三唑磷农药降解酶的定位及酶学性质研究

Distribution and properties of triazophos-degrading enzyme

采用超声波破碎法破碎三唑磷农药降解菌芽孢杆菌TAP-1,研究不同细胞组分对三唑磷农药的降解率,应用液相色谱法研究三唑磷农药降解酶的酶学性质。结果表明,胞外酶、胞间粗酶液和胞内酶对三唑磷的降解率分别为2.8(±0.34)%、12.8(±2.44)%和84.4(±6.71)%。菌株TAP-1降解三唑磷农药的主要活性物质位于降解菌细胞内,属于胞内酶组分。该酶最适作用pH值为6.5~7.0,在pH 5.0~7.5之间,酶活力均能保持在最高酶活力的68%以上。最适反应温度为30℃,在25~40℃温度范围内能保持75%以上的降解活性。不同化学试剂和金属离子对酶活性产生不同的影响。巯基修饰剂、EDTA、Hg^(2+)和Mn^(2+)对酶活性有抑制作用。Mg^(2+)、Co^(2+)和Fe^(3+)对酶活性有促进作用。该降解酶米氏常数(Km)为23.02μmol/L,最大反应速度(Vmax)为0.738μmol/mg·min。

To investigate the effects of different fractions of bacterial strain Bacillus TAP-1 on the degradation rate of tirazophos, sonication method was applied to lyse the TAP-1 cells to obtain triazophos-degrading enzyme, while HPLC （ High Performance Liquid Chromatography ） method was used to determine the enzymatic properties. The results showed that the degradation rate of triazophos by intracellular, intercellular and extracellular extracts were 84.4 （ ± 6.71 ） %, 12.8 （ ±2.44 ） %, and 2.8 （ ± 0.34 ） %, respectively. The active substances of triazophos-degrading were intracellular fractions. The enzyme demonstrated greatest enzymatic activity in the pH range of 6.5-7.0 with its highest activity occurring in the pH range of 5.0-7.5. Enzymatic activity occurred at an optimum temperature of 30℃ for the degradation of triazophos ; activity remained above 75% of the maximum over a temperature range of 25℃-40℃. Different chemicals and metallic ions also had different effects on its activity. Its activity can be strongly inhibited by thiol modifier, ETDA, Hg2＋ and Mn2＋, but stimulated by Mg2＋, Co2＋ and Fe3＋. The Kin value and the maximal degradation rate of triazophos-hydrolase were 23.02 μmol/L and 0.738μmol/mg ·min, respectively.