有机磷水解酶纳米胶囊的调控及其对酶活性和稳定性的影响  

Regulation of Organophosphorus Hydrolase Nanocapsules and the Effect on Enzyme Activity and Stability

作　　者：涂诚昭 雷美玲 叶霖[1,3] TU Cheng-Zhao;LEI Mei-Ling;YE Lin(School of Materials Science and Engineering,Beijing Institute of Technology,Beijing 100081,China;State Key Laboratory of NBC Protection for Civilian,Beijing 102205,China;Beijing Key Laboratory of Construction Tailorable Advanced Functional Materials and Green Applications,Beijing 100081,China)

机构地区：[1]北京理工大学材料学院,北京100081 [2]国民核生化灾害防护国家重点实验室,北京102205 [3]结构可控先进功能材料与绿色应用北京市重点实验室,北京100081

出　　处：《中国生物化学与分子生物学报》2021年第3期372-379,共8页Chinese Journal of Biochemistry and Molecular Biology

摘　　要：纳米胶囊化是提高有机磷水解酶(organic phosphorus hydrolase,OPH)稳定性,进而实现其实用化的最具前景的解决方案。纳米胶囊一方面能够有效保护有机磷水解酶免于失活,但另一方面胶囊的存在也会阻碍底物与酶活性中心的接近。因此,通过调节纳米胶囊的结构来调控有机磷水解酶纳米胶囊活性和稳定性是十分值得研究的课题。本文用N-羟基琥珀酰亚胺丙烯酸酯(N-succinimidyl acrylate,NAS)将有机磷水解酶表面丙烯酰化,以丙烯酰胺(acrylamide,AAM)和N-(3-氨丙基)甲基丙烯酰胺盐酸盐(N-(3-aminopropyl)methacrylamide hydrochloride,APM)为单体,在有机磷水解酶蛋白表面进行原位自由基聚合反应,制备了单分散的有机磷水解酶纳米胶囊(OPH nanocapsule,nOPH)。用透射电镜和傅里叶红外光谱对nOPH结构进行了表征。研究了不同NAS投料量在37℃下对有机磷水解酶纳米胶囊活性,及其热稳定性和有机溶剂稳定性的影响规律。当NAS投料量在75∶1以下时,纳米胶囊的存在对有机磷水解酶活性未见影响,而当大于这一投料比时,酶活性会因胶囊致密程度的上升而下降。在加热下或有机溶剂中,有机磷水解酶纳米胶囊的酶活性呈现出随着NAS投料比的增加先增加而后下降的趋势。以上结果表明,酶活性可以通过在由有机磷水解酶制得有机磷水解酶纳米胶囊的过程中,调节纳米胶囊至合适的致密程度来最大程度的保留。合适的致密程度还能显著提高有机磷水解酶纳米胶囊热稳定性和有机溶剂稳定性。这对进一步实现有机磷水解酶的实用化具有重要意义。Nanocapsuling organophosphorus hydrolase(OPH)is a promising strategy,which can improve OPH stability and put it into practical application.Nanocapsule can protect OPH from deactivation,but on the other hand it may block the contact of enzymes and substrates to some extent.Thus,it is worth exploring the influences of nanocapsule density on the enzyme activity and stability.In this study,OPH surface was modified by N-acryloxysuccinimide(NAS),and then the in situ radical polymerization technique was applied to construct a polymer shell around the surface to form the OPH nanocapsule(nOPH).Transmission electron microscope(TEM)and Fourier Transform Infrared(FTIR)was used to characterize the structure of nOPH.The nOPH activity is not influenced by the presence of nanocapsule when the feed ratio of NAS to OPH is below 75.On the contrary,it decreases with the increasing of NAS feed ratio when it is above 75.Furthermore,nOPH activities at high temperatures and in 20%DMSO solutions both first increase and then decrease with the increasing of NAS feed ratio.The results showed that the appropriate density of the nanocapsule could retain the enzyme activity to the maximum extent during the preparation of nOPH nanocapsule,and significantly improve its thermal stability and organic solvent stability.Hence,the results are of great significance to further realize the OPH practical application.

关 键 词：原位自由基聚合 纳米胶囊 有机磷水解酶 热稳定性 有机溶剂稳定性 

分 类 号：TB34[一般工业技术—材料科学与工程]