Zhejiang-Malaysia Joint Research Laboratory for Agricultural Product Processing and Nutrition, College of Food and Pharmaceutical Sciences, Ningbo University, Ningbo 315211, China.
School of Agriculture, Food and Ecosystem Sciences, Faculty of Science, University of Melbourne, Melbourne 3010, Australia.
ACS Appl Mater Interfaces. 2024 Aug 7;16(31):40836-40847. doi: 10.1021/acsami.4c05208. Epub 2024 Jul 25.
The stimulus-responsive regulation of enzyme catalytic activity and selectivity provides a new opportunity to extend the functionality and efficiency of immobilized enzymes. This work aims to design and synthesize a thermo-switchable enzyme@MOF for size-selective biocatalysis and biosensing through the immobilization of lipase (CRL) within ZIF-8 functionalized with thermally responsive polymer, poly(-isopropylacrylamide) (PNIPAM) (CRL@ZIF-8-PNIPAM). Unlike free CRL, which does not demonstrate substrate selectivity, we can reversibly tune the pore size of the ZIF-8-PNIPAM nanostructures (open pores or blocked pores) through temperature stimulus and subsequently modulate the substrate selectivity of CRL@ZIF-8-PNIPAM. CRL@ZIF-8-PNIPAM had the highest hydrolytic activity for small molecules (12 mM -nitrophenol/mg protein/min, 4-nitrophenyl butyrate (-NP Be)) and the lowest hydrolytic activity for large molecules (0.16 mM p-nitrophenol/mg protein/min, 4-nitrophenyl palmitate (-NP P)). In addition, CRL@ZIF-8-PNIPAM demonstrated thermo-switchable behavior for large molecules (-NP P). The -NP P hydrolytic activity of CRL@ZIF-8-PNIPAM was significantly lower at 40 °C (blocked pores) than at 27 °C (open pores). However, the transition of blocked pores and open pores is a gradual process that resulted in a delay in the "thermo-switchable" catalytic behavior of CRL@ZIF-8-PNIPAM during thermal cycling. CRL@ZIF-8-PNIPAM was also successfully used for the fabrication of electrochemical biosensors for the selective biosensing of pesticides with different molecular sizes.
通过将热响应聚合物聚（异丙基丙烯酰胺）（PNIPAM）功能化的 ZIF-8 内固定化脂肪酶（CRL），设计并合成了用于尺寸选择性生物催化和生物传感的热开关酶@MOF（CRL@ZIF-8-PNIPAM）。与没有表现出底物选择性的游离 CRL 不同，我们可以通过温度刺激来可逆地调节 ZIF-8-PNIPAM 纳米结构的孔径（打开的孔或封闭的孔），并随后调节 CRL@ZIF-8-PNIPAM 的底物选择性。CRL@ZIF-8-PNIPAM 对小分子（12 mM -硝基苯酚/毫克蛋白/分钟，4-硝基苯丁酸（-NP Be））具有最高的水解活性，对大分子（0.16 mM 对硝基苯酚/毫克蛋白/分钟，4-硝基苯棕榈酸（-NP P））具有最低的水解活性。此外，CRL@ZIF-8-PNIPAM 对大分子（-NP P）表现出热开关行为。与 27°C（打开的孔）相比，CRL@ZIF-8-PNIPAM 在 40°C（封闭的孔）下的 -NP P 水解活性显著降低。然而，封闭孔和打开孔的转变是一个逐渐的过程，导致 CRL@ZIF-8-PNIPAM 在热循环过程中“热开关”催化行为出现延迟。CRL@ZIF-8-PNIPAM 还成功地用于制备电化学生物传感器，用于选择性地检测不同分子大小的农药。
