A two-enzyme cascade reaction consisting of two reaction pathways. Studies in bulk solution for understanding the performance of a flow-through device with immobilised enzymes

We found that the lower dependency of 3503 yield observed for sequentially immobilised enzymes originates from a change in one of the two possible reaction pathways due to enzyme separation, which was not the case for enzymes that were co-immobilised

Nicolas Ghéczy; Kai Sasaki; Makoto Yoshimoto; Sajad Pour-Esmaeil; Martin Kröger; Pasquale Stano; Peter Walde

2020

Scholarcy highlights

  • ALaboratory for Multifunctional Materials, Department of Materials, ETH Zurich, Vladimir-Prelog-Weg 5, CH-8093 Zurich, Switzerland
  • We found that the lower dependency of 3503 yield observed for sequentially immobilised enzymes originates from a change in one of the two possible reaction pathways due to enzyme separation, which was not the case for enzymes that were co-immobilised
  • We analysed the reaction in bulk solution in great detail a er both enzymes were added from the beginning to a solution containing both substrates DCFH2-DA and H2O2, allowing the reaction to proceed along pathway 1 and along pathway 2
  • If bovine carbonic anhydrase would have been added before horseradish peroxidase to allow for complete hydrolysis of DCFH2-DA, progress along pathway 2 would have been prevented
  • The data shown in Fig. S-13 and S-14, Electronic supplementary information,† re ect the BCA-catalysed hydrolysis of DCFH2-DA, i.e., the time-dependent disappearance of DCFH2-DA, the transient formation of DCFH2-MA and the formation of the completely hydrolysed product DCFH2
  • 5600 5600 11 400 7200 19 800 analysed by taking into account the known absorption spectra of DCFH2-DA and DCFH2, the latter being obtained through complete hydrolysis of DCFH2-DA with hydroxide ions
  • The determination of the spectra of all components of the reaction and of the molar absorption coefficients of isosbestic points was crucial for understanding the changes in the absorption spectrum of the reaction mixture during the

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