Metabolic flux analysis of secondary metabolism in plants

• Published in: Metabolic engineering communications Vol. 10; p. e00123
• Main Authors: Shih, Meng-Ling, Morgan, John A.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.06.2020; Elsevier
• Subjects: biochemical pathways; carbon; dietary supplements; energy efficiency; enzymes; genes; isotope labeling; Metabolic channeling; metabolic engineering; Metabolic flux analysis; metabolism; microorganisms; Plant secondary metabolites; secondary metabolites; sensory properties; Stable isotopic labeling; Subcellular compartmentation; synthetic biology; Subcellular compartmentation; MS; IP; IPP; MFA; Phe; Plant secondary metabolites; 13C MFA; Metabolic channeling; INST-MFA; NMR; Metabolic flux analysis; LC; Stable isotopic labeling; MVA; MEP; DMAPP; GC; MVAP; MVAPP; BA; DMAPP, Dimethylallyl diphosphate; BA, Benzoic acid; MFA, Metabolic flux analysis; MVAP, Mevalonate 5-phosphate; NMR, Nuclear magnetic resonance; MVA, Mevalonic acid; 13C MFA, Steady state isotopically labeled metabolic flux analysis; INST-MFA, Isotopically non-steady state metabolic flux analysis; MVAPP, Mevalonate 5-diphosphate; LC, Liquid chromatography; MEP, Methylerythritol 4-phosphate; Phe, Phenylalanine; IP, Isopentenyl phosphate; IPP, Isopentenyl diphosphate; MS, Mass spectrometry; GC, Gas chromatography
• ISSN: 2214-0301; 2214-0301
• DOI: 10.1016/j.mec.2020.e00123

Numerous secondary metabolites from plants are important for their medicinal, nutraceutical or sensory properties. Recently, significant progress has been made in the identification of the genes and enzymes of plant secondary metabolic pathways. Hence, there is interest in using synthetic biology to enhance the production of targeted valuable metabolites in plants. In this article, we examine the contribution that metabolic flux analysis will have on informing the rational selection of metabolic engineering targets as well as analysis of carbon and energy efficiency. Compared to microbes, plants have more complex tissue, cellular and subcellular organization, making precise metabolite concentration measurements more challenging. We review different techniques involved in quantifying flux and provide examples illustrating the application of the techniques. For linear and branched pathways that lead to end products with low turnover, flux quantification is straightforward and doesn’t require isotopic labeling. However, for metabolites synthesized via parallel pathways, there is a requirement for isotopic labeling experiments. If the fed isotopically labeled carbons don’t scramble, one needs to apply transient label balancing methods. In the transient case, it is also necessary to measure metabolite concentrations. While flux analysis is not able to directly identify mechanisms of regulation, it is a powerful tool to examine flux distribution at key metabolic nodes in intermediary metabolism, detect flux to wasteful side pathways, and show how parallel pathways handle flux in wild-type and engineered plants under a variety of physiological conditions. •Plant secondary metabolites have high economic value to human health and pleasure.•Plant secondary metabolites are synthesized by pathways in subcellular compartments.•Metabolic flux analysis can guide the selection of metabolic engineering targets.