Effect of puuC overexpression and nitrate addition on glycerol metabolism and anaerobic 3-hydroxypropionic acid production in recombinant Klebsiella pneumoniae ΔglpKΔdhaT.

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  • Additional Information
    • Source:
      Publisher: Academic Press Country of Publication: Belgium NLM ID: 9815657 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1096-7184 (Electronic) Linking ISSN: 10967176 NLM ISO Abbreviation: Metab Eng Subsets: MEDLINE
    • Publication Information:
      Original Publication: Brugge, Belgium ; Orlando, FL : Academic Press, c1999-
    • Subject Terms:
      Aldehyde Oxidoreductases/*metabolism ; Genetic Enhancement/*methods ; Glycerol/*metabolism ; Klebsiella pneumoniae/*physiology ; Lactic Acid/*analogs & derivatives ; Nitrates/*metabolism ; Recombination, Genetic/*genetics; Aldehyde Oxidoreductases/genetics ; Anaerobiosis/physiology ; Klebsiella pneumoniae/classification ; Lactic Acid/biosynthesis ; Species Specificity ; Up-Regulation
    • Abstract:
      3-Hydroxypropionic acid (3-HP), an industrially important platform chemical, is used as a precursor during the production of many commercially important chemicals. Recently, recombinant strains of K. pneumoniae overexpressing an NAD(+)-dependent γ-glutamyl-γ-aminobutyraldehyde dehydrogenase (PuuC) enzyme of K. pneumoniae DSM 2026 were shown to produce 3-HP from glycerol without the addition coenzyme B(12), which is expensive. However, 3-HP production in K. pneumoniae is accompanied with NADH generation, and this always results in large accumulation of 1,3-propanediol (1,3-PDO) and lactic acid. In this study, we investigated the potential use of nitrate as an electron acceptor both to regenerate NAD(+) and to prevent the formation of byproducts during anaerobic production of 3-HP from glycerol. Nitrate addition could improve NAD(+) regeneration, but decreased glycerol flux towards 3-HP production. To divert more glycerol towards 3-HP, a novel recombinant strain K. pneumoniae ΔglpKΔdhaT (puuC) was developed by disrupting the glpK gene, which encodes glycerol kinase, and the dhaT gene, which encodes 1,3-propanediol oxidoreductase. This strain showed improved cellular NAD(+) concentrations and a high carbon flux towards 3-HP production. Through anaerobic cultivation in the presence of nitrate, this recombinant strain produced more than 40±3mM 3-HP with more than 50% yield on glycerol in shake flasks and 250±10mM 3-HP with approximately 30% yield on glycerol in a fed-batch bioreactor.
      (Copyright © 2012 Elsevier Inc. All rights reserved.)
    • Accession Number:
      0 (Nitrates)
      33X04XA5AT (Lactic Acid)
      C4ZF6XLD2X (hydracrylic acid)
      EC 1.2.- (Aldehyde Oxidoreductases)
      EC 1.2.1.19 (aminobutyraldehyde dehydrogenase)
      PDC6A3C0OX (Glycerol)
    • Publication Date:
      Date Created: 20121002 Date Completed: 20130801 Latest Revision: 20220317
    • Publication Date:
      20240829
    • Accession Number:
      10.1016/j.ymben.2012.09.004
    • Accession Number:
      23022570