPI), FK506 binding-protein 45 (BmFKBP45) and BmFKBP59 homologue, were isolated from the silkworm, Bm12 cell line following treatment with DA, which suggested that these proteins were possible DA-binding proteins. To validate the interaction between DA and the three immunophilins, we performed bio-layer interferometry (BLI) assay, and the results showed that DA has interaction with BmPPI, whose affinity constant value is 1.98 × 10 M and which has no affinity with FKBP45 and FKBP59 homologue in vitro. Furthermore, we investigated the affinity between DA and human PPI protein (HsPPIA) and the affinity constant (K) value is 2.22 × 10 M. Additionally, we compared the effects of silkworm and human PPI proteins produced by DA and immunosuppressants, cyclosporine A (CsA), and tacrolimus (FK506), by employing I2H (insect two-hybrid) in the SF-9 cell line. The results indicated that in silkworm, the effects created by DA and CsA were stronger than FK506. Furthermore, the effects created by DA in silkworm were stronger than those in humans. This study will offer new thinking to elucidate the molecular mechanism of DA in the immunity system of insects.
杀虫脒 A（DA）是一种主要的次生代谢产物，具有抗昆虫免疫作用。然而，其详细的作用机制及其与靶蛋白的相互作用仍不清楚。先前，从 DA 处理的家蚕 Bm12 细胞系中分离出三种免疫亲合素，即肽基脯氨酰顺反异构酶（BmPPI）、FK506 结合蛋白 45（BmFKBP45）和 BmFKBP59 同源物，这表明这些蛋白可能是 DA 结合蛋白。为了验证 DA 与这三种免疫亲合素之间的相互作用，我们进行了生物层干涉（BLI）测定，结果表明 DA 与 BmPPI 相互作用，其亲和力常数值为 1.98×10 M，并且在体外与 FKBP45 和 FKBP59 同源物没有亲和力。此外，我们研究了 DA 与人 PPI 蛋白（HsPPIA）之间的亲和力及其亲和力常数（K）值为 2.22×10 M。此外，我们通过 SF-9 细胞系中的昆虫双杂交（I2H）比较了 DA 和免疫抑制剂环孢素 A（CsA）和他克莫司（FK506）对家蚕和人 PPI 蛋白的影响。结果表明，在家蚕中，DA 和 CsA 产生的作用强于 FK506。此外，DA 在蚕中的作用强于人。这项研究将为阐明 DA 在昆虫免疫系统中的分子机制提供新的思路。
