一株抗JEV单抗可变区的计算机建模及其与JEVE蛋白的分子对接

Computerized modeling of variable regions of an anti-JEV monoclonal antibody and their molecular docking with JEV E protein

目的利用计算机建模获得单克隆抗体2F2可变区三维结构,再与日本脑炎病毒(JEV)E蛋白进行模拟分子对接,了解JEV E蛋白上与2F2作用的关键氨基酸位点,为阐明2F2的中和作用机制奠定基础。方法利用BLASTP软件在蛋白质数据库(PDB)中搜索2F2同源蛋白。以同源蛋白为模板,用Modeller软件对2F2可变区主链、侧链进行建模,组装出完整的2F2可变区三维结构模型并进行一系列分子动力学优化,得到能量最低的单抗可变区三维结构。最后利用Discovery Studio软件模拟2F2可变区与JEV E蛋白的分子对接。结果获得2F2可变区三维结构模型。模拟分子对接结果显示JEV E蛋白上与2F2可变区相互接触、作用的氨基酸位点有9个,即Ⅰ区的13位谷氨酸、16位丝氨酸、37位天冬氨酸和300位苏氨酸,Ⅲ区的336位赖氨酸、347位天冬氨酸、354位亮氨酸、387位精氨酸和388位甘氨酸残基。其中Ⅰ区13位谷氨酸、16位丝氨酸和37位天冬氨酸可能是中和表位必需的氨基酸。Ⅲ区的387位精氨酸和388位甘氨酸位于优势中和抗原表位区域,336位赖氨酸与已报道的337位中和表位非常接近。结论通过计算机建模确定JEV E蛋白上有9个与单克隆抗体2F2作用的主要氨基酸位点,为利用E蛋白突变体阐明2F2单抗的中和机制提供了依据。

Objectives To model the three-dimensional structure of the variable regions of anti-JEV monoclonal antibody 2F2 and to identify amino acid residues that are pivotal to neutralization via molecular docking with conformations of the JEV E glycoprotein. Methods Previously obtained sequences of the variable regions of the 2F2 McAb were analyzed with BLASTP, and the PDB database was searched for proteins with similar variable regions. The three-dimensional structure of variable regions of the 2F2 MeAb with the lowest energy was determined using MODELLER 9vl, and molec- ular docking of variable regions of the 2F2 McAb and JEV E protein was determined using DISCOVERY STUDIO v2.1 following energy minimization and thermodynamic optimization of the antigen-antibody complex models that were created. Results The three-dimensional structure of the variable regions of the 2F2 McAb with the lowest energy was selected and optimized, and docking with the E protein was modeled. The results of modeled docking revealed that nine unreported amino acid residues of the JEV E protein came in contact with 2F2, as defined by hydrogen bonds. These amino acid resi- dues are 13Glu, 16Ser, 37Asp, 300Thr （in domain I）, 336Lys, 347Asp, 354Leu, 387Arg, and 388Gly （in domain III）. 13Glu and 16Ser are adjacent to the reported neutralizing epitope 18Ala, suggesting that these residues may be essential to neutralization within domain I. In domain III, 387Arg and 388Gly lie in the known neutralizing epitope-rich area 373-- 399, and 336Lys is adjacent to the known 337Ile as well, whereas 354Leu has never been reported. Conclusion The nine crucial amino acid residues of JEV E protein that were recognized by 2F2 will help to elucidate the mechanism by which 2F2 neutralizes JEV infection.