Hyaluronic acid production is enhanced by harnessing the heme-induced respiration in recombinant Lactococcus lactis cultures

Abstract

Lactococcus lactis is a promising host for pathway-engineered production of high molecular weight hyaluronic acid (HA). This work investigates the effect of hemin-supplemented respiratory metabolism in an engineered L. lactis ldh mutant strain (Δldh L. lactis) as a strategy to enhance HA titer in the batch fermentation process. L. lactis lacks a functional electron transport chain (ETC) and ultimately relies on substrate-level phosphorylation for energy production. We conducted detailed investigations under different dissolved oxygen (DO) conditions and found a steady increase in HA titer with increased DO levels. We found a strong correlation between cofactor availability (NAD+ and acetyl-CoA) and the intracellular concentration of HA precursors, UDP-GlcUA and UDP-GlcNAc. We correlated the intracellular precursor levels with the HA titer obtained for the four conditions. We found that these were higher in hemin-supplemented cultures, with the best HA titer of 4.6 g/l. Finally, we observed that the molecular weight of HA (MWHA) correlates strongly with the ratio of HA precursors concentrations. We also found an inverse correlation between MWHA and HA titer across these experiments. These results can be used in optimizing the trade-off between molecular weight and HA production in bioprocesses involving recombinant L. lactis cultures.