title: One-pot synthesis of glutathione by a two-enzyme cascade using a thermophilic ATP regeneration system.
abstract: In vitro cascade catalysis using enzyme-based system is becoming a promising biomanufacturing platform for biofuels and biochemicals production. Glutathione is a pivotal non-protein thiol compound and widely applied in food and pharmaceutical industries. In this study, glutathione was synthesized by a bifunctional glutathione synthetase together with a thermophilic ATP regeneration system through a two-enzyme cascade in vitro. Four bifunctional glutathione synthetases from Streptococcus sanguinis, S. gordonii, S. uberis and Bacillus cereus were applied for glutathione synthesis. The bifunctional glutathione synthetase from S. sanguinis was selected and coupled with the polyphosphate kinase from Thermosynechococcus elongatus BP-1 for regenerating ATP to produce glutathione in one pot. In the optimized system, 28.5mM glutathione was produced within 5h due to efficient ATP regeneration from low-cost polyphosphate. The yield based on added l-cysteine reached 81.4% and the productivity of glutathione achieved 5.7mM/h. The one-pot system indicated a potential biotransformation platform for industrial production of glutathione.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China.
authors.forename: Xing
authors.initials: X
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China.
authors.forename: Bing
authors.initials: B
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China; Shanghai Collaborative Innovation Center for Biomanufacturing Technology, 130 Meilong Road, Shanghai 200237, China. Electronic address: lizm@ecust.edu.cn.
authors.forename: Zhimin
authors.initials: Z
authors.lastname: Li
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Ye
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D001409:Bacillus cereus / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005978:Glutathione / Q000032:analysis / Q000378:metabolism*; D005981:Glutathione Synthase / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering; D017856:Phosphotransferases (Phosphate Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D011122:Polyphosphates / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D013291:Streptococcus / Q000201:enzymology / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D011122:Polyphosphates; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D017856:Phosphotransferases (Phosphate Group Acceptor); C022424:polyphosphate kinase; D005981:Glutathione Synthase; D005978:Glutathione
keywords: ATP regeneration; Bifunctional glutathione synthetase; Glutathione; Polyphosphate; Polyphosphate kinase