High Enzymatic Activity of CO2 Conversion Achieved by Controllable Enzyme Immobilization and Microenvironment

Enzyme immobilization has greatly facilitated the development of biocatalysts in industry. Lack of control over the enzymatic orientation and microenvironment often results in low enzymatic activity, which, in turn, limits their applications on a large scale. In this study, the immobilization of a His-tagged enzyme was performed at a pH of 7.0 using a vinyl sulfone-activated resin, and varied microenvironments were selected through the screening of blocking reagents characterized by diverse charges and chain lengths. The enzyme 2,3-dihydroxybenzoic acid decarboxylase (200 μg) was used for CO2 conversion under specified conditions (2.7 M KHCO3, 0.2 MPa of CO2, temperatures of 30/45 °C). Remarkably, the relative activity of the enzyme, when it was immobilized and blocked by mercaptoethylamine, reached 97.2%, even exceeding the activity of the free enzyme by achieving a 107.3% conversion of CO2. When the immobilized enzyme was blocked using polyethylene glycol (PEG), its tolerance to heat, pH fluctuations, and exposure to organic solvents was enhanced. Moreover, the immobilized enzyme demonstrated a consistent catalytic activity over seven catalysis cycles. The mechanistic study unraveled the synergistic effects of the enzymatic orientation and microenvironment, which reduce mass transfer resistance and increases stability. Such high activity of the immobilized enzyme could be applied for CO2 fixation as well as for the rational design of enzyme immobilization at the molecular level.