| 摘要: | 本實驗室於 2015 年利用原位創新合成法 (de novo approach) ，成功以類沸石咪唑骨架材料-90 (Zeolitic Imidazolate Framework-90；ZIF-90) 包覆酵素。ZIF-90 提供內部酵素孔洞篩選性，使小於窗口大小之反應物 (Substrate) 能進入材料供酵素催化，而大於窗口之蛋白質水解?-K 則無法通過並水解酵素。此外，ZIF-90 還可提供空間侷限性質，防止酵素在高熱或具有尿素環境下展開，使酵素能夠在嚴苛條件下進行催化反應。然而，酵素固定化往往使材料與酵素間產生相互作用，使酵素脫離原始狀態，造成活性下降；因此，本實驗室於2020年將酵素包覆於 MOFs 的不同種形態──中空金屬有機骨架材料 (Hollow MOFs；HMOFs) 中。因其擁有內部空腔，酵素結構得以獲得足夠空間，雖然會失去空間侷限性的優點，但 HMOFs 不僅改善酵素被固定後所造成的活性下降問題，且仍保有 MOFs 孔洞篩選的特性。但原先的在蝕刻步驟需要較長時間 (20 hr)，就固定酵素而言，若能縮短合成時間，則可以降低酵素的自然崩解對催化效率的影響；本實驗採用兩種不同的蝕刻方法，分別為添加蝕刻劑單寧酸崩解而得的 enzyme@HZIF-90-ZnTA，以及運用 MOFs 對氫離子配位能力差異，崩解 enzyme@ZIF-67@ZIF-8 內層ZIF-67所合成的 enzyme@HZIF-8，兩者皆能快速完成蝕刻步驟，以降低酵素自然降解的影響，使 HMOFs 於酵素固定化上有更良好的活性表現。;In 2015, our laboratory successfully embeded enzymes in zeolitic imidazolate framework-90 (ZIF-90), a sub group of Metal-organic Frameworks (MOF), via mild water-based de novo approach. ZIF-90 provides internal enzyme size-selective sheltering which makes substrates smaller than the aperture delivery into MOF materials for enzymatic catalysis. By contrast, larger proteins, like proteinase K, which is an enzyme that catalyzes proteolysis to digest proteins into smaller polypeptides or single amino acids, cannot pass through to digest the enzyme. In addition, ZIF-90 can also provide shielding confinement properties against enzyme unfolding under high temperatures or urea environments. Thus, the enzyme can remain its functions under severe conditions. However, there are always interactions between the material and the enzyme after the enzyme immobilization process, which alters the enzymes from their native states, resulting in the decrease in activity. To solve this problem, in 2020, our laboratory embed enzymes in one of different forms of MOFs, which calls hollow metal-organic frameworks (HMOFs). Everything has its pros and cons. In advantages, using HMOFs not only improves the problem of decreased activity caused by the enzyme immobilization but also remains the characteristic of MOFs size-selective sheltering. Although it will lose the shielding confinement properties, the enzyme structure can have more space due to HMOFs with more freedom flexibility. In disadvantages, the original etching steps (hollowing steps) take longer time for about 20 hours. For enzymes, the less synthesis time, the less catalytic efficiency caused by nature decay will be reduced.
Therefore, we use two different approaches to accelerate synthetic processes. One is to add tannic acid as an etching agent to form the enzyme@HZIF -90-ZnTA, and the other one is to decompose the inner ZIF-67 structure of enzyme @ ZIF-67 @ ZIF-8 by different coordination abilities of the hydrogen ion between two MOFs to synthesize enzyme@HZIF-8. Both of them can quickly complete the etching steps to reduce the impact of natural enzyme decay, so that HMOFs can have the better activity performance on enzyme immobilization. |
