title: [Cloning and expression of the genes of glutathione synthetases].
abstract: The genes(gsh-I,gsh-II) for gamma-glutamyl-cysteine synthetase(GSH-I) and glutathione synthetase(GSH-II) from Escherichia coli B were amplified by PCR and then subcloned into plasmid pUC19 respectively. The DNA fragments harboring gshII and gsh I were inserted into plasmid pTrc99A one by one to get a hybrid plasmid pTrc-gsh. E. coli BL21 was transformed by pTrc-gsh for expression of the related enzymes. Analysis of SDS-PAGE showed that the expected products were expressed. E. coli BL21(pTrc-gsh) were incubated at 37 degrees C and pH 7.2 to OD550 = 0.5. The conditions were then switched to 34 degrees C and pH6.7 after the addition of 0.1 mmol/L IPTG. The expressed products were up to 25% of the total protein of the bacteria. Acetone-treated cells of the engineered strain could synthesize GSH efficiently.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Institute of Biochemistry, East China University of Science & Technology, Shanghai 200237, China.
authors.forename: L X
authors.initials: LX
authors.lastname: Shen
authors.forename: D Z
authors.initials: DZ
authors.lastname: Wei
authors.forename: Z F
authors.initials: ZF
authors.lastname: Zhao
authors.forename: S L
authors.initials: SL
authors.lastname: Zhang
authors.forename: E L
authors.initials: EL
authors.lastname: Wang
mesh_terms: D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005981:Glutathione Synthase / Q000096:biosynthesis / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D011994:Recombinant Proteins; D005981:Glutathione Synthase