Green surface modification of nylon membrane to site-specifically co-immobilize xylanase and lichenase for clean production of reducing sugar and juice clarification

Abstract

The scarcity of simple and green functionalization protocols for carriers and the high purification cost of enzymes greatly hindered the application of multi-enzymes. Herein, a universal green and mild (25 °C, 1 h) carrier modification strategy for targeted immobilization of multi-enzymes without complex purification was proposed, in which only biocompatible tannic acid and Elastin-like polypeptides-SpyCatcher (ELPs-SC) were fed. The biofunctionalized nylon membranes (ELPs-SC-NMs) exhibited excellent stability and only 8.5% loss after being incubated at 4 °C for 15 days. Then, ELPs-SC-NMs were adopted to site-specifically immobilize target multi-enzymes (xylanase-SpyTag-lichenase, XTL) directly from the crude solution with the load of 46.2 μg/cm². The retention of the XTL was up to 95.2% after 6 days of incubation at 4 °C, and the activity of xylanase in XTL was still 79.8% of the initial value after 18 days of storage, which was increased by 278.2% compared with free xylanase. In addition, the covalent immobilized XTL showed excellent reusability, which remained 74.7% (xylanase) and 77.5% (lichenase) of the initial activity after 8 uses, respectively. Moreover, the immobilized multi-enzymes were applied for clean production of reducing sugars from biomass, the substrate conversion rate and total reducing sugar content were close to those of free enzymes. They also exhibited more excellent catalytic properties than free ones in the clarification of fruit juices. The universality of our strategy was also demonstrated on poly(vinylidene fluoride) membranes. The green site-specific covalent immobilization avoids the complex enzyme purification process and significantly saves the cost, which will shield light both on co-immobilized multi-enzymes for cleaning biocatalytic applications and mild bio-based material surface functionalization.