表面等离子体子共振传感器研究谷胱甘肽过氧化物酶模拟物与底物的结合作用

Study of Interaction between Glutathione Peroxidase Mimics and Substrates by Surface Plasmon Resonance Biosensor

采用表面等离子体子共振(SPR)传感装置,固定入射角,以波长为变量,以CCD为检测系统,用对金有较强吸附作用的谷胱甘肽(GSH)为基底膜,研究了GSH分别与谷胱甘肽硫转移酶(GST)与小分子GPX模拟酶2-位-碲桥联-β-环糊精(2-TeCD)的动力学过程,计算得GSH与GST的结合常数为2.82×106L/mol;GSH与2-TeCD的结合常数为3.92×102L/mol。结果表明:本方法可以用来评价小分子GPX模拟酶与底物的结合程度,简便快捷,不需标记样品,可以实时监测反应过程,作用后酶与底物易于分离,分离后的酶可以继续使用等优点。

The surface plasmon resonance （SPR） biosensor was designed on the basis of fixing the angle of incidence and measuring the reflected intensities. A charge coupled device （CCD） was used to measure the SPR spectra. Glutathione （GSH） self-assembling in solution was used to form the sensing membrane on the gold substrate. Interaction of glutathione-S-transferase （GST） and 2, 2-ditelluro bis-（2-deoxy-cyclodextrin） （2-TeCD） with substrate GSH were studied by this SPR sensor and the kinetic processes of sensing monolayer formation were studied. The kinetic constants of GSH and GST was 2. 82 × 10^6 L/mol, GSH and 2-TeCD was 3.92 × 10^2 L/mol. These results validate the SPR sensor technology can be used to study interaction of the small-molecule glutathione peroxidase （GPX） mimics and substrate GSH in real time. Under the experimental conditions, the sensor is a labeling free, simultaneous observation of the interaction among enzyme and substrates can be realized and, therefore, kinetic constants of reactions can be obtained. Such instrumentation is to be used in fields such as on-line process monitoring.