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应用光谱法观测酶的结构变化,紫外差光谱显示在233nm有一较明显的差吸收负峰,在286nm、300nm左右有不大的负峰和在265nm的正峰;荧光谱显示327nm的相对荧光强度随SDS浓度的增高而递减;发射峰位置红移,酶受SDS>4.1mmol/1作用1h,可引起310nm新荧光发射肩。动力学资料表明,SDS对酶的变性过程为一级反应;酶的失活分快慢相,呈现二个一级反应,同时测定在SDS作用下actinidin的变性与失活速度,表明酶的失活速度大于变性速度,似可认为低浓度SDS能使结构域相错开,从而敏感地改变活性部位的微区结构,导致酶快速失活,提高SDS浓度则使酶的结构域发生构象变化,引起光谱法所显示的特征。
The changes of the structure of the enzyme were observed by spectrophotometry. The UV spectrum showed a significant negative absorption peak at 233 nm, a small negative peak at 286 nm and 300 nm and a positive peak at 265 nm. The fluorescence spectrum showed the relative fluorescence intensity at 327 nm Decreased with the increase of SDS concentration. The emission peak shifted to red, and the enzyme was exposed to SDS> 4.1 mmol / l for 1 h, which caused a new fluorescence emission shoulder of 310 nm. Kinetic data show that SDS denaturation process for the first-order reaction; enzyme inactivation of fast and slow phase, showing two first-order reaction, and determination of actinidin degeneration and deactivation rate in the SDS, indicating that the enzyme inactivation The rate of denaturation is higher than that of denaturation. It seems that low concentrations of SDS can make the domains staggered, thus sensitively changing the micro-structure of the active site, resulting in rapid inactivation of the enzyme. Increasing the concentration of SDS causes the conformational changes of the enzyme domain, Law shows the characteristics.