粉尘螨水通道蛋白DerfAQP2分子模拟和自由能计算

Simulation of the molecular dynamics of mite aquaporin DerfAQP2 from the dust mite Dermatophagoides farinae (Acariformes: Pyroglyphidae)

目的探讨粉尘螨水通道蛋白DerfAQP2的工作机制。方法用SWISS-MOEL软件对DerfAQP2进行同源建模,用VMD软件建立分子模拟体系,用NAMD2.9软件包进行动力学模拟并对体系进行优化,等温等压系综环境下计算水分子通过时的吉布斯自由能。结果以大肠埃希菌水甘油通道蛋白(PDB code:1LDF)晶体结构作为模板构建DerfAQP2蛋白质空间构像,使用VMD软件生成psf文件,将DerfAQP2蛋白单聚体放置于80×80×100?~3大小的盒子中,加入TIP3型水分子、55个钠离子、53个氯离子,最终体系共计60 184个原子。经过20 ns的分子动力学模拟,DerfAQP2蛋白结构趋于稳定。取dcd文件进行结构分析,发现8个水分子在通道内构成单列水,当水分子进入的通道外前庭时,DerfAQP2的通道残基由Arg203、Tyr44和Ile188残基构成;在通道的中间部分,可见NPA结构的Asn200和Asn64;在细胞质与通道交界附近,水分子与周边的His62以及Ala61、Ile63、Ser52残基间相互作用。将单列水起始位置的能量定义为0,绘制自由能曲线,可见水分子通过通道的能量屏障约为2.0 kcal/mol。结论成功建立了粉尘螨水通道蛋白DerfAQP2的三维空间结构和分子模拟体系,解析了其运输水分子的关键氨基酸。

Objective To examine the mechanism of action of mite aquaporin DerfAQP2 from the dust mite Dermatophagoides farinae(Acariformes:Pyroglyphidae).Methods A homology structural model of DerfAQP2 was constructed using the software Swiss-model,and a molecular dynamics(MD)simulation system was created using the software Visual Molecular Dynamics(VMD).NAMD 2.9 was used to simulate the dynamics and to optimize the MD system.Gibbs free energy was calculated for water transport through the channel at a constant temperature of 298 K and a constant pressure of 1 atm(NPT ensemble).Results A spatial structure was constructed via homology modelling of DerfAQP2 from the X-ray structure of E.coli glycerol facilitator GLPF(PDB code No.1 LDF).A psf file was generated by the software VMD.The monomer DerfAQP2 was placed in a box with 80×80×100?3.TIP3 water molecules,55 Na^+,and 53 Cl^-were added,for 60,184 atoms in total.After simulation for 20 ns,the conformation of DerfAQP2 stabilized.The dcd file for the 20 ns simulation was used in structural analysis.Eight water molecules passed through the pore in a single file.When water molecules entered the extracellular vestibule of the pore,the pore residues were Arg203,Tyr44,and Ile188.In the middle of the channel,the two conserved Asn-Pro-Ala(NPA)motifs were found in the residues Asn-200 and Asn-64.Near the junction of the cytoplasm and the channel,the water molecules interacted with the residues His62,Ala61,Ile63,and Ser52.The overall free energy profile for DerfAQP2 water transport was determined using 0 for the energy at the starting point,when water molecules formed a single file.This yielded an energy barrier of approximately 2.0 kcal/mol.Conclusion The three-dimensional spatial structure and an MD simulation system were successfully constructed for DerfAQP2,and key residues for water molecules passing through DerfAQP1 aquaporin were determined.