Bacillus thuringiensis subsp israelensis Cyt1Aa synergizes Cry11Aa toxin by functioning as a membrane-bound receptor
Bacillus thuringiensis subsp israelensis Cyt1Aa synergizes Cry11Aa toxin by functioning as a membrane-bound receptor
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DOI：
10.1073/pnas.0505494102
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发表时间：
2005-12-20
影响因子：
11.1
通讯作者：
Bravo, A
中科院分区：
文献类型：
--
作者：
Pérez, C;Fernandez, LE;Bravo, A
苏云金芽胞Israelensis产生晶体蛋白，Cry（4Aa、4 Ba、10Aa和11 Aa）和Cyt（1Aa和2Ba）蛋白，对人类疾病的蚊子媒介有毒。Cyt 1Aa克服了昆虫对Cry 11 Aa和Cry 4毒素的抗性，并协同这些毒素的毒性。然而，协同作用的分子机制仍然没有解决。在这里，我们提供的证据表明，Cyt 1Aa的功能作为Cry 11 Aa的受体。Cyt 1Aa和Cry 11 Aa的序列结合分析显示，Cyt 1Aa与埃及伊蚊刷状缘膜囊泡的结合增强了生物素化的Cry 11 Aa的结合。Cyt 1Aa-和Cry 11 Aa-结合表位的酵母双杂交系统，肽阵列，异源竞争试验与合成肽映射。Cyt 1Aa中的两个暴露区域，环β 6-α E和部分β 7，结合Cry 11 Aa。另一方面，Cry 11 Aa通过结构域II环α 8和β 4结合Cyt 1Aa蛋白，这也参与了中肠受体相互作用。对Cry 11 Aa和Cyt 1Aa中的单点突变的表征揭示了关键的Cry 11 Aa（S259和E266）和Cyt 1Aa（K198、E204和K225）残基参与两种蛋白质的相互作用和协同作用。此外，还分离出了对Cry 11 Aa具有增强协同作用的Cyt 1Aa环β 6-α E突变体（K198 A）。这里提供的数据强烈表明，Cyt 1Aa协同或抑制抗性Cry 11 Aa毒素作为膜结合受体的功能。苏云金芽胞Israelensis是一种高效的致病菌，因为它产生毒素及其功能受体，促进毒素与靶膜结合并引起毒性。
Bacillus thuringiensis subsp. israelensis produces crystal proteins, Cry (4Aa, 4Ba, 10Aa, and 11Aa) and Cyt (1Aa and 2Ba) proteins, toxic to mosquito vectors of human diseases. Cyt1Aa overcomes insect resistance to Cry11Aa and Cry4 toxins and synergizes the toxicity of these toxins. However, the molecular mechanism of synergism remains unsolved. Here, we provide evidence that Cyt1Aa functions as a receptor of Cry11Aa. Sequential-binding analysis of Cyt1Aa and Cry11Aa revealed that Cyt1Aa binding to Aedes aegypti brush border membrane vesicles enhanced the binding of biotinylated-Cry11Aa. The Cyt1Aa- and Cry11Aa-binding epitopes were mapped by means of the yeast two-hybrid system, peptide arrays, and heterologous competition assays with synthetic peptides. Two exposed regions in Cyt1Aa, loop beta 6-alpha E and part of beta 7, bind Cry11Aa. On the other side, Cry11Aa binds Cyt1Aa proteins by means of domain II-loop alpha 8 and beta-4, which are also involved in midgut receptor interaction. Characterization of single-point mutations in Cry11Aa and Cyt1Aa revealed key Cry11Aa (S259 and E266) and Cyt1Aa (K198, E204 and K225) residues involved in the interaction of both proteins and in synergism. Additionally, a Cyt1Aa loop beta 6-alpha E mutant (K198A) with enhanced synergism to Cry11Aa was isolated. Data provided here strongly indicates that Cyt1Aa synergizes or suppresses resistance to Cry11Aa toxin by functioning as a membrane-bound receptor. Bacillus thuringiensis subsp. israelensis is a highly effective pathogenic bacterium because it produces a toxin and also its functional receptor, promoting toxin binding to the target membrane and causing toxicity.