Abstract
Lactic acid is mainly used to produce bio-based, bio-degradable polylactic acid. For industrial production of lactic acid, engineered Saccharomyces cerevisiae can be used. To avoid cellular toxicity caused by lactic acid accumulation, pH-neutralizing agents are used, leading to increased production costs. In this study, lactic acid-producing S. cerevisiae BK01 was developed with improved lactic acid tolerance through adaptive laboratory evolution (ALE) on 8% lactic acid. The genetic basis of BK01 could not be determined, suggesting complex mechanisms associated with lactic acid tolerance. However, BK01 had distinctive metabolomic traits clearly separated from the parental strain, and lactic acid production was improved by 17% (from 102 g/L to 119 g/L). To the best of our knowledge, this is the highest lactic acid titer produced by engineered S. cerevisiae without the use of pH neutralizers. Moreover, cellulosic lactic acid production by BK01 was demonstrated using acetate-rich buckwheat husk hydrolysates. Particularly, BK01 revealed improved tolerance against acetic acid of the hydrolysates, a major fermentation inhibitor of lignocellulosic biomass. In short, ALE with a high concentration of lactic acid improved lactic acid production as well as acetic acid tolerance of BK01, suggesting a potential for economically viable cellulosic lactic acid production.
Highlights
Lactic acid is an organic acid widely used in the food, pharmaceutical, textile, and chemical industries, especially for the production of polylactic acid [1,2], a biodegradable and compostable bioplastic [3,4,5]
Strain, cells were subjected to adaptive laboratory evolution (ALE) by serial sub-cultures in complex medium containing a high concentration of lactic acid (8%), which critically limits cell growth [40,41]
S. cerevisiae strain was subjected to ALE with a high conlacticThe acidlactic to itsacid-producing toxic level (8%), BK01 might have a higher tolerance to fermentation centration of lactic acid, and a lactic acid-tolerant BK01 strain was successfully isolated
Summary
Lactic acid is an organic acid widely used in the food, pharmaceutical, textile, and chemical industries, especially for the production of polylactic acid [1,2], a biodegradable and compostable bioplastic [3,4,5]. LAB is not considered as a suitable host for industrial fermentations due to phage contamination issues and high nutritional requirements [9,10]. Engineered Saccharomyces cerevisiae expressing a heterologous lactate dehydrogenase gene (ldh) is a promising workhorse for industrial lactic acid production [11,12]. Various metabolic engineering approaches have been performed to improve lactic acid production by the yeast [13]. Expression of multiple copies of ldh genes increased lactate dehydrogenase expression levels and lactic acid production [14,15,16,17,18]. Xylose-metabolizing SR8 expressing the ldh gene from Lactobacillus acidophilus [8] Evolved strain BK01. SR8LDH strain evolved on complex medium containing 20 g/L glucose and 8% lactic acid This study BK_D1 S. cerevisiae D452-2 expressing the xylose oxidoreductase pathway derived from Pichia stipitis (XYL1, XYL2, and XYL3), ∆ald, adaptive laboratory evolution on xylose [29] Parental strain SR8LDH
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