树脂固定化海洋脂肪酶ADM47601的研究

Research on immobilization of lipase ADM47601 from marine microorganisms by resin

以海洋脂肪酶ADM47601为研究对象,以树脂为载体,进行了固定化酶的研究。筛选了21种树脂,确定D316型阴离子交换树脂为载体。通过正交试验得到了固定化的最佳条件,每克载体加脂肪酶10 000 U,pH7.5,25℃,150 r/min,固定化时间3 h,固定化酶活力回收率可达65.53%±1.06%。固定化脂肪酶最适作用pH为7.5,在pH6.0~9.0的范围内能够保持85%的活力。最适作用温度40℃,在50℃下贮存1 h后依然保持66%的活力。25℃保存半衰期为55 d,操作8次仍然能维持60%以上的活性,且在多种有机试剂中能够稳定的保持活性。固定化脂肪酶Km和Vmax值分别为4.86×10^(–5) mol/L和1.31×10^(–5) mol/(L·min),对底物亲和性较好。

The effect of different resins on immobilizing lipase ADM47601 was investigated in this study. A total of 21 types of resins were screened, and the anion-exchange resin D316 demonstrated the best immobilization. After the orthogonal test, the recovery ratio of enzyme activity reached up to 65.53% ± 1.06% under optimal conditions： 10000U lipase per 1g resin, pH7.5, 25℃, 150r/min and 3 h for immobilization. The optimal pH of immobilized lipase was 7.5, and it exhibited 85% of the maximum activity in the range of pH6.0–9.0. The optimal temperature was 40℃, and the activity was maintained at 66% after 1 h of storage at 50℃. At 25℃, the half-life was 55 days and the residual activity was still higher than 60% after utilizing the lipase 8 times. The immobilized lipase can maintain activity in various organic reagents, and its Km and Vmax values were 4.86 × 10^–5mol/L and 1.31 × 10^–5mol/（L·min）, respectively, indicating high affinity to the substrate.