title: Modification of glycolysis and its effect on the production of L-threonine in Escherichia coli.
abstract: High concentrations of acetate, the main by-product of Escherichia coli (E. coli) high cell density culture, inhibit bacterial growth and L-threonine production. Since metabolic overflux causes acetate accumulation, we attempted to reduce acetate production by redirecting glycolysis flux to the pentose phosphate pathway by deleting the genes encoding phosphofructokinase (pfk) and/or pyruvate kinase (pyk) in an L-threonine-producing strain of E. coli, THRD. pykF, pykA, pfkA, and pfkB deletion mutants produced less acetate (9.44 ± 0.83, 3.86 ± 0.88, 0.30 ± 0.25, and 6.99 ± 0.85 g/l, respectively) than wild-type THRD cultures (19.75 ± 0.93 g/l). THRDΔpykF and THRDΔpykA produced 11.05 and 5.35 % more L-threonine, and achieved a 10.91 and 5.60 % higher yield on glucose, respectively. While THRDΔpfkA grew more slowly and produced less L-threonine than THRD, THRDΔpfkB produced levels of L-threonine (102.28 ± 2.80 g/l) and a yield on glucose (0.34 g/g) similar to that of THRD. The dual deletion mutant THRDΔpfkBΔpykF also achieved low acetate (7.42 ± 0.81 g/l) and high L-threonine yields (111.37 ± 2.71 g/l). The level of NADPH in THRDΔpfkA cultures was depressed, whereas all other mutants produced more NADPH than THRD did. These results demonstrated that modification of glycolysis in E. coli THRD reduced acetate production and increased accumulation of L-threonine.
authors.affiliation: National and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science and Technology, Tianjin, 300457, People's Republic of China.
authors.forename: Xixian
authors.initials: X
authors.lastname: Xie
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liang
authors.forename: Hongliang
authors.initials: H
authors.lastname: Liu
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
authors.forename: Qingyang
authors.initials: Q
authors.lastname: Xu
authors.forename: Chenglin
authors.initials: C
authors.lastname: Zhang
authors.forename: Ning
authors.initials: N
authors.lastname: Chen
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005285:Fermentation; D017353:Gene Deletion; D005947:Glucose / Q000378:metabolism; D006019:Glycolysis; D009249:NADP / Q000378:metabolism; D010427:Pentose Phosphate Pathway; D025221:Phosphofructokinases / Q000235:genetics; D011770:Pyruvate Kinase / Q000235:genetics; D013912:Threonine / Q000096:biosynthesis*
chemical_list: D013912:Threonine; D009249:NADP; D025221:Phosphofructokinases; D011770:Pyruvate Kinase; D005947:Glucose