中华蜜蜂普通气味结合蛋白ASP2的气味结合功能模式分析

【目的】研究中华蜜蜂（Apis cerana cerana）体外重组普通气味结合蛋白AcerASP2与不同气味信息的结合功能和模式。【方法】通过诱导条件的优化和纯化，在获得的重组AcerASP2蛋白基础上，研究竞争性荧光探针1-NPN及不同结构气味信息与蛋白的相互作用关系，并通过同源建模和分子对接解析蛋白与气味信息的结合模式和机理。【结果】通过条件摸索获得了可溶性表达的重组中蜂ASP2蛋白，荧光光谱分析1-NPN与AcerASP2的解离常数K1-NPN为7.38 μmol•L-1，结合位点数n为1.0321。在7种气味信息中，4-烯丙基藜芦醚亲和力最强，其IC50和解离常数KD分别达到7.09和3.46 μmol•L-1。分子对接显示AcerASP2具有1个呈狭长的口袋状的疏水性结合内腔，4-烯丙基藜芦醚绝大部分位于该预测疏水性内腔中，且与Lys74产生2个氢键。【结论】中华蜜蜂AcerASP2由于特殊的空间结构而具有较强的气味信息结合能力，且易通过与疏水内腔中的赖氨酸残基产生氢键而促进结合。

Interpretation of Odorant Binding Function and Mode of General Odorant Binding Protein ASP2 in Chinese Honeybee (Apis cerana cerana)

【Objective】 The objective of this study is to research the binding function and mode of different odors with general odorant binding protein AcerASP2 of Chinese honeybees (Apis cerana cerana). 【Method】 With the optimization of induction conditions, the purified recombinant AcerASP2 protein was obtained, then the competitive fluorescence assay was used to determine the binding function of AcerASP2 with odors having different structures, finally the homology modeling and molecular docking were applied to elucidate the binding mode and mechanism. 【Result】After the soluble recombinant AcerASP2 protein purified, in the competitive fluorescence assay between AcerASP2 and 1-NPN, the dissociation constants K1-NPN and the number of binding sites n were 7.38 μmol•L-1 and 1.0321, respectively. In selective 7 kinds of odors, the affinity of 4-allylveratrole seemed to be the strongest, and the IC50 and dissociation constant KD were 7.09 and 3.46 μmol•L-1, respectively. The molecular docking results showed that AcerASP2 had one elongated pocket-like hydrophobic cavity, 4-allylveratrole just existed in the cavity, and two hydrogen bonds were found with Lys74 of AcerASP2. 【Conclusion】Because of the particular spatial structure of binding cavity, AcerASP2 could bind diverse odors with different structures, and the binding interaction could be easily to be promoted by the hydrogen bonds between odors with lysines in AcerASP2.