荧光素酶nanoKAZ与发光底物的分子对接及对接复合体的全原子动力学模拟

The docking of a luciferase nanoKAZ with its substrate and the molecular dynamics simulation of the docked complex

nanoKAZ是一种体量小且底物专一性低的新型荧光素酶,具有高潜在应用价值,然而nanoKAZ与底物的作用方式未见报道.这里使用分子对接技术构建了nanoKAZ与底物coelenterazine结合后形成的复合体构象,使用软件GROMACS对该复合体进行常温水溶液态的全原子动力学模拟并综合模拟过程中的酶主链运动特性,酶与底物间结合自由能变化与形成氢键情况分析nanoKAZ与底物作用方式.结果显示coelenterazine进入nanoKAZ空腔后借由与β桶区域内7个氨基酸残基产生强结合能并与其中Q12形成氢键以稳定分子朝向,同时coelentazine还促使催化腔口区域片段V27-V38(V27与coelenterazine形成氢键)和片段P61-H87在空间上彼此靠近从而关闭催化腔,稳定疏水的底物分子以促进催化反应发生.以上结果阐明了底物与nanoKAZ活性腔的结合方式,提出了nanoKAZ-底物反应中间体的构象,为nanoKAZ的理性改造提供了理论基础.

nanoKAZ is a novel luciferase, which has a small molecular size and low coelenterazine specificity therefore it was considered as an ideal reporter protein in bio-imaging. However, the catalysis mechanism of nanoKAZ is still unclear. Here a docking simulation was used for binding a coelenterazine to a nanoKAZ followed with an all-atoms dynamics simulation in water using GROMACS software for the structural and dynamics characterization of the nanoKAZ-coelenterazine complex. As the result, we found that the coelenterazine, which entranced into the putative catalytic cavity of nanoKAZ, is griped by 7 residues on the β-barrel, in which Q12 was further formed a hydrogen bond that held the coelenterazine. Furthermore, coelenterazine gathered and stabled(decrease the flexibility) two fragments L18-F31 and P61-H87, they are located on the cavity entrance of nanoKAZ and normally flexible when coelenterazine absence, resulting the entrance of cavity closed thereby building an entire hydrophobic environment of catalytic cavity facilitating to coelenterazine oxidation. Our finding indicates the binding property between nanoKAZ and coelenterazine, and is significant for the rational molecular redesign of nanoKAZ.