title: Manipulating pyruvate to acetyl-CoA conversion in Escherichia coli for anaerobic succinate biosynthesis from glucose with the yield close to the stoichiometric maximum.
abstract: Efficient succinate production in Escherichia coli is attained during anaerobic glucose fermentation in biosynthetic processes combining the reductive branch of the TCA cycle and the glyoxylate bypass. Pyruvate dehydrogenase (PDH) or pyruvate formate lyase (PFL) serves in E. coli as a source of acetyl-CoA, a substrate for the glyoxylate bypass. Depending on enzymes responsible for acetyl-CoA generation, the contribution of the glyoxylate bypass to the anaerobic succinate biosynthesis may vary to support redox balance resulting in diverse maximum achievable yield values. Anaerobic succinate biosynthesis from glucose was studied using E. coli strains with altered expression of genes encoding PFL and PDH. For acetyl-CoA formation by PFL, the yield of 1.32 mol succinate per mole of glucose was achieved with the theoretical value of 1.6 mol/mol. Involvement of PDH in anaerobic acetyl-CoA synthesis increased succinate yield up to 1.49 mol/mol, which is 89.8% of the predicted maximum (1.6(6) mol/mol). The maximum yield of 1.69 mol succinate per mol glucose, amounting to 98.8% of the stoichiometric maximum (1.71 mol/mol), was achieved with the strain possessing PDH as the primary anaerobic source of acetyl-CoA. During high cell density fermentation, the best engineered strain produced high amounts of succinate (570.7 mM) and only small quantities of acetate (11.9 mM).
authors.affiliation: Research Institute for Genetics and Selection of Industrial Microorganisms, 1-st Dorozhniy pr., 1, 117545 Moscow, Russia. Electronic address: skorokhodova@genetika.ru.
authors.forename: Alexandra Yu
authors.initials: AY
authors.lastname: Skorokhodova
authors.affiliation: Research Institute for Genetics and Selection of Industrial Microorganisms, 1-st Dorozhniy pr., 1, 117545 Moscow, Russia.
authors.forename: Anastasiya A
authors.initials: AA
authors.lastname: Morzhakova
authors.affiliation: Research Institute for Genetics and Selection of Industrial Microorganisms, 1-st Dorozhniy pr., 1, 117545 Moscow, Russia.
authors.forename: Andrey Yu
authors.initials: AY
authors.lastname: Gulevich
authors.affiliation: Research Institute for Genetics and Selection of Industrial Microorganisms, 1-st Dorozhniy pr., 1, 117545 Moscow, Russia.
authors.forename: Vladimir G
authors.initials: VG
authors.lastname: Debabov
mesh_terms: D000105:Acetyl Coenzyme A / Q000235:genetics / Q000378:metabolism; D000123:Acetyltransferases / Q000235:genetics / Q000378:metabolism; D000693:Anaerobiosis; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways; D019289:Pyruvic Acid / Q000378:metabolism*; D019802:Succinic Acid / Q000032:analysis / Q000378:metabolism*
chemical_list: D029968:Escherichia coli Proteins; D000105:Acetyl Coenzyme A; D019289:Pyruvic Acid; D019802:Succinic Acid; D000123:Acetyltransferases; C022560:formate C-acetyltransferase; D005947:Glucose
keywords: Acetyl-CoA; E. coli; Pyruvate; Pyruvate dehydrogenase; Pyruvate formate lyase; Succinic acid