Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo, Zhejiang 315211, PR China.
Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo, Zhejiang 315211, PR China.
Int J Biol Macromol. 2018 Oct 1;117:1188-1194. doi: 10.1016/j.ijbiomac.2018.06.041. Epub 2018 Jun 15.
Phospholipase D (PLD) was effectively immobilized on a ZnO nanowires/macroporous SiO composite support through an in-situ cross-linking method. An anionic and long-chained bi-epoxy cross-linker was used by adsorbing on the surface of ZnO nanowires through static interaction before cross-linking. Under the fine control of in-situ cross-linking the immobilized PLD has loading amount as high as 113.7 mg/g, possessing high specific activity from 13,987 to 16,142 U/g in all the range of loading amount. The immobilized PLD showed high activity and stability in catalyzing the conversion of phosphatidylcholine (PC) to phosphatidylserine (PS). The reaction conditions such as loading amount of PLD, substrate molar ratio, temperature, solution pH, and reaction time were optimized for the finding of best synthetic process. Under optimized conditions and the PS yield reached 94.8% within 40 min at 50 °C. The immobilized PLD exhibited not only better thermostability and resistance to pH inactivation than free PLD but also the greatly improved storage stability and reusability. It was found that 81.5% of initial activity retained after incubation at 4 °C for 60 days and that 80.4% of PS yield retained after 13 cycling reuses.
通过原位交联方法，将磷脂酶 D（PLD）有效地固定在 ZnO 纳米线/大孔 SiO 复合材料载体上。通过静电相互作用吸附在 ZnO 纳米线表面，在交联前使用阴离子和长链双环氧交联剂。在原位交联的精细控制下，固定化 PLD 的载量高达 113.7mg/g，在所有载量范围内具有高比活性，从 13987 到 16142U/g。固定化 PLD 在催化磷脂酰胆碱（PC）转化为磷脂酰丝氨酸（PS）方面表现出高活性和稳定性。优化了反应条件，如 PLD 的负载量、底物摩尔比、温度、溶液 pH 值和反应时间，以找到最佳的合成工艺。在优化条件下，在 50°C 下反应 40min，PS 产率达到 94.8%。固定化 PLD 不仅表现出比游离 PLD 更好的热稳定性和抗 pH 失活能力，而且还具有大大提高的储存稳定性和可重复使用性。结果发现，在 4°C 下孵育 60 天后，初始活性保留了 81.5%，在 13 次循环重复使用后，PS 的产率保留了 80.4%。
