Wang Huatang, Chen Huiling, Wang Zhengbing, Liu Jiali, Zhang Xingyan, Li Chaofeng, Zeng Xinnian
Guangdong Engineering Research Center for Insect Behavior Regulation, College of Agriculture South China Agricultural University, Guangzhou, China.
Guangdong Key Laboratory of Animal Conservation and Resource Utilization, Guangdong Public Laboratory of Wild Animal Conservation and Utilization, Guangdong Institute of Applied Biological Resources, Guangzhou, Guangdong, China.
Environ Entomol. 2019 Feb 13;48(1):245-252. doi: 10.1093/ee/nvy179.
For insects, odorant-binding proteins (OBPs) play an essential role in binding and transporting semiochemicals through the sensillum lymph to olfactory receptor neurons within the antennal sensilla. In the present study, the full-length cDNA encoding a general odorant-binding protein 1 (DcitOBP1, accession number KY475564) was cloned from the antennae of Diaphorina citri using RACE-PCR, and qRT-PCR analysis revealed that the DcitOBP1 gene was expressed mainly in the antennae of D. citri. In molecular docking assay, the results showed that DcitOBP1 protein has better binding affinities to the 12 selected host-plant volatile compounds. Then, the recombinant DcitOBP1 protein was expressed in Escherichia coli. After removed His-Tag, the binding properties of purified DcitOBP1 protein to the selected host-plant volatile compounds were investigated in a fluorescence ligand-binding assay, similar, but more obviously binding properties of DcitOBP1 protein result were obtained, the dissociation constant (KD) value of DcitOBP1/1-NPN complex was 6.440 ± 0.521, and the DcitOBP1 protein showed high binding affinities (IC50 < 100 μM) to six of the selected ligands, namely methyl salicylate, α-phellandrene, (1R)-(+)-α-pinene, 3-carene, β-caryophyllene, and α-caryophyllene. Additionally, the behavior bioassays were also showed that D. citri had significant behavioral responses toward to α-caryophyllene, β-caryophyllene, (1R)-(+)-α-pinene, and α-phellandrene. Our investigation infer that the DcitOBP1 protein might play a crucial role in host-plant volatile odorants' perception in D. citri, and these results also have been supplied previous insight evidence into the physiological functions of the DcitOBP1 protein of D. citri.
对于昆虫而言，气味结合蛋白（OBPs）在结合和运输信息化学物质方面发挥着至关重要的作用，这些物质通过感器淋巴传递至触角感器内的嗅觉受体神经元。在本研究中，利用RACE-PCR从柑橘木虱触角中克隆出编码一种普通气味结合蛋白1（DcitOBP1，登录号KY475564）的全长cDNA，qRT-PCR分析表明DcitOBP1基因主要在柑橘木虱的触角中表达。在分子对接试验中，结果显示DcitOBP1蛋白对12种选定的寄主植物挥发性化合物具有更好的结合亲和力。然后，重组DcitOBP1蛋白在大肠杆菌中表达。去除His标签后，在荧光配体结合试验中研究了纯化的DcitOBP1蛋白与选定的寄主植物挥发性化合物的结合特性，获得了相似但更明显的DcitOBP1蛋白结合特性结果，DcitOBP1/1-NPN复合物的解离常数（KD）值为6.440±0.521，并且DcitOBP1蛋白对六种选定配体，即水杨酸甲酯、α-水芹烯、（1R）-（+）-α-蒎烯、3-蒈烯、β-石竹烯和α-石竹烯表现出高结合亲和力（IC50<100μM）。此外，行为生物测定还表明柑橘木虱对α-石竹烯、β-石竹烯、（1R）-（+）-α-蒎烯和α-水芹烯有显著的行为反应。我们的研究推断DcitOBP1蛋白可能在柑橘木虱对寄主植物挥发性气味的感知中起关键作用，这些结果也为柑橘木虱DcitOBP1蛋白的生理功能提供了先前的深入证据。
