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Production of Ophthalmic Acid Using Engineered Escherichia coli.

Abstract：

:Ophthalmic acid (OA; l-γ-glutamyl-l-2-aminobutyryl-glycine) is an analog of glutathione (GSH; l-γ-glutamyl-l-cysteinyl-glycine) in which the cysteine moiety is replaced by l-2-aminobutyrate. OA is a useful peptide for the pharmaceutical and/or food industries. Herein, we report a method for the production of OA using engineered Escherichia coli cells. yggS-deficient E. coli, which lacks the highly conserved pyridoxal 5'-phosphate-binding protein YggS and naturally accumulates OA, was selected as the starting strain. To increase the production of OA, we overexpressed the OA biosynthetic enzymes glutamate-cysteine ligase (GshA) and glutathione synthase (GshB), desensitized the product inhibition of GshA, and eliminated the OA catabolic enzyme γ-glutamyltranspeptidase. The production of OA was further enhanced by the deletion of miaA and ridA with the aim of increasing the availability of ATP and attenuating the unwanted degradation of amino acids, respectively. The final strain developed in this study successfully produced 277 μmol/liter of OA in 24 h without the formation of by-products in a minimal synthetic medium containing 1 mM each glutamate, 2-aminobutyrate, and glycine.IMPORTANCE Ophthalmic acid (OA) is a peptide that has the potential for use in the pharmaceutical and/or food industries. An efficient method for the production of OA would allow us to expand our knowledge about its physiological functions and enable the industrial/pharmaceutical application of this compound. We demonstrated the production of OA using Escherichia coli cells in which OA biosynthetic enzymes and degradation enymes were engineered. We also showed that unique approaches, including the use of a ΔyggS mutant as a starting strain, the establishment of an S495F mutation in GshA, and the deletion of ridA or miaA, facilitated the efficient production of OA in E. coli.

journal_name

Appl Environ Microbiol

journal_title

Applied and environmental microbiology

authors

Ito T,Tokoro M,Hori R,Hemmi H,Yoshimura T

doi

10.1128/AEM.02806-17

subject

Has Abstract

pub_date

2018-03-19 00:00:00

issue

eissn

0099-2240

issn

1098-5336

pii

AEM.02806-17

journal_volume

pub_type

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Production of Ophthalmic Acid Using Engineered Escherichia coli.