Breeding of Monofluoroacetate-resistant Strains of Actinobacillus succinogenes and the Mechanism Based on Metabolic Flux Analysis

Abstract:

Succinic acid has received a great deal of attention as an important green chemical stock for the manufacture of synthetic resins, biodegradable polymers and chemical intermediates. In this paper, the breeding mechanism of Actinobacillus succinogenes based on metabolic flux analysis was demonstrated to improve the yield of succinic acid by fermentation. After the NTG treatment, mutants from A. succinogenes CGMCC 1593 which were able to grow in medium containing concentrations of about 50~100 mmol/L of sodium monofluoroacetate were obtained. Among them, a mutant SF-9 was selected for producing more succinic acid and less acetic acid. When fermentations were conducted in a 5 L bioreactors, the final succinic acid concentration of SF-9 (34.8 g/L) increased 23.4%, and the mass ratio of succinic acid/acetic acid increased from 3.3 to 9 compared with those of the parent strain. Based on the metabolic flux analysis of A. succinogenes, PEP was found to be a key node which has an important effect on the production of succinic acid, and the flux ratio of by-productions (acetic, formic, lactic acid) was influenced by PYR node. Compared with the parent strain, the flux to succinic acid of mutant (A. succinogenes SF-9) was significantly increased, while the flux to by-productions had an obvious decline. Therefore, PEP and PYR are not rigid nodes in the metabolic regulation of A. succinogenes.

Key words:succinic acid, breed, monofluoroacetate, metabolic flux analysis