title: Efficient production of the Nylon 12 monomer ω-aminododecanoic acid methyl ester from renewable dodecanoic acid methyl ester with engineered Escherichia coli.
abstract: The expansion of microbial substrate and product scopes will be an important brick promoting future bioeconomy. In this study, an orthogonal pathway running in parallel to native metabolism and converting renewable dodecanoic acid methyl ester (DAME) via terminal alcohol and aldehyde to 12-aminododecanoic acid methyl ester (ADAME), a building block for the high-performance polymer Nylon 12, was engineered in Escherichia coli and optimized regarding substrate uptake, substrate requirements, host strain choice, flux, and product yield. Efficient DAME uptake was achieved by means of the hydrophobic outer membrane porin AlkL increasing maximum oxygenation and transamination activities 8.3 and 7.6-fold, respectively. An optimized coupling to the pyruvate node via a heterologous alanine dehydrogenase enabled efficient intracellular L-alanine supply, a prerequisite for self-sufficient whole-cell transaminase catalysis. Finally, the introduction of a respiratory chain-linked alcohol dehydrogenase enabled an increase in pathway flux, the minimization of undesired overoxidation to the respective carboxylic acid, and thus the efficient formation of ADAME as main product. The completely synthetic orthogonal pathway presented in this study sets the stage for Nylon 12 production from renewables. Its effective operation achieved via fine tuning the connectivity to native cell functionalities emphasizes the potential of this concept to expand microbial substrate and product scopes.
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Ladkau
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany.
authors.forename: Miriam
authors.initials: M
authors.lastname: Assmann
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany; Department of Solar Materials, Helmholtz Centre for Environmental Research - UFZ, Permoserstrasse 15, 04318 Leipzig, Germany.
authors.forename: Manfred
authors.initials: M
authors.lastname: Schrewe
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany.
authors.forename: Mattijs K
authors.initials: MK
authors.lastname: Julsing
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany; Department of Solar Materials, Helmholtz Centre for Environmental Research - UFZ, Permoserstrasse 15, 04318 Leipzig, Germany. Electronic address: andreas.schmid@ufz.de.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schmid
authors.affiliation: Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227 Dortmund, Germany; Department of Solar Materials, Helmholtz Centre for Environmental Research - UFZ, Permoserstrasse 15, 04318 Leipzig, Germany. Electronic address: bruno.buehler@ufz.de.
authors.forename: Bruno
authors.initials: B
authors.lastname: Bühler
mesh_terms: D000409:Alanine / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000502:physiology; D003247:Conservation of Natural Resources / Q000379:methods*; D004926:Escherichia coli / Q000502:physiology*; D024861:Genetic Enhancement / Q000379:methods*; D007850:Lauric Acids / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000502:physiology; D009757:Nylons / Q000302:isolation & purification / Q000378:metabolism*; D019289:Pyruvic Acid / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D007850:Lauric Acids; D009757:Nylons; D011994:Recombinant Proteins; C036222:nylon 12; D019289:Pyruvic Acid; C046049:12-aminododecanoic acid; D000409:Alanine
keywords: Dodecanedioic acid monomethyl ester (PubChem CID: 231998); In vivo cascade biocatalysis; Methyl 12-aminododecanoate (PubChem CID: 3665980); Methyl 12-hydroxydodecanoate (PubChem CID: 522424); Methyl dodecanoate (PubChem CID: 8139); Nylon 12; Orthogonal pathway engineering; Renewable plastics; Substrate uptake