Formate and Nitrate Utilization in Enterobacter aerogenes for Semi‐Anaerobic Production of Isobutanol

Anaerobic bioprocessing is preferred because of its economic advantages. However, low productivity and decreased growth of the host strain have limited the use of the anaerobic process. Anaerobic respiration can be applied to anoxic processing using formate and nitrate metabolism to improve the productivity of value‐added metabolites. A isobutanol‐producing strains is constructed using Enterobacter aerogenes as a host strain by metabolic engineering approaches. The byproduct pathway (ldhA, budA, and pflB) is knocked out, and heterologous keto‐acid decarboxylase (kivD) and alcohol dehydrogenase (adhA) are expressed along with the L‐valine synthesis pathway (ilvCD and budB). The pyruvate formate‐lyase mutant shows decreased growth rates when cultivated in semi‐anaerobic conditions, which results in a decline in productivity. When formate and nitrate are supplied in the culture medium, the growth rates and amount of isobutanol production is restored (4.4 g L−1, 0.23 g g−1 glucose, 0.18 g L−1 h−1). To determine the function of the formate and nitrate coupling reaction system, the mutant strains that could not utilize formate or nitrate is contructed. Decreased growth and productivity are observed in the nitrate reductase (narG) mutant strain. This is the first report of engineering isobutanol‐producing E. aerogenes to increase strain fitness via augmentation of formate and nitrate metabolism during anaerobic cultivation. Microorganisms can generate cellular energy by electron donor and acceptor in oxygen‐deficient condition. In this study, Enterobacter aerogenes is metabolically engineered for isobutanol production. When formate and nitrate are supplied as electron donor and acceptor, respectively, titer and productivity of isobutanol increased with improved energy supply in semi‐anaerobic condition. This study suggest that anaerobic respiration assists cell growth as well as isobutanol productivity. This article is part of an AFOB (Asian Federation of Biotechnology) Special issue. To learn more about the AFOB visit www.afob.org.