Effect of the Gad system on Actinobacillus succinogenes during acid stress

Abstract

The glutamate decarboxylase (Gad) system is an important amino acid-dependent acid resistance system commonly found in microorganisms. Actinobacillus succinogenes is one of the best natural producers of succinic acid (SA) but lacks glutamate decarboxylase. This study assessed the effects of Gad system introduction into A. succinogenes. The recombinant strains gadB-SW and gadBC-SW were constructed by heterologous expression of gadB alone, or gadB together with gadC, respectively. After 1.0 and 1.5 h of acid stress at pH 4.6, cell survival of gadBC-SW was greater than gadB-SW. The growth of gadB-SW and gadBC-SW was both affected by the expression of heterologous proteins and by γ-aminobutyric acid, with gadBC-SW growth reduced at a neutral pH. SA production in acidic conditions was evaluated by a shake flask and by 3-L bioreactor fermentation. The results showed gadBC-SW to increase SA production by 8.4% in shake flask compared to the parent strain, SW. For a 3-L bioreactor batch fermentation under acidic environment, the highest conversion rate of sugar to SA was observed for gadBC-SW, reaching 96%. However, SA concentration by gadBC-SW was only 47 g/L and 31 g/L at pH 6.5 and pH 6.0, respectively. In summary, the introduction of heterologous gadB and gadC into A. succinogenes not only improved acid tolerance but also influenced the synthesis of SA and added a metabolic burden.