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  • Title: Improvement of d-Lactic Acid Production in Saccharomyces cerevisiae Under Acidic Conditions by Evolutionary and Rational Metabolic Engineering.
    Author: Baek SH, Kwon EY, Bae SJ, Cho BR, Kim SY, Hahn JS.
    Journal: Biotechnol J; 2017 Oct; 12(10):. PubMed ID: 28731533.
    Abstract:
    Microbial lactic acid (LA) production under acidic fermentation conditions is favorable to reduce the production cost, but circumventing LA toxicity is a major challenge. A d-LA-producing Saccharomyces cerevisiae strain JHY5610 is generated by expressing d-lactate dehydrogenase gene (Lm. ldhA) from Leuconostoc mesenteroides, while deleting genes involved in ethanol production (ADH1, ADH2, ADH3, ADH4, and ADH5), glycerol production (GPD1 and GPD2), and degradation of d-LA (DLD1). Adaptive laboratory evolution of JHY5610 lead to a strain JHY5710 having higher LA tolerance and d-LA-production capability. Genome sequencing of JHY5710 reveal that SUR1I245S mutation increases LA tolerance and d-LA-production, whereas a loss-of-function mutation of ERF2 only contributes to increasing d-LA production. Introduction of both SUR1I245S and erf2Δ mutations into JHY5610 largely mimic the d-LA-production capability of JHY5710, suggesting that these two mutations, which could modulate sphingolipid production and protein palmitoylation, are mainly responsible for the improved d-LA production in JHY5710. JHY5710 is further improved by deleting PDC1 encoding pyruvate decarboxylase and additional integration of Lm. ldhA gene. The resulting strain JHY5730 produce up to 82.6 g L-1 of d-LA with a yield of 0.83 g g-1 glucose and a productivity of 1.50 g/(L · h) in fed-batch fermentation at pH 3.5.
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