A novel platform for automated high-throughput fluxome profiling of metabolic variants

Advances in metabolic engineering are enabling the creation of a large number of cell factories. However, high-throughput platforms do not yet exist for rapidly analyzing the metabolic network of the engineered cells. To fill the gap, we developed an integrated solution for fluxome profiling of larg...

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Bibliographic Details
Published in:	Metabolic engineering Vol. 25; pp. 8 - 19
Main Authors:	Heux, Stéphanie, Poinot, Juliette, Massou, Stéphane, Sokol, Serguei, Portais, Jean-Charles
Format:	Journal Article
Language:	English
Published:	Belgium Elsevier Inc 01-09-2014 Elsevier
Subjects:	Automation Batch Cell Culture Techniques - instrumentation Batch Cell Culture Techniques - methods Equipment Design Equipment Failure Analysis Escherichia coli - physiology Escherichia coli Proteins - physiology Flux calculation Fluxomics Gene Expression Profiling - instrumentation High-throughput Life Sciences Metabolic Flux Analysis - instrumentation Metabolome - physiology Mutation - genetics NMR Peptide Mapping - instrumentation Peptide Mapping - methods Robotics - instrumentation Statistical analysis Systems Integration Flux calculation DO HT HEPA PPP Fluxomics LiHa CDW WT ED R5P:C3 High-throughput TSP-d4 RSD Automation Statistical analysis MS R5P:C1 G6PDH HCA PYR PCA NMR TCA OD600 FTBL AKG:C4, C4 of alpha-ketoglutarate, OAAA/PYR:C3 INFORMATION-CONTENT ISOTOPE LABELING EXPERIMENTS ZWF GENE-KNOCKOUT ESCHERICHIA-COLI MICROBIAL CULTIVATIONS CLOSTRIDIUM-ACETOBUTYLICUM QUANTITATIVE 2D NMR MAMMALIAN-CELL CULTURES CENTRAL CARBON METABOLISM CORYNEBACTERIUM-GLUTAMICUM
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Summary:	Advances in metabolic engineering are enabling the creation of a large number of cell factories. However, high-throughput platforms do not yet exist for rapidly analyzing the metabolic network of the engineered cells. To fill the gap, we developed an integrated solution for fluxome profiling of large sets of biological systems and conditions. This platform combines a robotic system for 13C-labelling experiments and sampling of labelled material with NMR-based isotopic fingerprinting and automated data interpretation. As a proof-of-concept, this workflow was applied to discriminate between Escherichia coli mutants with gradual expression of the glucose-6-phosphate dehydrogenase. Metabolic variants were clearly discriminated while pathways that support metabolic flexibility towards modulation of a single enzyme were elucidating. By directly connecting the data flow between cell cultivation and flux quantification, considerable advances in throughput, robustness, release of resources and screening capacity were achieved. This will undoubtedly facilitate the development of efficient cell factories. •Integrated solution for fluxome profiling of large sets of biological systems and conditions.•Suite of high-throughput compatible methods from cell cultivation to flux calculation.•Discrimination of metabolic variants on the basis of their metabolic fluxes.•In depth understanding of biological processes.•Exceeding existing methods in terms of throughput, robustness, release of resources and screening capacity.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1096-7176 1096-7184
DOI:	10.1016/j.ymben.2014.06.001