摘要

晶体蛋白(crystal protein, Cry)毒素是苏云金芽胞杆菌(Bacillus thuringiensis, Bt)分泌的主要杀虫蛋白,其中Cry4Ba和Cry11Aa对埃及伊蚊(Aedes aegypti)幼虫具有特异的毒杀活性。Cry毒素作用于昆虫肠道上皮细胞需要细胞膜受体的介导,而昆虫肠道的免疫应答因子通过识别Cry毒素,进而驱动免疫系统抵御毒素的侵染。为了系统探究Cry4Ba和Cry11Aa毒素与埃及伊蚊的分子互作机制,本研究通过制备GST-Cry4Ba和GST-Cry11Aa融合蛋白,提取Cry4Ba和Cry11Aa晶体蛋白,利用GST-pull down和免疫共沉淀技术挖掘埃及伊蚊中肠刷状缘膜泡囊蛋白中与Cry4Ba和Cry11Aa互作的结合蛋白。通过液相色谱串联质谱鉴定和数据库比对,发现133个与Cry4Ba互作蛋白和237个与Cry11Aa互作蛋白。通过生物信息学分析,从中筛选到16个膜结合或跨膜蛋白,包括5个氨肽酶N、1个碱性磷酸酶、1个钙黏蛋白、2个α-淀粉酶、1个B型清道夫受体、2个电压依赖性阴离子通道以及4个肌动蛋白。此外,筛选到7个与免疫或维持细胞稳态相关蛋白,包括1个肽聚糖识别蛋白、1个载脂蛋白、1个血粘蛋白、1个丝氨酸蛋白酶抑制剂以及3个热激蛋白。本研究为后续埃及伊蚊Bt受体的功能验证以及免疫应答机制的研究提供必要的靶标参考。

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