ACS Appl Mater Interfaces. 2019 Nov 27;11(47):44913-44921. doi: 10.1021/acsami.9b16274. Epub 2019 Nov 12.
Exploring a suitable immobilization strategy to improve catalytic efficiency and reusability of cellulase is of great importance to lowering the cost and promoting the industrialization of cellulose-derived bioethanol. In this work, a layered structure with a thin PEG hydrogel as the inner layer and sodium polyacrylate (PAANa) brush as the outer layer was fabricated on low density polyethylene (LDPE) film by visible-light-induced graft polymerization. Two enzymes, β-glucosidase (BG) and cellulase, were separately coimmobilized onto this hierarchical film. As supplementary to cellulase for improving catalytic efficiency, BG was in situ entrapped into the inner PEG hydrogel layer during the graft polymerization from the LDPE surface. After graft polymerization of sodium acrylate on the PEG hydrogel layer was reinitiated, cellulase was covalently attached on the outer PAANa brush layer. Owing to the mild reaction condition (visible-light irradiation and room temperature), the immobilized BG could retain a high activity after the graft polymerization. The immobilization did not alter the optimal pH and temperature of BG or the optimal temperature of cellulase. However, the optimal pH of cellulase shifts to 5.0 after immobilization. Compared with the original activity of single cellulase system and isolated BG/cellulase immobilization system, the dual-enzyme system exhibited 82% and 20% increase in catalytic activity, respectively. The dual-enzyme system could maintain 93% of carboxymethylcellulose sodium salt (CMC) activity after repeating 10 cycles of hydrolysis and 89% of filter paper activity after 6 cycles relative to original activity, exhibiting excellent reusability. This layer coimmobilization system of BG and cellulase on the polymer film displays tremendous potential for practical application in a biorefinery.
探索合适的固定化策略以提高纤维素酶的催化效率和可重复使用性对于降低成本和促进纤维素衍生生物乙醇的工业化具有重要意义。在这项工作中，通过可见光诱导接枝聚合，在低密度聚乙烯（LDPE）薄膜上制备了具有薄聚乙二醇（PEG）水凝胶内层和聚丙烯酸钠（PAANa）刷外层的层状结构。两种酶，β-葡萄糖苷酶（BG）和纤维素酶，分别被共固定在这个分级薄膜上。作为补充纤维素酶以提高催化效率，BG 在从 LDPE 表面进行接枝聚合时被原位包埋在PEG 水凝胶内层中。在PEG 水凝胶层上重新引发丙烯酸钠接枝聚合后，纤维素酶通过共价键结合到PAANa 刷外层上。由于温和的反应条件（可见光照射和室温），固定化 BG 在接枝聚合后仍保持高活性。固定化不会改变 BG 或纤维素酶的最适 pH 和温度，但最适 pH 值会从 5.0 左右移动到 5.0。与原始的单一纤维素酶系统和分离的 BG/纤维素酶固定化系统的原始活性相比，双酶系统的催化活性分别提高了 82%和 20%。与原始活性相比，双酶系统在重复水解 10 次后，仍能保持 93%的羧甲基纤维素钠（CMC）活性，在重复 6 次后，仍能保持 89%的滤纸活性，表现出优异的可重复使用性。这种聚合物薄膜上 BG 和纤维素酶的层共固定化系统在生物炼制中的实际应用中具有巨大的潜力。
