Fig. 6: Spatiotemporal regulation of bi-enzymatic cascade reactions through compartmentalization of GOX and HRP in PNIPAM/DEX systems.

a Schematic diagram and chemical reaction equations demonstrating the generation of resorufin catalyzed by GOX and HRP. Colocalization of GOX and HRP in the small second-level compartments from the PNIPAM (5 wt%)/DEX (5 wt%) under (b) rhodamine channel, (c) FITC channel, and (d) merged channel at 35 °C. The confocal images show the colocalization of GOX and HRP on the (e) surface and (f) cross-section of the colloidosome-like compartments under the merged channel at 35 °C. g Partitioning fraction of GOX in PNIPAM-rich phase from PNIPAM/DEX systems at 25 °C and 35 °C. h Partitioning fraction of HRP in DEX-rich phase from PNIPAM/DEX systems at 25 °C and 35 °C. i Partitioning fraction of HRP in PNIPAM-rich phase from PNIPAM/DEX systems at 25 °C and 35 °C. Michaelis–Menten plots of tandem reactions catalyzed by GOX and HRP in (j) a 50 mM sodium phosphate buffer (pH 7.4), (k) a 50 mM sodium phosphate buffer (pH 7.4) containing PEG (1 wt%)/DEX (9 wt%), and (l) a 50 mM sodium phosphate buffer (pH 7.4) containing PNIPAM (5 wt%)/DEX (5 wt%) at 25 and 35 °C. Lineweaver–Burk plots of tandem reactions catalyzed by GOX and HRP in (m) a 50 mM sodium phosphate buffer (pH 7.4), (n) a 50 mM sodium phosphate buffer (pH 7.4) containing PEG (1 wt%)/DEX (9 wt%), and (o) a 50 mM sodium phosphate buffer (pH 7.4) containing PNIPAM (5 wt%)/DEX (5 wt%) at 25 and 35 °C. Error bars indicate mean ± SD (n  =  3 independent samples).