D-lactic acid-producing Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 has been long-term adapted to unfavorable conditions where high concentrations of end-product lactic acid inhibit a balanced growth. Evolving force has been applied into L. mesenteroides ATCC 8293 by growing in MRS media containing increasing lactic acid concentrations of 30 g/L to 95 g/L over 8 months. Of 20 adapted L. mesenteroides mutants, LMA92A, which was isolated on media containing 95 g/L lactic acid, was characterized at systems level. A half maximal inhibitory concentration (IC50) of LMA92A was 33.7 ± 5.1 g/L, which is twice higher than 18.2 ± 2.4 g/L of wild-type (WT). Proteomic analysis of WT and LMA92A shows that expressions of several proteins including chaperonin GroEL and F0F1 ATP synthase subunit beta had been up-regulated in LMA92A. GC/MS analysis of fatty acid compositions of WT and LMA92A shows that concentrations of unsaturated and long-chain fatty acids increased in LMA92A, which help maintain cell membrane integrity. All these positive new traits drove LMA92A to produce 1.6-fold lactic acid titer compared to WT in pH-controlled fed-batch fermentation.
