title: Combining De Ley-Doudoroff and methylerythritol phosphate pathways for enhanced isoprene biosynthesis from D-galactose.
abstract: An engineered Escherichia coli strain was developed for enhanced isoprene production using D-galactose as substrate. Isoprene is a valuable compound that can be biosynthetically produced from pyruvate and glyceraldehyde-3-phosphate (G3P) through the methylerythritol phosphate pathway (MEP). The Leloir and De Ley-Doudoroff (DD) pathways are known existing routes in E. coli that can supply the MEP precursors from D-galactose. The DD pathway was selected as it is capable of supplying equimolar amounts of pyruvate and G3P simultaneously. To exclusively direct D-galactose toward the DD pathway, an E. coli ΔgalK strain with blocked Leloir pathway was used as the host. To obtain a fully functional DD pathway, a dehydrogenase encoding gene (gld) was recruited from Pseudomonas syringae to catalyze D-galactose conversion to D-galactonate. Overexpressions of endogenous genes known as MEP bottlenecks, and a heterologous gene, were conducted to enhance and enable isoprene production, respectively. Growth test confirmed a functional DD pathway concomitant with equimolar generation of pyruvate and G3P, in contrast to the wild-type strain where G3P was limiting. Finally, the engineered strain with combined DD-MEP pathway exhibited the highest isoprene production. This suggests that the equimolar pyruvate and G3P pools resulted in a more efficient carbon flux toward isoprene production. This strategy provides a new platform for developing improved isoprenoid producing strains through the combined DD-MEP pathway.
authors.affiliation: Department of Energy and Biotechnology (DEB), Energy and Environment Fusion Technology Center (E2FTC), Myongji University, 116 Myongji-ro, Cheoin-gu, Yongin-si, Gyeonggi-do, 449-728, Republic of Korea.
authors.forename: Kristine Rose M
authors.initials: KR
authors.lastname: Ramos
authors.forename: Kris Niño G
authors.initials: KN
authors.lastname: Valdehuesa
authors.forename: Huaiwei
authors.initials: H
authors.lastname: Liu
authors.forename: Grace M
authors.initials: GM
authors.lastname: Nisola
authors.forename: Won-Keun
authors.initials: WK
authors.lastname: Lee
authors.forename: Wook-Jin
authors.initials: WJ
authors.lastname: Chung
mesh_terms: D001709:Biotechnology / Q000379:methods*; D002070:Butadienes / Q000737:chemistry; D002244:Carbon / Q000737:chemistry; D002384:Catalysis; D004247:DNA / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry; D004896:Erythritol / Q000031:analogs & derivatives* / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D005690:Galactose / Q000737:chemistry*; D005986:Glyceraldehyde 3-Phosphate / Q000737:chemistry; D045782:Hemiterpenes / Q000096:biosynthesis* / Q000737:chemistry; D010420:Pentanes / Q000737:chemistry; D010710:Phosphates / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D044224:Pseudomonas syringae / Q000201:enzymology; D019289:Pyruvic Acid / Q000737:chemistry; D013403:Sugar Phosphates / Q000737:chemistry*
chemical_list: C114232:2-C-methylerythritol 4-phosphate; D002070:Butadienes; D017931:DNA Primers; D045782:Hemiterpenes; D010420:Pentanes; D010710:Phosphates; D013403:Sugar Phosphates; C005059:isoprene; D005986:Glyceraldehyde 3-Phosphate; D002244:Carbon; D019289:Pyruvic Acid; D004247:DNA; D004896:Erythritol; D005690:Galactose