(560av) Polyelectrolytes Enabled Co-Localization of Enzyme Cascades for One-Pot Synthesis

Authors: 
Wang, Z., Tsinghua University
Wang, J., Tsinghua University
Chen, G., Tsinghua University
Xu, W., Tsinghua University
Fu, Z., Tsinghua University
Jiang, G., Key Lab of Industrial Biocatalysis, Ministry of Education, Tsinghua University
Wu, J., University of California Riverside
Liu, Z., Key Lab of Industrial Biocatalysis, Ministry of Education, Tsinghua University
Co-localization of enzyme cascades for one-pot synthesis hold great promise for the green production of fine chemicals because it circumvents the intermediate purifications, and meanwhile, facilitates the substrate transport to the subsequent enzyme, mimicking that occurs a living cell in nature. However enzymes from different origin may differ dramatically in their pH window of stability and/or activity, and thus hinder the operation and the performance of the enzyme cascade.

Here we present a method of using polyelectrolyte to co-localize enzyme cascades, in which the polyelectrolyte generates a favorable microenvironment in terms of pH in the vicinity region of the conjugated enzyme and thus enables a high activity and stability.

To demonstrate the effectiveness of this method, we chose chloroperoxidase (CPO) from Caldariomyces fumago and xanthine oxidase (XO) from bovine milk as a model cascade and tested its performance in the oxidization of p-xylene to p-toluic acid. After the conjugation of CPO to polymethacrylic acid (PMAA) whereas XO to chitosan quaternary ammonium salt, respectively, CPO-PMAA@XO-chitosan was obtained by electrostatic interactions between oppositely charged polyelectrolytes. The co-localization of enzymes was confirmed by confocal laser scanning microscopy, which showed the average distance between CPO and XO was 6.2 nm. The oxidization of p-xylene to p-toluic acid operated at various pH showed that, compared to the native counterpart, the CPO-PMAA@XO-chitosan received a 7.6-fold increase in throughput and moreover a wider pH window of activity and stability. The effectiveness of this method was also demonstrated in GOx-PMAA@HRP-chitosan cascade. The enhanced activity and expanded operation window suggests that this assembly method is promising for one-pot enzyme cascade reaction.