Fig. 6.

The role of LlPC in growth and milk acidification in Lactococcus and c-di-AMP–regulated aspartate biosynthesis. (A) Schematic diagram of L. lactis central metabolism and truncated TCA cycle. Solid lines indicate enzymatic steps present in L. lactis, while dotted lines with a red “x” indicate the enzyme is missing. For Icd, the gene is present, but enzymatic activity is absent in L. lactis. Metabolites written in gray are not formed in this metabolic pathway. (B) Growth of L. lactis strains in CDM with or without amino acid supplementation after 20-h incubation. (C) Growth (solid lines) and acidification (dotted lines) of milk supplemented with glucose. (D) Growth (solid lines) and acidification (dotted lines) of milk supplemented with glucose with or without additional amino acids. Note that aspartate addition slightly lowers the milk pH. (E) Intracellular aspartate levels in WT, the high c-di-AMP gdpP mutant strain OS2, and OS2 strain containing plasmids with or without WT or mutated pycA. Different letters indicate statistically significant differences at P < 0.0001 (Tukey’s multiple-comparisons test). Data presented in B–E are mean ± SD from three independent biological replicates.