Biosynthesis of poly(2-hydroxyisovalerate-co-lactate) by metabolically engineered Escherichia coli.
Polyhydroxyalkanoates (PHAs) containing 2-hydroxyacids such as lactate (LA) and 2-hydroxybutyrate (2HB) have recently been produced by metabolically engineered microorganisms. Here, we further expanded 2-hydroxyacid monomer spectrum of PHAs by engineering Escherichia coli to produce PHAs containing 2-hydroxyisovalerate (2HIV). To generate 2HIV in vivo, feedback resistant ilvBNmut genes encoding acetohydroxyacid synthase and ilvCD genes encoding ketol-acid reductoisomerase and dihydroxyacid dehydratase, respectively, and panE gene encoding d-2-hydroxyacid dehydrogenase are overexpressed. Also, pct540 gene encoding evolved propionyl-CoA transferase and phaC1437 gene encoding evolved PHA synthase are overexpressed along with ilvBNmut, ilvCD, and panE genes in E. coli strain for in vivo synthesis of 2HIV containing PHAs. E. coli strain expressing all of these genes can produce poly(13.2 mol% 2HIV-co-7.5 mol% 2HB-co-42.5 mol% 3HB-co-36.8 mol% LA) when it is cultured in a chemically defined medium containing 20 g/L of glucose and 2 g/L of sodium 3-hydroxybutyrate (3HB). To produce PHA containing only 2HIV and LA monomers, poxB, pflB, adhE and frdB genes encoding enzymes involved in competing pathways for pyruvate are deleted so that cells can generate more 2HIV and LA. When this engineered E. coli strain expressing ilvBNmut, ilvCD, panE, pct540 and phaC1437 genes is cultured in the medium containing 20 g/L of glucose and 2 mM l-isoleucine, which can inhibit l-threonine dehydratase responsible for in vivo 2HB generation, poly(20 mol% 2HIV-co-80 mol% LA) can be produced to the polymer content of 9.6% w/w. These results suggest that novel PHAs containing 2HIV can be produced by engineering branched-chain amino acid metabolism.
Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), BioProcess Engineering Research Center, Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea.
Jung Eun
JE
Yang
Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), BioProcess Engineering Research Center, Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea.
Je Woong
JW
Kim
Center for Bio-based Chemistry, Division of Convergence Chemistry, Korea Research Institute of Chemical Technology, Daejeon, Republic of Korea.
Young Hoon
YH
Oh
Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), BioProcess Engineering Research Center, Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea.
So Young
SY
Choi
Division of Drug Discovery Research, Korea Research Institute of Chemical Technology, Daejeon, Republic of Korea.
Hyuk
H
Lee
Division of Drug Discovery Research, Korea Research Institute of Chemical Technology, Daejeon, Republic of Korea.
A-Reum
AR
Park
Center for Bio-based Chemistry, Division of Convergence Chemistry, Korea Research Institute of Chemical Technology, Daejeon, Republic of Korea.
Jihoon
J
Shin
Department of Environmental Engineering and Energy, Myongji University, Gyeonggido, Republic of Korea.
Si Jae
SJ
Park
Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), BioProcess Engineering Research Center, Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea.
Sang Yup
SY
Lee
D000596:Amino Acids / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D004798:Enzymes / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D016680:Genome, Bacterial; D005947:Glucose / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D011091:Polyesters / Q000378:metabolism*; D054813:Polyhydroxyalkanoates / Q000378:metabolism; D019289:Pyruvic Acid / Q000378:metabolism; D014631:Valerates / Q000378:metabolism; D014633:Valine / Q000378:metabolism
D000596:Amino Acids; D001426:Bacterial Proteins; D003470:Culture Media; D004798:Enzymes; D011091:Polyesters; D054813:Polyhydroxyalkanoates; D014631:Valerates; C000613723:poly(2-hydroxyisovalerate-co-lactate); C035864:2-hydroxyisovaleric acid; D019289:Pyruvic Acid; D014633:Valine; D005947:Glucose
2-Hydroxyacids; Escherichia coli; Metabolic engineering; Poly(2-hydroxyisovalerate-co-lactate); Polyhydroxyalkanoates