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L-Cysteine production by metabolically engineered Corynebacterium glutamicum.

Abstract

L-Cysteine is a commercially important amino acid. Here, we report the construction of L-cysteine-producing Corynebacterium glutamicum using a metabolic engineering approach. L-Serine O-acetyltransferase (SAT), encoded by cysE gene, is a key enzyme of L-cysteine biosynthesis, because of its feedback inhibition by L-cysteine. Therefore, we introduced a mutation into the C. glutamicum cysE gene, which appeared to desensitize SAT against feedback inhibition by L-cysteine. We successfully produced L-cysteine by overexpressing this mutant cysE gene in C. glutamicum, while the wild-type strain scarcely produced L-cysteine. To enhance the biosynthesis of L-serine (a substrate for SAT), a mutant serA gene, encoding D-3-phosphoglycerate dehydrogenase to desensitize it against feedback inhibition by L-serine, was additionally overexpressed in the mutant cysE-overexpressing strain and its L-cysteine production was indeed improved. Moreover, we disrupted the ldh gene encoding L-lactate dehydrogenase and the aecD gene encoding cysteine desulfhydrase to prevent the formation of lactic acid as a by-product and degradation of L-cysteine produced at the stationary phase, respectively, which resulted in enhanced L-cysteine production. However, since the concentration of L-cysteine produced still decreased at the stationary phase despite the aecD disruption, NCgl2463 encoding a possible cystine importer protein was further disrupted to prevent cystine import, because the produced L-cysteine is immediately oxidized to cystine. As a result, the time before the start of the decrease in L-cysteine concentration was successfully prolonged. Approximately 200 mg/L of L-cysteine production was achieved by overexpression of mutant cysE and serA genes and disruption of aecD and NCgl2463 genes in C. glutamicum.

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  • Authors+Show Affiliations

    ,

    School of Life Science and Technology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midoriku, Yokohama, Kanagawa, 226-8501, Japan.

    School of Life Science and Technology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midoriku, Yokohama, Kanagawa, 226-8501, Japan. thirasawa@bio.titech.ac.jp.

    Source

    Applied microbiology and biotechnology 103:6 2019 Mar pg 2609-2619

    Pub Type(s)

    Journal Article

    Language

    eng

    PubMed ID

    30729285

    Citation

    Kondoh, Mariko, and Takashi Hirasawa. "L-Cysteine Production By Metabolically Engineered Corynebacterium Glutamicum." Applied Microbiology and Biotechnology, vol. 103, no. 6, 2019, pp. 2609-2619.
    Kondoh M, Hirasawa T. L-Cysteine production by metabolically engineered Corynebacterium glutamicum. Appl Microbiol Biotechnol. 2019;103(6):2609-2619.
    Kondoh, M., & Hirasawa, T. (2019). L-Cysteine production by metabolically engineered Corynebacterium glutamicum. Applied Microbiology and Biotechnology, 103(6), pp. 2609-2619. doi:10.1007/s00253-019-09663-9.
    Kondoh M, Hirasawa T. L-Cysteine Production By Metabolically Engineered Corynebacterium Glutamicum. Appl Microbiol Biotechnol. 2019;103(6):2609-2619. PubMed PMID: 30729285.
    * Article titles in AMA citation format should be in sentence-case
    TY - JOUR T1 - L-Cysteine production by metabolically engineered Corynebacterium glutamicum. AU - Kondoh,Mariko, AU - Hirasawa,Takashi, Y1 - 2019/02/07/ PY - 2018/10/09/received PY - 2019/01/22/accepted PY - 2019/01/20/revised PY - 2019/2/8/pubmed PY - 2019/2/8/medline PY - 2019/2/8/entrez KW - Corynebacterium glutamicum KW - Cystine import protein homologs KW - L-Cysteine KW - Metabolic engineering SP - 2609 EP - 2619 JF - Applied microbiology and biotechnology JO - Appl. Microbiol. Biotechnol. VL - 103 IS - 6 N2 - L-Cysteine is a commercially important amino acid. Here, we report the construction of L-cysteine-producing Corynebacterium glutamicum using a metabolic engineering approach. L-Serine O-acetyltransferase (SAT), encoded by cysE gene, is a key enzyme of L-cysteine biosynthesis, because of its feedback inhibition by L-cysteine. Therefore, we introduced a mutation into the C. glutamicum cysE gene, which appeared to desensitize SAT against feedback inhibition by L-cysteine. We successfully produced L-cysteine by overexpressing this mutant cysE gene in C. glutamicum, while the wild-type strain scarcely produced L-cysteine. To enhance the biosynthesis of L-serine (a substrate for SAT), a mutant serA gene, encoding D-3-phosphoglycerate dehydrogenase to desensitize it against feedback inhibition by L-serine, was additionally overexpressed in the mutant cysE-overexpressing strain and its L-cysteine production was indeed improved. Moreover, we disrupted the ldh gene encoding L-lactate dehydrogenase and the aecD gene encoding cysteine desulfhydrase to prevent the formation of lactic acid as a by-product and degradation of L-cysteine produced at the stationary phase, respectively, which resulted in enhanced L-cysteine production. However, since the concentration of L-cysteine produced still decreased at the stationary phase despite the aecD disruption, NCgl2463 encoding a possible cystine importer protein was further disrupted to prevent cystine import, because the produced L-cysteine is immediately oxidized to cystine. As a result, the time before the start of the decrease in L-cysteine concentration was successfully prolonged. Approximately 200 mg/L of L-cysteine production was achieved by overexpression of mutant cysE and serA genes and disruption of aecD and NCgl2463 genes in C. glutamicum. SN - 1432-0614 UR - https://www.unboundmedicine.com/medline/citation/30729285/L-Cysteine_production_by_metabolically_engineered_Corynebacterium_glutamicum L2 - https://dx.doi.org/10.1007/s00253-019-09663-9 DB - PRIME DP - Unbound Medicine ER -