中红侧沟茧蜂化学感受蛋白MmedCSP1的结合特征

化学感受蛋白是一类存在于昆虫化学感受器中的可溶性蛋白, 被认为与昆虫识别外界化学信息有关。本研究使用pGEX-4T-1表达载体在BL21 (DE3)异源表达系统中表达中红侧沟茧蜂Microplitis mediator化学感受蛋白MmedCSP1, 并通过亲和层析法纯化得到去表达标签的MmedCSP1；使用bis-ANS作为荧光配基, 在荧光分光光度计上研究它与50种气味标样的结合特征, 从而得到此类化学感受蛋白在中红侧沟茧蜂嗅觉识别中识别气味的种类。结果表明: MmedCSP1只能与水杨酸甲酯、戊烷、罗勒烯、β-紫罗兰酮、3, 4-二甲基苯甲醛、2-己酮和叶醇结合。但只有脂类化合物β-紫罗兰酮能在浓度为1 mmol/L下将bis-ANS从MmedCSP1中替换50%, β-紫罗兰酮与MmedCSP1的结合常数为16.89 μmol/L。这些结果提示MmedCSP1参与中红侧沟茧蜂对水杨酸甲酯、戊烷、罗勒烯、β-紫罗兰酮、3, 4-二甲基苯甲醛、2-己酮和叶醇等气味的识别过程, 且在不同气味中的识别过程中对于气味的运输能力有差异。

Binding characterization of chemosensory protein MmedCSP1 in Microplitis mediator (Hymenoptera: Braconidae)

Chemosensory proteins (CSPs), which are assumed to be related with the process of storing and releasing the chemical odors for insects, belong to a class of small soluble polypeptides in insect chemical receptors. In this study, chemosensory protein of Microplitis mediator MmedCSP1 was expressed using pGEX-4T-1/BL21 (DE3) prokaryotic expression system, and then purified using GSTrap FF, and the binding characteristics of MmedCSP1 to 50 kinds of chemical molecules were further studied using bis-ANS (4, 4′-dianilino-1, 1′- binaphthyl- 5, 5′-sulfonate) as fluorescent probes. The results showed that MmedCSP1 could bind to methyl salicylate, pentane, ocimene, β-ionone, 3, 4-dimethylbenzaldehyde, 2-hexanone and cis-3-hexen-l-ol. However, only β-ionone replaced the bis-ANS of MmedCSP1 by 50%, and the binding constant was 16.89 μmol/L. These results suggest that MmedCSP1 is involved in the process of storing and releasing methyl salicylate, pentane, ocimene, β-ionone, 3, 4-dimethylbenzaldehyde, 2-hexanone and cis-3-hexen-l-ol for M. mediator, and has different transport capability in storing and releasing those odorants.