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

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Abstract

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.
Original languageEnglish
Article number116
JournalMicrobial Cell Factories
Volume18
ISSN1475-2859
DOIs
Publication statusPublished - 2019

Keywords

  • Escherichia coli
  • Renewable feedstocks
  • Sucrose
  • Adaptive laboratory evolution
  • Platform strains

Cite this

@article{d4d8bb1390c94b20adce5a40097690ff,
title = "Generation of an E. coli platform strain for improved sucrose utilization using adaptive laboratory evolution",
abstract = "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.",
keywords = "Escherichia coli, Renewable feedstocks, Sucrose, Adaptive laboratory evolution, Platform strains",
author = "Mohamed, {Elsayed Tharwat Tolba} and Hemanshu Mundhada and Landberg, {Jenny Marie} and Isaac Cann and Mackie, {Roderick I.} and Nielsen, {Alex Toftgaard} and Herrgard, {Markus J.} and Feist, {Adam M.}",
year = "2019",
doi = "10.1186/s12934-019-1165-2",
language = "English",
volume = "18",
journal = "Microbial Cell Factories",
issn = "1475-2859",
publisher = "BioMed Central",

}

Generation of an E. coli platform strain for improved sucrose utilization using adaptive laboratory evolution. / Mohamed, Elsayed Tharwat Tolba; Mundhada, Hemanshu; Landberg, Jenny Marie; Cann, Isaac; Mackie, Roderick I.; Nielsen, Alex Toftgaard; Herrgard, Markus J.; Feist, Adam M.

In: Microbial Cell Factories, Vol. 18, 116, 2019.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Generation of an E. coli platform strain for improved sucrose utilization using adaptive laboratory evolution

AU - Mohamed, Elsayed Tharwat Tolba

AU - Mundhada, Hemanshu

AU - Landberg, Jenny Marie

AU - Cann, Isaac

AU - Mackie, Roderick I.

AU - Nielsen, Alex Toftgaard

AU - Herrgard, Markus J.

AU - Feist, Adam M.

PY - 2019

Y1 - 2019

N2 - 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.

AB - 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.

KW - Escherichia coli

KW - Renewable feedstocks

KW - Sucrose

KW - Adaptive laboratory evolution

KW - Platform strains

U2 - 10.1186/s12934-019-1165-2

DO - 10.1186/s12934-019-1165-2

M3 - Journal article

VL - 18

JO - Microbial Cell Factories

JF - Microbial Cell Factories

SN - 1475-2859

M1 - 116

ER -