title: Increased production of L-serine in Escherichia coli through Adaptive Laboratory Evolution.
abstract: L-serine is a promising building block biochemical with a high theoretical production yield from glucose. Toxicity of L-serine is however prohibitive for high-titer production in E. coli. Here, E. coli lacking L-serine degradation pathways was evolved for improved tolerance by gradually increasing L-serine concentration from 3 to 100g/L using adaptive laboratory evolution (ALE). Genome sequencing of isolated clones revealed multiplication of genetic regions, as well as mutations in thrA, thereby showing a potential mechanism of serine inhibition. Additional mutations were evaluated by MAGE combined with amplicon sequencing, revealing role of rho, lrp, pykF, eno, and rpoB on tolerance and fitness in minimal medium. Production using the tolerant strains resulted in 37g/L of L-serine with a 24% mass yield. The resulting titer is similar to the highest production reported for any organism thereby highlighting the potential of ALE for industrial biotechnology.
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Hemanshu
authors.initials: H
authors.lastname: Mundhada
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Jose M
authors.initials: JM
authors.lastname: Seoane
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Konstantin
authors.initials: K
authors.lastname: Schneider
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Anna
authors.initials: A
authors.lastname: Koza
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Hanne B
authors.initials: HB
authors.lastname: Christensen
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Tobias
authors.initials: T
authors.lastname: Klein
authors.affiliation: Department of Bioengineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Patrick V
authors.initials: PV
authors.lastname: Phaneuf
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Markus
authors.initials: M
authors.lastname: Herrgard
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark; Department of Bioengineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA. Electronic address: afeist@ucsd.edu.
authors.forename: Adam M
authors.initials: AM
authors.lastname: Feist
authors.affiliation: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark. Electronic address: atn@biosustain.dtu.dk.
authors.forename: Alex T
authors.initials: AT
authors.lastname: Nielsen
mesh_terms: D053898:Biosynthetic Pathways / Q000235:genetics; D019020:Directed Molecular Evolution / Q000379:methods*; D004926:Escherichia coli / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D024861:Genetic Enhancement / Q000379:methods; D005947:Glucose / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics; D012694:Serine / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D015854:Up-Regulation / Q000235:genetics*
chemical_list: D012694:Serine; D005947:Glucose
keywords: Adaptive laboratory evolution (ALE); E. coli; Fermentation; Serine production; Serine tolerance