Abstract：
Enzymes play a vital role in synthesizing complex biological molecules like hyaluronic acid (HA). Immobilizing enzymes on support materials is essential for their efficient use and reuse in multiple cycles. Microgels, composed of cross-linked, highly swollen polymer networks, are ideal for enzyme uptake owing to their high porosity. This study demonstrates the immobilization of His6-tagged hyaluronan synthase from Pasteurella multocida (PmHAS) onto nitrilotriacetic acid functionalized microgels using different bivalent ions (Ni2+, Co2+, Mn2+, Mg2+, and Fe2+) via metal affinity binding. The results indicate that using Ni2+ yields the microgels with the highest enzyme uptake and HA formation. The immobilized PmHAS enables repetitive enzymatic production, producing high molecular weight HAs with decreasing dispersities in each step. Furthermore, the highest reported yield of HA with high molecular weight for immobilized PmHAS is achieved. This system establishes a foundation for continuous HA formation, with future works potentially enhancing PmHAS stability through protein engineering.
摘要：
酶在合成透明质酸（HA）等复杂生物分子中起着至关重要的作用。将酶固定在载体材料上对于它们在多个循环中的有效使用和再利用是必不可少的。微凝胶,由交联组成，高度溶胀的聚合物网络，是理想的酶吸收由于其高孔隙率。这项研究证明了使用不同的二价离子（Ni2，Co2+,Mn2+,Mg2+,和Fe2+)通过金属亲和结合。结果表明,使用Ni2+产生具有最高酶摄取和HA形成的微凝胶。固定化PmHAS能够重复酶促生产，生产高分子量的HA，在每个步骤中的分散性降低。此外，对于固定化PmHAS,实现了具有高分子量的HA的最高报道产率。该系统为连续形成HA奠定了基础，未来的工作可能会通过蛋白质工程增强PmHAS的稳定性。
