title: Effects of eliminating pyruvate node pathways and of coexpression of heterogeneous carboxylation enzymes on succinate production by Enterobacter aerogenes.
abstract: Lowering the pH in bacterium-based succinate fermentation is considered a feasible approach to reduce total production costs. Newly isolated Enterobacter aerogenes strain AJ110637, a rapid carbon source assimilator under weakly acidic (pH 5.0) conditions, was selected as a platform for succinate production. Our previous work showed that the ΔadhE/PCK strain, developed from AJ110637 with inactivated ethanol dehydrogenase and introduced Actinobacillus succinogenes phosphoenolpyruvate carboxykinase (PCK), generated succinate as a major product of anaerobic mixed-acid fermentation from glucose under weakly acidic conditions (pH <6.2). To further improve the production of succinate by the ΔadhE/PCK strain, metabolically engineered strains were designed based on the elimination of pathways that produced undesirable products and the introduction of two carboxylation pathways from phosphoenolpyruvate and pyruvate to oxaloacetate. The highest production of succinate was observed with strain ES04/PCK+PYC, which had inactivated ethanol, lactate, acetate, and 2,3-butanediol pathways and coexpressed PCK and Corynebacterium glutamicum pyruvate carboxylase (PYC). This strain produced succinate from glucose with over 70% yield (gram per gram) without any measurable formation of ethanol, lactate, or 2,3-butanediol under weakly acidic conditions. The impact of lowering the pH from 7.0 to 5.5 on succinate production in this strain was evaluated under pH-controlled batch culture conditions and showed that the lower pH decreased the succinate titer but increased its yield. These findings can be applied to identify additional engineering targets to increase succinate production.
authors.affiliation: Institute for Innovation, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan yoshinori_tajima@ajinomoto.com.
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Tajima
authors.affiliation: Institute for Innovation, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Yamamoto
authors.affiliation: Institute for Innovation, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan.
authors.forename: Keita
authors.initials: K
authors.lastname: Fukui
authors.affiliation: Institute for Innovation, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan.
authors.forename: Yousuke
authors.initials: Y
authors.lastname: Nishio
authors.affiliation: Research Institute for Bioscience Products & Fine Chemicals, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan.
authors.forename: Kenichi
authors.initials: K
authors.lastname: Hashiguchi
authors.affiliation: Institute for Innovation, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Usuda
authors.affiliation: Department of Biotechnology, Graduate School of Engineering, Tokyo University of Agriculture & Technology, Koganei, Tokyo, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Sode
mesh_terms: D000188:Actinobacillus / Q000201:enzymology / Q000235:genetics; D048230:Corynebacterium glutamicum / Q000201:enzymology / Q000235:genetics; D003470:Culture Media / Q000737:chemistry; D021902:Enterobacter aerogenes / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015870:Gene Expression; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D060847:Metabolic Engineering; D019764:Phosphoenolpyruvate Carboxykinase (ATP) / Q000235:genetics / Q000378:metabolism*; D011766:Pyruvate Carboxylase / Q000235:genetics / Q000378:metabolism*; D019289:Pyruvic Acid / Q000378:metabolism*; D019802:Succinic Acid / Q000378:metabolism*
chemical_list: D003470:Culture Media; D019289:Pyruvic Acid; D019802:Succinic Acid; D019764:Phosphoenolpyruvate Carboxykinase (ATP); D011766:Pyruvate Carboxylase; D005947:Glucose