荧光假单胞杆菌磷脂酶B界面催化识别机制及其定向调控规律研究

To make clear the reason that there was a shiftting of the catalytic selectivity of the phospholipase B from Pseudomonas fluorescens BIT-18 when used in the Degumming of vegetable oils, it's needed to research the Mechanism of molecular recognition. After prepared, purified and crystallizing, high-quality Pf-PLB monocrystal and enzymesubstrate complex was obtained, and Then their three-dimensional structurethe was inspected, and the mode substrate of Pf-PLB was prepared. After Investigating the change of crystal structure of Pf-PLB monocrystal and its crystalloid enzymesubstrate complex with the change of different external factor, the relation between the Structure and Function of Pf-PLB was summarized. And then, what shiftting the catalytic selectivity of Pf-PLB was Known by analyzing the polarity of substrate binding region, comformation character and thermodynamicpr inciples. The molecular mechanism, catalytic recognition and directed control during the interfacial catalyzing of Pf-PLB were clarified, which lay the foundation of phospholipase industrial and directed Molecular Modification of enzyme.

针对利用荧光假单胞杆菌磷脂酶B（Pf-PLB）在油水界面对植物油进行脱胶时表现出的催化选择性迁移现象，拟开展Pf-PLB对底物分子识别机制与定向调控研究。首先制备Pf-PLB粗酶液，分离纯化后制备高质量Pf-PLB单晶和Pf-PLB-底物复合物结晶，并对其三维结构进行解析，同时制备其模式底物；然后通过考察不同条件下的Pf-PLB单晶、Pf-PLB-底物复合物结晶的结构变化，分析各种外因对Pf-PLB催化特异性的影响，总结出Pf-PLB结构与功能之间的关系；最后根据底物结合区的极性、构象特征和热力学原理等探讨Pf-PLB催化选择性迁移的原因，进而从分子水平揭示Pf-PLB界面催化识别的分子机制和定向调控规律，为磷脂酶工业和酶分子改造提供理论依据。

针对荧光假单胞杆菌磷脂酶B（Pf-PLB）在油水界面对植物油进行脱胶时表现出的催化选择性迁移现象，对Pf-PLB与底物分子的识别机制与定向调控规律进行了研究。首先制备了Pf-PLB粗酶液，分离纯化、结晶后得到高质量Pf-PLB单晶，制备了Pf-PLB-底物复合物结晶，并对其三维结构进行了解析，并制备了模式底物；然后通过考察不同条件下的Pf-PLB单晶、Pf-PLB-底物复合物结晶的结构变化，分析各种外因对Pf-PLB催化特异性的影响，总结出Pf-PLB结构与功能之间的关系；最后根据底物结合区的构象特征探讨Pf-PLB催化选择性迁移的原因，从分子水平上揭示了Pf-PLB界面催化识别的分子机制，探讨了选择性定向调控规律，为磷脂酶工业化推广应用和酶分子改造提供了基础依据。