保护剂对重组大肠杆菌NMN转移酶稳定性的影响

通过向NMN转移酶( Nmnat)中添加保护剂以提高其热稳定性及储存稳定性,扩大酶的使用范围。研究了固体醇类(山梨醇、甘露醇)、糖类(海藻酸钠、蔗糖、甘露糖)和有机溶剂( DMSO、丙二醇-单甲醚、乙醇、甘油)对Nmnat的稳定性的作用,通过正交实验得到一种复合保护剂,并研究复合保护剂对Nmnat最适反应温度、pH稳定性、储存稳定性的影响。结果表明,山梨醇、海藻酸钠和DMSO能够显著提高Nmnat的热稳定性。复合保护剂配方为山梨醇1.5 g/L,海藻酸钠1.0 g/L, DMSO 0.5%。复合保护剂的添加使Nmnat的最适反应温度从37℃提高到50℃；50℃保温2 h酶活提高了24.5%；pH使用范围由7.0~8.0扩大到6.0~8.0；4℃储存28 d后,酶活保留率提高了15.65%。

Effects of protective agents on stability of NMN adenylyltransferase from Escherichia coli

In order to enhance the stability and storage stability of NMN adenylyltransferase ( Nmnat) ,protective additives were added into the preparation of Nmnat, thereby expanding the scope of the use of enzymes. Some kinds of protective agents:solid alcohols (sorbitol, mannitol), sugar (sodium alginate, sucrose, mannose) and organic solvent (DMSO, propylene glycol-single ether, ethanol, glycerol) were investigated. The optimal combination additive (1. 5 g/L sorbitol, 1. 0 g/L alginate, 0. 5% DMSO) was obtained using the orthogonal experimental design. And the thermal stability, optimum reaction temperature, optimum reaction pH, the storage stability of Nmnat with combination additive were discussed. These additives might increase the optimal reaction temperature from 37 ℃ to about 50 ℃ due to their good protection effects, the activity increased by 24. 5% after preservation for 2 hours at 50 ℃, the range of pH stability from 7 . 0~8 . 0 expanded to 6 . 0~8 . 0 , the residual activity of Nmnat containing combination additive increased 15 . 65% after storing at 4 ℃ for 28 days.