loading page

High production of glutathione by in vitro enzymatic cascade after thermostability enhancement
  • Xiangwei Cui,
  • Zhimin Li
Xiangwei Cui
East China University of Science and Technology

Corresponding Author:[email protected]

Author Profile
Zhimin Li
East China University of Science and Technology
Author Profile


The cell free system has been paid more attention due to its potential of facilitating more efficient catalysis of multistep reactions. In this study, an efficient enzymatic cascade of GSH production was developed through the evolution of bifunctional glutathione synthetase (GshF), coupled with polyphosphate kinase (PPK). First, the stability and activity of GshF were enhanced by loop interchange and site-directed mutagenesis. As a result, the GshF half-value period increased 163.3-fold, and its activity raised 18 %. PPK from Jhaorihella thermophile (PPKJT) was characterized and used to regenerate ATP in the GSH synthesis, with hexametaphosphate (PolyP(6)) as the phosphate donor. After the process optimization, 99.9 mM GSH and 7.6 mM oxidized glutathione (GSSG) were produced within 2 h. The molar yield was 95.9 mol/mol based on the amino acid added, while the productivities of GSH achieved 49.95 mM/h, which was the highest yield and productivity ever reported about GSH synthesis.
23 Jun 2020Submitted to AIChE Journal
30 Jun 2020Submission Checks Completed
30 Jun 2020Assigned to Editor
04 Jul 2020Reviewer(s) Assigned
31 Jul 2020Editorial Decision: Revise Major
18 Aug 20201st Revision Received
01 Sep 2020Submission Checks Completed
01 Sep 2020Assigned to Editor
02 Sep 2020Reviewer(s) Assigned
08 Sep 2020Editorial Decision: Accept
Jan 2021Published in AIChE Journal volume 67 issue 1. 10.1002/aic.17055