Generation of an E. coli platform strain for improved sucrose utilization using adaptive laboratory evolution

Microbial Cell Factories

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[1] 0000 0001 2181 8870, grid.5170.3, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Building 220, Kemitorvet, 2800 Kgs, Lyngby, Denmark;0000 0001 2181 8870, grid.5170.3, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Building 220, Kemitorvet, 2800 Kgs, Lyngby, Denmark;0000 0001 2107 4242, grid.266100.3, Department of Bioengineering, University of California, 9500 Gilman Drive La Jolla, 92093, San Diego, CA, USA;0000 0004 1936 9991, grid.35403.31, Department of Animal Sciences, Institute for Genomic Biology and Energy Biosciences Institute, University of Illinois, 61801, Urbana, IL, USA;

关键词: Escherichia coli;    Renewable feedstocks;    Sucrose;    Adaptive laboratory evolution;    Platform strains;   

DOI  :  10.1186/s12934-019-1165-2

来源: publisher

【 摘 要 】

BackgroundSucrose is an attractive industrial carbon source due to its abundance and the fact that it can be cheaply generated from sources such as sugarcane. However, only a few characterized Escherichia coli strains are able to metabolize sucrose, and those that can are typically slow growing or pathogenic strains.MethodsTo generate a platform strain capable of efficiently utilizing sucrose with a high growth rate, adaptive laboratory evolution (ALE) was utilized to evolve engineered E. coli K-12 MG1655 strains containing the sucrose utilizing csc genes (cscB, cscK, cscA) alongside the native sucrose consuming E. coli W.ResultsEvolved K-12 clones displayed an increase in growth and sucrose uptake rates of 1.72- and 1.40-fold on sugarcane juice as compared to the original engineered strains, respectively, while E. coli W clones showed a 1.4-fold increase in sucrose uptake rate without a significant increase in growth rate. Whole genome sequencing of evolved clones and populations revealed that two genetic regions were frequently mutated in the K-12 strains; the global transcription regulatory genes rpoB and rpoC, and the metabolic region related to a pyrimidine biosynthetic deficiency in K-12 attributed to pyrE expression. These two mutated regions have been characterized to confer a similar benefit when glucose is the main carbon source, and reverse engineering revealed the same causal advantages on M9 sucrose. Additionally, the most prevalent mutation found in the evolved E. coli W lineages was the inactivation of the cscR gene, the transcriptional repression of sucrose uptake genes.ConclusionThe generated K-12 and W platform strains, and the specific sets of mutations that enable their phenotypes, are available as valuable tools for sucrose-based industrial bioproduction in the facile E. coli chassis.

【 授权许可】

CC BY