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Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli

Strain engineering and bioprocessing strategies were applied for biobased production of porphobilinogen (PBG) using Escherichia coli as the cell factory. The non-native Shemin/C4 pathway was first implemented by heterologous expression of hemA from Rhodopseudomonas spheroids to supply carbon flux fr...

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Autores principales: Lall, Davinder, Miscevic, Dragan, Bruder, Mark, Westbrook, Adam, Aucoin, Marc, Moo-Young, Murray, Perry Chou, C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Singapore 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8668860/
https://www.ncbi.nlm.nih.gov/pubmed/34970474
http://dx.doi.org/10.1186/s40643-021-00482-3
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author Lall, Davinder
Miscevic, Dragan
Bruder, Mark
Westbrook, Adam
Aucoin, Marc
Moo-Young, Murray
Perry Chou, C.
author_facet Lall, Davinder
Miscevic, Dragan
Bruder, Mark
Westbrook, Adam
Aucoin, Marc
Moo-Young, Murray
Perry Chou, C.
author_sort Lall, Davinder
collection PubMed
description Strain engineering and bioprocessing strategies were applied for biobased production of porphobilinogen (PBG) using Escherichia coli as the cell factory. The non-native Shemin/C4 pathway was first implemented by heterologous expression of hemA from Rhodopseudomonas spheroids to supply carbon flux from the natural tricarboxylic acid (TCA) pathways for PBG biosynthesis via succinyl-CoA. Metabolic strategies were then applied for carbon flux direction from the TCA pathways to the C4 pathway. To promote PBG stability and accumulation, Clustered Regularly Interspersed Short Palindromic Repeats interference (CRISPRi) was applied to repress hemC expression and, therefore, reduce carbon flowthrough toward porphyrin biosynthesis with minimal impact to cell physiology. To further enhance PBG biosynthesis and accumulation under the hemC-repressed genetic background, we further heterologously expressed native E. coli hemB. Using these engineered E. coli strains for bioreactor cultivation based on ~ 30 g L(−1) glycerol, we achieved high PBG titers up to 209 mg L(−1), representing 1.73% of the theoretical PBG yield, with improved PBG stability and accumulation. Potential biochemical, genetic, and metabolic factors limiting PBG production were systematically identified for characterization. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s40643-021-00482-3.
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spelling pubmed-86688602021-12-28 Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli Lall, Davinder Miscevic, Dragan Bruder, Mark Westbrook, Adam Aucoin, Marc Moo-Young, Murray Perry Chou, C. Bioresour Bioprocess Research Strain engineering and bioprocessing strategies were applied for biobased production of porphobilinogen (PBG) using Escherichia coli as the cell factory. The non-native Shemin/C4 pathway was first implemented by heterologous expression of hemA from Rhodopseudomonas spheroids to supply carbon flux from the natural tricarboxylic acid (TCA) pathways for PBG biosynthesis via succinyl-CoA. Metabolic strategies were then applied for carbon flux direction from the TCA pathways to the C4 pathway. To promote PBG stability and accumulation, Clustered Regularly Interspersed Short Palindromic Repeats interference (CRISPRi) was applied to repress hemC expression and, therefore, reduce carbon flowthrough toward porphyrin biosynthesis with minimal impact to cell physiology. To further enhance PBG biosynthesis and accumulation under the hemC-repressed genetic background, we further heterologously expressed native E. coli hemB. Using these engineered E. coli strains for bioreactor cultivation based on ~ 30 g L(−1) glycerol, we achieved high PBG titers up to 209 mg L(−1), representing 1.73% of the theoretical PBG yield, with improved PBG stability and accumulation. Potential biochemical, genetic, and metabolic factors limiting PBG production were systematically identified for characterization. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s40643-021-00482-3. Springer Singapore 2021-12-13 2021 /pmc/articles/PMC8668860/ /pubmed/34970474 http://dx.doi.org/10.1186/s40643-021-00482-3 Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Research
Lall, Davinder
Miscevic, Dragan
Bruder, Mark
Westbrook, Adam
Aucoin, Marc
Moo-Young, Murray
Perry Chou, C.
Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title_full Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title_fullStr Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title_full_unstemmed Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title_short Strain engineering and bioprocessing strategies for biobased production of porphobilinogen in Escherichia coli
title_sort strain engineering and bioprocessing strategies for biobased production of porphobilinogen in escherichia coli
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8668860/
https://www.ncbi.nlm.nih.gov/pubmed/34970474
http://dx.doi.org/10.1186/s40643-021-00482-3
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