School of Life Science, Beijing Institute of Technology, Beijing, China.
PLoS One. 2013 Dec 4;8(12):e81335. doi: 10.1371/journal.pone.0081335. eCollection 2013.
Several crystal (Cry) proteins are known to occur as DNA-protein complexes. However, the role of the DNA associated with the activated toxin in the mechanism of action of the Cry toxin has long been ignored. Here, we focused on the DNA-activated Cry toxin complex. Both forms of the Cry8Ca2 and Cry8Ea1 toxins, i.e., with or without bound DNA, were separately obtained. Size-exclusion chromatography analysis indicated that the Cry8Ca2 toxin-DNA complex has a tight or compact structure. The Cry8Ca2 toxin-DNA complex is more likely to move toward the air/water interface and is more hydrophobic than the toxin without DNA. Competitive binding assays indicated that the Cry8Ca2 and Cry8Ea1 toxins without DNA specifically bind to the midgut of Anomala corpulenta and Holotrichia parallela larvae, respectively. In contrast, the association of DNA with each toxin might result in the nonspecific recognition of the Cry toxin and its target receptor in the insect midgut. The association of the DNA fragment with the Cry8 toxin was shown to protect the Cry protein from digestion by proteases. Based on our results, we propose an additional step in the mechanism of action of the Cry8 toxin and elucidate the function of the associated DNA as well as the importance of the removal of this DNA for the insecticidal activity of the toxin.
几种晶体（Cry）蛋白已知以 DNA-蛋白复合物的形式存在。然而，与激活毒素相关的 DNA 在 Cry 毒素作用机制中的作用长期以来一直被忽视。在这里，我们专注于 DNA 激活的 Cry 毒素复合物。分别获得了两种形式的 Cry8Ca2 和 Cry8Ea1 毒素，即与 DNA 结合或不结合的形式。凝胶排阻色谱分析表明，Cry8Ca2 毒素-DNA 复合物具有紧密或紧凑的结构。Cry8Ca2 毒素-DNA 复合物更有可能向空气/水界面移动，并且比没有 DNA 的毒素更疏水。竞争性结合测定表明，没有 DNA 的 Cry8Ca2 和 Cry8Ea1 毒素分别特异性结合到 Anomala corpulenta 和 Holotrichia parallela 幼虫的中肠。相比之下，DNA 与每种毒素的结合可能导致 Cry 毒素及其在昆虫中肠的靶受体的非特异性识别。与 DNA 片段的结合显示 Cry8 毒素蛋白免受蛋白酶的消化。基于我们的结果，我们提出了 Cry8 毒素作用机制的一个额外步骤，并阐明了相关 DNA 的功能以及去除该 DNA 对毒素杀虫活性的重要性。
