基于三维定量构效关系的计算机辅助设计环糊精谷胱甘肽过氧化物模拟酶的研究

Glutathione peroxidase is an important mammalian selenoenzyme that function in cellular redox reactions and plays an essential role in the detoxification of hydroperoxides in vivo. Fabrication of GPX models offers an ideal alternative for elucidating the origin of substrate binding and catalysis of enzyme. Mimicking the natures of molecular recognition and catalysis of enzymes by artificial enzymes is very essential for exploring the evolved biological process of enzymes as well as their properties of structures and functions. On the basis of structure and.catalytic mechanism understanding for GPX, we choose cyclodextrins as the scaffolds of enzyme models to establish the systems of GPX models. QSAR analysis was performed on a series of cyclodextrin GPX models. The best model with the most suitable molecule orientation was obtained by using all-space search. To understand the hypostasis of structures and catalysis of natural GPX and mechanism of molecular recognition in enzymatic two-substrate reaction, we design and construct direct assays systems to investigate cyclodextrin GPX mimics. We also attempt to elucidate the relationships between substrate specificity and catalytic capacity of cyclodextrin GPX models through establishing the assay systems using structurally distinct substrates. Through studying the relationships between substrate recognition and catalysis of GPX mimics, we provide some useful information on the catalysis of enzymes. Our work not only perfects the development of the research areas of cyclodextrins, but also promotes the understanding of catalytic properties of natural enzymes.

谷胱甘肽过氧化物酶（GPX）是构成生物体抗氧化防御体系的三大酶之一，对GPX的人工模拟不仅能对天然GPX的结构和催化功能提供重要信息，而且还可以开发抗氧化药物，具有重大的理论与实际应用价值。已有的研究表明，环糊精GPX模拟物展现了较高的催化活性，但其高活性的来源并不清楚，急需全面系统的研究模拟物活性与结构的关系，为构建更高活性GPX模拟酶提供理论指导。本项目拟通过计算机辅助设计的方法构建高活性环糊精GPX酶模拟物体系。首先，通过分子模拟、分子动力学等计算化学方法，建立环糊精GPX模拟物活性与结构的三维定量构效关系模型；其次，通过计算机辅助对模拟酶结构进行分子再设计，构建环糊精GPX模拟酶库来筛选高活性的酶模型；最后，基于环糊精选择性修饰策略，进行高活性环糊精GPX模拟酶的制备、表征与活性验证。本项目的顺利实施，以期为完善GPX模拟酶体系的合理设计提供重要的酶模拟信息。

谷胱甘肽过氧化物酶是构成生物体抗氧化防御体系的三大酶之一，对GPX的人工模拟，不仅能对天然GPX的结构和催化功能提供重要信息，而且还可以开发抗氧化药物，具有重大的理论与实际应用价值。本项目运用计算化学的理论与方法，开展对GPX模拟物的分子设计工作。通过计算机辅助设计构建GPX模拟酶库，筛选天然酶活性级别的GPX模拟酶。在此基础上，以高活性GPX模拟酶作为研究平台，通过建立模拟酶的催化能力与分子识别评测体系，探究GPX模型的底物识别与催化作用的关系，进一步阐释酶实施催化反应的循环机制。最后，拟通过建立线粒体的Fe2+/Vc诱导损伤体系，研究GPX模拟物的抗氧化活性。经过三年的研究，我们合成了8种新型的具有GPX催化活性的环糊精GPX模拟物：2-Te-α-CD、2-Te-γ-CD、6-Te-γ-CD、2-Se-γ-CD、6-Se-γ-CD、6-diSe -γ-CD、6-diTe-γ-CD和6-disecys-γ-CD，其中6-diTe -γ-CD表现了目前报道环糊精GPX模拟物酶中最高的生物酶活；我们从环糊精空腔大小角度考察酶-底物尺寸匹配效应在催化过程中的作用，发现了γ-环糊精作为具有底物结合部位的受体在构建GPX模拟酶方面较β-环糊精更具优势；另外，我们还成功利用了ESI-MS/MS技术成功捕捉环糊精GPX模拟物——2-Te-γ-CD催化GSH还原氢过氧化物反应过程中的关键中间体——碲醇、次碲酸和碲硫化合物，揭示了2-Te -γ-CD催化GSH还原过氧化物反应的催化循环机制。