Department of Cell Biology and Neuroscience, University of California, Riverside, CA 92521, USA.
Appl Environ Microbiol. 2011 Jan;77(1):24-31. doi: 10.1128/AEM.01852-10. Epub 2010 Oct 29.
Cry11Ba is one of the most toxic proteins to mosquito larvae produced by Bacillus thuringiensis. It binds Aedes aegypti brush border membrane vesicles (BBMV) with high affinity, showing an apparent dissociation constant (K(d)) of 8.2 nM. We previously reported that an anticadherin antibody competes with Cry11Ba binding to BBMV, suggesting a possible role of cadherin as a toxin receptor. Here we provide evidence of specific cadherin repeat regions involved in this interaction. Using cadherin fragments as competitors, a C-terminal fragment which contains cadherin repeat 7 (CR7) to CR11 competed with Cry11Ba binding to BBMV. This binding was also efficiently competed by the CR9, CR10, and CR11 peptide fragments. Moreover, we show CR11 to be an important region of interaction with Cry11Ba toxin. An alkaline phosphatase (AaeALP1) and an aminopeptidase-N (AaeAPN1) also competed with Cry11Ba binding to Ae. aegypti BBMV. Finally, we found that Cry11Ba and Cry4Ba share binding sites. Synthetic peptides corresponding to loops α8, β2-β3 (loop 1), β8-β9, and β10-β11 (loop 3) of Cry4Ba compete with Cry11Ba binding to BBMV, suggesting Cry11Ba and Cry4Ba have common sites involved in binding Ae. aegypti BBMV. The data suggest that three different Ae. aegypti midgut proteins, i.e., cadherin, AaeALP1, and AaeAPN1, are involved in Cry11Ba binding to Ae. aegypti midgut brush border membranes.
Cry11Ba 是苏云金芽孢杆菌产生的对蚊子幼虫最具毒性的蛋白质之一。它与埃及伊蚊刷状缘膜小泡（BBMV）具有高亲和力，表现出明显的解离常数（K(d)）为 8.2 nM。我们之前报道过一种抗钙粘蛋白抗体与 Cry11Ba 结合 BBMV 竞争，表明钙粘蛋白可能作为毒素受体发挥作用。在这里，我们提供了涉及这种相互作用的特定钙粘蛋白重复区域的证据。使用钙粘蛋白片段作为竞争物，包含钙粘蛋白重复 7（CR7）至 CR11 的 C 端片段与 Cry11Ba 结合 BBMV 竞争。该结合也被 CR9、CR10 和 CR11 肽片段有效地竞争。此外，我们表明 CR11 是与 Cry11Ba 毒素相互作用的重要区域。碱性磷酸酶（AaeALP1）和氨肽酶-N（AaeAPN1）也与 Cry11Ba 结合 Ae. 竞争aegypti BBMV。最后，我们发现 Cry11Ba 和 Cry4Ba 共享结合位点。Cry4Ba 的α8、β2-β3（环 1）、β8-β9 和 β10-β11（环 3）环对应的合成肽与 Cry11Ba 结合 BBMV 竞争，表明 Cry11Ba 和 Cry4Ba 具有共同的结合位点参与结合 Ae. 埃及伊蚊 BBMV。数据表明，三种不同的埃及伊蚊中肠蛋白，即钙粘蛋白、AaeALP1 和 AaeAPN1，参与 Cry11Ba 与埃及伊蚊中肠刷状缘膜的结合。
