Immobilization of Glycosyltransferase into a Hydrophilic Metal-organic Framework for Efficient Biosynthesis of Chondroitin Sulfate.

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Chondroitin sulfate (CS) is a structurally complex anionic polysaccharide widely used in medical, cosmetic and food applications. Enzymatic catalysis is an important strategy for synthesizing CS with uniform chain lengths and well-defined structures. However, the industrial application of glycosyltransferases is hindered by limitations such as low expression yields, poor stability, and challenges in reuse. We developed a mild and rapid one-step synthetic method for the efficient immobilization of chondroitin synthase (KfoC). The resulting KfoC@ZIF-90 composite exhibits high catalytic activity, thermal stability, and pH adaptability. Notably, KfoC@ZIF-90 exhibited 5-fold enhanced thermal stability at 40°C and retained 86% relative activity at pH 10, while also maintaining 90% activity in organic solvents, surpassing the performance of free KfoC. Molecular docking analysis revealed that encapsulating KfoC within hydrophilic ZIF-90 reduced its binding energy to the substrate, thereby improving catalytic performance. Furthermore, KfoC@ZIF-90 can be easily separated from the reaction solution by centrifugation, simplifying product isolation and purification while enabling enzyme reuse. These attributes significantly enhance operability and reduce processing costs, making enzymatic CS synthesis more feasible for industrial applications.