다양한 나노fiber를 활용한 효소의 고정화 및 재사용

Abstract

효소는 종종 재사용 및 복잡한 회수와 장기간의 작동 안정성 부족으로 인해 방해를 받고 있다. 이러한 단점은 효소의 고정화에 의해 극복 될 수 있다. 이 연구에서, OPH, LDC, GAD 그리고 ω-TA 효소를 foam-PANI, foam-PAMP, foam-PDA 그리고 foam-TA 담체에 고정화하여 높은 안정성을 보였다. 3개월 동안의 연속적인 안정성 측정을 통해서, 고정화 된 효소 foam-PANI-, foam-PAMP-, foam-PDA- 그리고 foam-TA-OPH는 여전히 각각 초기 활성의 87 %, 79 %，62% 그리고 83 %를 유지하였고, 70시간의 연속적인 반응 후에도 70 %, 72 %, 63 % 그리고 78 %의 전율이 유지되었다. LDC을 고정화 한 담체의 경우, 3개월 동안의 연속적인 안정성 측정에서, 초기 활성의 0 ~ 6 %만 감소하였고, 80시간동안 연속반응 후에, foam-PANI-, foam-PAMP-, foam-PDA- 그리고 foam-TA-LDC는 여전히 83 %, 80 %, 64 % 및 75 %의 전환율을 유지하였다. 고정화 효소 GAD의 연속적인 3개월 안정성 측정에서, 고정화 효소 foam-PANI-, foam -PAMP-, foam-PDA- 그리고 foam-TA-GAD는 여전히 초기 활성의 89 %, 88 %, 42 % 그리고 81 %을 유지하였다. 연속 생산반응 중에 H+의 축적으로 효소 활성에 영향을 미치기 때문에 30 시간 후에 생 촉매반응이 감소했다. 이 실험에서 사용된 ω-TA 생성 균주는 야생 균주이며, 효소활성이 낮음에도 불구하고 10회 연속반응 측정에서 고정화 효소 foam-PANI-, foam-PAMP-, foam-PDA- 그리고 foam-TA-(ω-TA)는 42 %, 28 %, 22 %, 및 41 %의 전환율을 꾸준히 유지하였다. 4 가지 재료인 foam-PANI, foam-PAMP, foam-PDA 그리고 foam-TA는 효소 OPH, LDC, GAD 그리고 ω-TA의 고정화 담체로 사용할 수 있음을 확인하였다. 고정화 효소는 생 촉매반응에서 재사용 할 수 있다. |Enzymes are often hampered by their reuse and cumbersome recovery and by lack of their long-term operational stability. These drawbacks can be overcome by immobilization of enzymes. Therefore, four different polymer nanofibers, foam-polyaniline nanofiber (foam-PANI), foam-magnetically separable polyaniline nanofiber (foam-PAMP), foam-polydopamine nanospheres (foam-PDA), and foam-tannic acid & FeCl3 (foam-TA) have been used for the surface coating of nickel foams and they have used for the immobilization of organophosphorus hydrolase (OPH), lysine decarboxylase (LDC), glutamate decarboxylase (GAD) and ω-transaminases (ω-TA). These four polymers have shown excellent properties. 1. Immobilization of OPH onto nickel foam coated with PANI, PAMP, PDA, and TA in the following ways: by cross-linking randomly OPH to polymer nanofibers-coated nickel foam using glutaraldehyde. After 70 cycles reuse of foam-PANI-, foam-PAMP-, foam-PDA-, and foam-TA-OPH under relevant process conditions, they showed high stability and maintained high conversion (70 %, 72 %, 63 %, and 78 %). The relative activities of foam-PANI-, foam-PAMP-, foam-PDA-, and foam-TA-OPH were 87 %, 79 %, 62 %, and 83 % of the initial activity retained, respectively after 3 months of vigorous shaking conditions at room temperature (25℃). The OPH immobilization onto four polymer nanofibers provides a practical method for the detoxification of organophosphate pesticides. 2. Cadaverine was produced by decarboxylation of lysine using LDC. Foam-PANI, foam-PAMP, foam-PDA, and foam-TA have been used as LDC immobilization carriers for stabilization and recycle. The free LDC lost 40 % of the its initial activity after 3 months under vigorous shaking conditions at room temperature, but the immobilized LDCs on foam-PANI, foam-PAMP, foam-PDA, and foam-TA lost only 0 to 6 % their initial activity. The immobilized LDCs with foam-PANI, foam-PAMP, foam-PDA, and foam-TA showed a high percentage of conversion (83 %, 80 %, 64 %, and 75 %) after 80 reaction cycles reuse. 3. The immobilized GADs on foam-PANI, foam-PAMP, foam-PDA, and foam-TA have been used for the production of gamma-aminobutyric acid (GABA) from glutamate. The immobilization yields after immobilization onto four different carriers, foam-PANI, foam-PAMP, foam-PDA, and foam-TA were 67 %, 70 %, 69 %, and 74 %, respectively. 62 %, 58 %, 47 %, and 54 % of monosodium glutamate (MSG) were converted to GABA after 48 h reaction. The immobilized GADs retained 89 %, 88 %, 42 %, and 81 % of their initial activities after 3 months of vigorous shaking conditions at room temperature. 4. ω-TA is one of important biocatalyst and is widely used for the production of chiral amines. The immobilized ω-TAs on foam-PANI, foam-PAMP, foam-PDA, and foam-TA were prepared and used to enhance the stability and recyclability. The immobilized ω-TAs on foam-PANI, foam-PAMP, foam-PDA, and foam-TA retained 42 %, 28 %, 22 %, and 41 % of their initial conversion after 10 cycles. Immobilized ω-TAs on foam-PANI- and foam-TA-(ω-TA) showed 97 % and 85 % relative activity, respectively.