Applying asymptotic methods to synthetic biology: Modelling the reaction kinetics of the mevalonate pathway

Dalwadi, Mohit P.; Garavaglia, Marco; Webb, Joseph P.; King, John R.; Minton, Nigel P.

doi:10.1016/j.jtbi.2017.11.022

Cited by 6 publications

(4 citation statements)

References 26 publications

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“…Several modeling efforts focused on analyzing the biosynthetic and signaling dynamics of complex terpenoids in plants ( Latowski et al., 2000 ; Bruggeman et al., 2001 ; Rios-Estepa et al., 2008 ; Rios-Estepa et al, 2010 ; Band et al., 2012 ; Beguerisse-Diaz et al., 2012 ; Pokhilko et al., 2013 ; Pokhilko et al., 2015 ; Allen and Ptashnyk, 2017 ; Nazareno and Hernandez, 2017 ; Dalwadi et al., 2018 ; Rizza et al., 2021 ). For example, Band et al.…”

Section: Discussionmentioning

confidence: 99%

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Changing biosynthesis of terpenoid percursors in rice through synthetic biology

Basallo,

Perez,

Lucido

et al. 2023

Front. Plant Sci.

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Many highly valued chemicals in the pharmaceutical, biotechnological, cosmetic, and biomedical industries belong to the terpenoid family. Biosynthesis of these chemicals relies on polymerization of Isopentenyl di-phosphate (IPP) and/or dimethylallyl diphosphate (DMAPP) monomers, which plants synthesize using two alternative pathways: a cytosolic mevalonic acid (MVA) pathway and a plastidic methyleritritol-4-phosphate (MEP) pathway. As such, developing plants for use as a platform to use IPP/DMAPP and produce high value terpenoids is an important biotechnological goal. Still, IPP/DMAPP are the precursors to many plant developmental hormones. This creates severe challenges in redirecting IPP/DMAPP towards production of non-cognate plant metabolites. A potential solution to this problem is increasing the IPP/DMAPP production flux in planta. Here, we aimed at discovering, understanding, and predicting the effects of increasing IPP/DMAPP production in plants through modelling. We used synthetic biology to create rice lines containing an additional ectopic MVA biosynthetic pathway for producing IPP/DMAPP. The rice lines express three alternative versions of the additional MVA pathway in the plastid, in addition to the normal endogenous pathways. We collected data for changes in macroscopic and molecular phenotypes, gene expression, isoprenoid content, and hormone abundance in those lines. To integrate the molecular and macroscopic data and develop a more in depth understanding of the effects of engineering the exogenous pathway in the mutant rice lines, we developed and analyzed data-centric, line-specific, multilevel mathematical models. These models connect the effects of variations in hormones and gene expression to changes in macroscopic plant phenotype and metabolite concentrations within the MVA and MEP pathways of WT and mutant rice lines. Our models allow us to predict how an exogenous IPP/DMAPP biosynthetic pathway affects the flux of terpenoid precursors. We also quantify the long-term effect of plant hormones on the dynamic behavior of IPP/DMAPP biosynthetic pathways in seeds, and predict plant characteristics, such as plant height, leaf size, and chlorophyll content from molecular data. In addition, our models are a tool that can be used in the future to help in prioritizing re-engineering strategies for the exogenous pathway in order to achieve specific metabolic goals.

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Section: Discussionmentioning

confidence: 99%

“…Regarding the MVA pathway, we found no model in plants. Still, this pathway was modeled in bacteria ( Weaver et al., 2015 , Dalwadi et al., 2018 ). These MVA models study the dynamics of the pathway in the context of introducing it in bacteria using synthetic biology.…”

Section: Discussionmentioning

confidence: 99%

Changing biosynthesis of terpenoid percursors in rice through synthetic biology

Basallo,

Perez,

Lucido

et al. 2023

Front. Plant Sci.

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“…Although many efforts have been devoted to expanding the toolkit, there remains an urgent need for more advanced methods, especially the CRISPR-based genome editing and gene regulation tools, e.g., CRISPRi [ 178 ] and base editor [ 179 ], to facilitate more complex metabolic engineering applications. Besides, computational simulation tools for C. necator are rather limited nowadays [ 180 , 181 ]. To guide metabolic engineering, more in silico design tools, such as flux balance analysis (FBA) and elementary mode analysis (EMA), should be developed.…”

Section: Discussionmentioning

confidence: 99%

Synthetic biology toolkit for engineering Cupriviadus necator H16 as a platform for CO2 valorization

Pan

Wang

et al. 2021

Biotechnol Biofuels

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Background CO2 valorization is one of the effective methods to solve current environmental and energy problems, in which microbial electrosynthesis (MES) system has proved feasible and efficient. Cupriviadus necator (Ralstonia eutropha) H16, a model chemolithoautotroph, is a microbe of choice for CO2 conversion, especially with the ability to be employed in MES due to the presence of genes encoding [NiFe]-hydrogenases and all the Calvin–Benson–Basham cycle enzymes. The CO2 valorization strategy will make sense because the required hydrogen can be produced from renewable electricity independently of fossil fuels. Main body In this review, synthetic biology toolkit for C. necator H16, including genetic engineering vectors, heterologous gene expression elements, platform strain and genome engineering, and transformation strategies, is firstly summarized. Then, the review discusses how to apply these tools to make C. necator H16 an efficient cell factory for converting CO2 to value-added products, with the examples of alcohols, fatty acids, and terpenoids. The review is concluded with the limitation of current genetic tools and perspectives on the development of more efficient and convenient methods as well as the extensive applications of C. necator H16. Conclusions Great progress has been made on genetic engineering toolkit and synthetic biology applications of C. necator H16. Nevertheless, more efforts are expected in the near future to engineer C. necator H16 as efficient cell factories for the conversion of CO2 to value-added products.

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“…Moreover, as exhibited in recent examples (Dalwadi et al. 2018a , b ; Kumar and Josić 2011 ; Eilertsen et al. 2018 ; Eilertsen and Schnell 2018 ), singular perturbation theory is particularly well suited to understanding systems of chemical reactions exhibiting a significant separation of timescales, so we will use a similar approach here.…”

Section: Introductionmentioning

confidence: 99%

Using singular perturbation theory to determine kinetic parameters in a non-standard coupled enzyme assay

et al. 2020

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We investigate how to characterize the kinetic parameters of an aminotransaminase using a non-standard coupled (or auxiliary) enzyme assay, where the peculiarity arises for two reasons. First, one of the products of the auxiliary enzyme is a substrate for the primary enzyme and, second, we explicitly account for the reversibility of the auxiliary enzyme reaction. Using singular perturbation theory, we characterize the two distinguished asymptotic limits in terms of the strength of the reverse reaction, which allows us to determine how to deduce the kinetic parameters of the primary enzyme for a characterized auxiliary enzyme. This establishes a parameter-estimation algorithm that is applicable more generally to similar reaction networks. We demonstrate the applicability of our theory by performing enzyme assays to characterize a novel putative aminotransaminase enzyme, CnAptA (UniProtKB Q0KEZ8) from Cupriavidus necator H16, for two different omega-amino acid substrates.

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Applying asymptotic methods to synthetic biology: Modelling the reaction kinetics of the mevalonate pathway

Cited by 6 publications

References 26 publications

Changing biosynthesis of terpenoid percursors in rice through synthetic biology

Changing biosynthesis of terpenoid percursors in rice through synthetic biology

Synthetic biology toolkit for engineering Cupriviadus necator H16 as a platform for CO2 valorization

Using singular perturbation theory to determine kinetic parameters in a non-standard coupled enzyme assay

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