脲酶与矿物-腐殖酸复合物的界面反应及其对酶构象和生物活性的影响

Urease, one of important enzymes participating nitrogen biogeochemical cycle, has high affinity with soil minerals and humic acids. Minerals and humic acids are usually bound with each other as an important adsorbent in soil, affecting substance transportation and fate in the environment. Though the interaction of enzyme with minerals or humic acids has been deeply researched, the interaction mechnism of enzyme with minerals-humic acids complexes existing in soil is not clear. Further, the effect essence of their interaction on enzyme biological activity should be explored in depth. In our studies, Kaolinite-humin acid、Kaolinite-fulvic acid、Montmorillonite-humic acid、Montmorillonite-fulvic acid are applied as representative mineral–organic complexes. On the basis of investigated complex surface properties (e.g. morphology, variable charge and hydrophobicity), attenuated total reflectance infrared spectroscopy (ATR-FTIR) and sum frequency vibrational spectra (SFG) are used to study the change of urease conformation. The effects of surface properties on urease conformation and active sites are specially considered. Combination with adsorbed urease activity and stability, the influence of urease conformation on its biological properties will be elucidated. As a result, this work will be beneficial for the recoganition of urease fate and functions in soil.

脲酶是参与氮素生物地球化学循环的重要酶类之一，与土壤矿物、腐殖酸具有较强的亲和力。土壤中矿物与腐殖酸相互胶结组成重要的吸附载体，影响环境物质的迁移与归宿。目前对矿物、腐殖酸与酶的作用特征与机制已有深入的研究，而更逼近真实土壤的矿物-腐殖酸复合物与脲酶的作用机制及其对酶生物活性的影响机制尚不明确。本项目选择高岭石-胡敏酸、高岭石-富里酸、蒙脱石-胡敏酸、蒙脱石-富里酸等几种典型的矿物-有机质复合物，在分析复合物形貌、电荷量、疏水性等表面特性的基础上，采用衰减全反射红外光谱（ATR-FTIR）及和频振动光谱（SFG）等技术研究吸附态脲酶构象的变化特征，明确不同复合物表面特性对脲酶构象及活性中心结构的影响；结合吸附态脲酶活性、稳定性等生物特性的变化规律，阐明脲酶构象变化对其生物特性的影响机制，为理解脲酶在土壤中的生物功能和环境行为提供科学理论依据。

土壤酶在氮、磷等营养元素地球化学循环中起着重要的作用，与土壤矿物、有机质具有较强的亲和力。腐殖酸等有机质可以直接与土壤酶发生交互作用，同时还可影响土壤矿物对其的吸附固定、生物活性和结构特性，并受土壤有机质含量、矿物类型、环境条件等多种因素的影响。本项目选择模式蛋白质溶菌酶、脲酶两种蛋白质，通过腐殖酸对两种酶生物活性、微环境及二级结构的影响，探讨酶活性变化与其结构变化的内在关联性，通过腐殖酸对高岭石、蒙脱石对模式蛋白质吸附固定的影响，揭示有机质对土壤酶在土壤中迁移转化的影响规律。研究结果表明，腐殖酸与溶菌酶的交互作用过程中ɑ-螺旋结构的变化可能是导致其活性发生变化的一个重要原因，蒙脱石对溶菌酶的吸附量约是高岭石的10倍，少量腐殖酸能够促进高岭石、蒙脱石对溶菌酶的吸附，但当pH=5且腐殖酸/高岭石质量比增加至1:50后，溶菌酶在高岭石上的吸附表现出先吸附后解吸的现象，蒙脱石在高pH值条件下表现为对溶菌酶先絮凝后吸附，这些结果为认识酶在土壤中的环境行为及调控其生物功能提供科学依据。