Dataset Information

Metabolic Dynamics in Escherichia coli-Based Cell-Free Systems

ABSTRACT: The field of metabolic engineering has yielded remarkable accomplishments in using cells to produce valuable molecules, and cell-free expression (CFE) systems have the potential to push the field even further. However, CFE systems still face some outstanding challenges, including endogenous metabolic activity that is poorly understood yet has a significant impact on CFE productivity. Here, we use metabolomics to characterize the temporal metabolic changes in CFE systems and their constituent components, including significant metabolic activity in central carbon and amino acid metabolism. We find that while changing the reaction starting state via lysate preincubation impacts protein production, it has a comparatively small impact on metabolic state. We also demonstrate that changes to lysate preparation have a larger effect on protein yield and temporal metabolic profiles, though general metabolic trends are conserved. Finally, while we improve protein production through targeted supplementation of metabolic enzymes, we show that the endogenous metabolic activity is fairly resilient to these enzymatic perturbations. Overall, this work highlights the robust nature of CFE reaction metabolism as well as the importance of understanding the complex interdependence of metabolites and proteins in CFE systems to guide optimization efforts.

INSTRUMENT(S): Tandem Gas Chromatography MS - positive, Tandem Gas Chromatography MS -

SUBMITTER: April Miguez

PROVIDER: MTBLS2630 | MetaboLights | 2023-02-22

REPOSITORIES: MetaboLights

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Dataset's files

Source:

			Action	DRS
	MTBLS2630	Other
	FILES	Other
	a_MTBLS2630_Exp1_GCxGC-MS_metabolite_profiling.txt	Txt
	a_MTBLS2630_Exp2_GCxGC-MS_metabolite_profiling.txt	Txt
	a_MTBLS2630_Exp3_GCxGC-MS_metabolite_profiling.txt	Txt

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Publications

Metabolic Dynamics in <i>Escherichia coli</i>-Based Cell-Free Systems.

Miguez April M AM Zhang Yan Y Piorino Fernanda F Styczynski Mark P MP

ACS synthetic biology 20210903 9

The field of metabolic engineering has yielded remarkable accomplishments in using cells to produce valuable molecules, and cell-free expression (CFE) systems have the potential to push the field even further. However, CFE systems still face some outstanding challenges, including endogenous metabolic activity that is poorly understood yet has a significant impact on CFE productivity. Here, we use metabolomics to characterize the temporal metabolic changes in CFE systems and their constituent com ...[more]

PMID: 34478281

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