A kinetic model based approach to understand the heterologous acrylate pathway functioning in lactococcus lactis and lactobacillus plantarum for production of propionic acid

Abstract

Engineering microbes with heterologous pathway for production of high demand products has received considerable attention. One such platform chemical with robust demand growth especially in food and feed industry (66 % of total demand) is propionic acid. At present, propionate is industrially synthesized by petrochemical processes however; recent market needs demand bio-propionic acid mainly due to its ability to serve as a natural preservative . Among fermentation pathways for propionic acid production, acrylate pathway was considered for its ability to promote propionic acid production by minimizing substrate carbon loss. However, multitude of factors have limited in utilizing native microbes to industrial level and necessitate an alternate host. Previously, E. coli, a high lactate precursor production host was chosen and engineered with the acrylate pathway genes from Clostridium propionicum for propionic acid production. Productivity levels of 0.3 g/L could only be achieved and factors such as inclusion body formation, insufficient enzymes, draining of acrylate pathway intermediates to undesired products and regulatory hurdles were attributed to this low product titre. Therefore, an alternative user-friendly biosystem was proposed to facilitate propionate production. Consequently, the study was perpetuated in LAB (Lactococcus lactis and Lactobacillus plantarum), a simple metabolic host with high acid tolerance than E.coli, no inclusion body formation, nonfunctional metabolic networks associated with acrylate pathway intermediates, high lactate production and easy amenable in food industry. Initially, the acrylate pathway was constructed in the chosen LAB strains and were analysed under respective standard glucose fermentation for propionic acid synthesis. Upon induction, the heterologous acrylate pathway resulted in propionic acid production in the constructed strain L. lactis RS3 whereas L. plantarum AP3 did not produce propionic acid. In order tounderstand and quantify constraints of the flux in the