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Metabolomic analysis on Microcystis aeruginosa cells under various amino acid supplementation conditions

ABSTRACT: Threonine, an essential amino acid for translation, exhibits toxicity towards slow-growing cyanobacteria. This prompted us to perform transcriptomic and proteomic analyses using Microcystis aeruginosa cells treated with threonine, revealing a noteworthy upregulation of thrC, annotated as pyridoxal 5’-phosphate (PLP)-dependent threonine synthase. PyMoL-based structural prediction and in vitro biochemical assays unveiled the moonlighting functions of ThrC in both threonine biosynthesis and deamination. The production of α-ketobutyrate from threonine, both in vivo and in vitro, provided strong support for the role of ThrC as a threonine deaminase. The kinetics of ThrC as a threonine synthase using O-phospho-homoserine were consistent with the Michaelis-Menten kinetics of ThrC in Thermus thermophilus, albeit with a slightly lower Vmax value. However, the in vitro deaminase activity of ThrC, exhibiting sigmoidal kinetics, suggests the potential production of α-ketobutyrate under threonine oversupply conditions in slow-growing cyanobacteria. The phylogenetic lineage of our ThrC is positioned distinctively apart from canonical threonine synthase and threonine deaminase. Gene duplication and subsequent divergence of thrC, followed by gene deletion, could have contributed to the possession of moonlighting ThrC protein in aquatic bacteria including cyanobacterial lineages. Metabolomic data revealed that the presence of threonine and α-ketobutyrate disrupted the balance of amino acids (alanine and methionine) and DNA biosynthetic pathways via feedback inhibition. Interestingly, the addition of exogenous alanine or methionine could alleviate threonine toxicity. Our data revealed that threonine does not uniformly support the growth of freshwater cyanobacteria to the same extent, but rather can be toxic to certain groups within this bacterial family.

INSTRUMENT(S): Liquid Chromatography MS - positive - hilic, Liquid Chromatography MS - negative - hilic

PROVIDER: MTBLS11852 | MetaboLights | 2025-04-02

REPOSITORIES: MetaboLights

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			Action	DRS
	Data230314_KU_4samples(neg)-Blank_IS(neg)-1.mzML	Mzml
	Data230314_KU_4samples(neg)-Blank_IS(neg)-2.mzML	Mzml
	Data230314_KU_4samples(neg)-Korea_C(neg)-2.mzML	Mzml
	Data230314_KU_4samples(neg)-Korea_C(neg)-3.mzML	Mzml
	Data230314_KU_4samples(neg)-Korea_C(neg).mzML	Mzml

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Project description:This a model from the article: A kinetic model of the branch-point between the methionine and threonine biosynthesis pathways in Arabidopsis thaliana. Curien G, Ravanel S, Dumas R Eur. J. Biochem. 2003 Dec; Volume: 270 (Issue: 23 )]:4615-27 14622248 , Abstract: This work proposes a model of the metabolic branch-point between the methionine and threonine biosynthesis pathways in Arabidopsis thaliana which involves kinetic competition for phosphohomoserine between the allosteric enzyme threonine synthase and the two-substrate enzyme cystathionine gamma-synthase. Threonine synthase is activated by S-adenosylmethionine and inhibited by AMP. Cystathionine gamma-synthase condenses phosphohomoserine to cysteine via a ping-pong mechanism. Reactions are irreversible and inhibited by inorganic phosphate. The modelling procedure included an examination of the kinetic links, the determination of the operating conditions in chloroplasts and the establishment of a computer model using the enzyme rate equations. To test the model, the branch-point was reconstituted with purified enzymes. The computer model showed a partial agreement with the in vitro results. The model was subsequently improved and was then found consistent with flux partition in vitro and in vivo. Under near physiological conditions, S-adenosylmethionine, but not AMP, modulates the partition of a steady-state flux of phosphohomoserine. The computer model indicates a high sensitivity of cystathionine flux to enzyme and S-adenosylmethionine concentrations. Cystathionine flux is sensitive to modulation of threonine flux whereas the reverse is not true. The cystathionine gamma-synthase kinetic mechanism favours a low sensitivity of the fluxes to cysteine. Though sensitivity to inorganic phosphate is low, its concentration conditions the dynamics of the system. Threonine synthase and cystathionine gamma-synthase display similar kinetic efficiencies in the metabolic context considered and are first-order for the phosphohomoserine substrate. Under these conditions outflows are coordinated. SBML level 2 code generated for the JWS Online project by Jacky Snoep using PySCeS Run this model online at http://jjj.biochem.sun.ac.za To cite JWS Online please refer to: Olivier, B.G. and Snoep, J.L. (2004) Web-based modelling using JWS Online , Bioinformatics, 20:2143-2144 Biomodels Curation The model simulates the flux for TS and CGS under conditions given in Table 2 and reproduces the dotted lines given in Table 3 of the paper. There is a typo in the equation for the apparent specificity constant for Phser, Kts (equation13). This was changed after communication with the authors to be: Kts = 5.9E-4+6.2E-2*pow(AdoMet,2.9)/(pow(32,2.9)+pow(AdoMet,2.9)). The model was successfully tested on Jarnac and Copasi. Due to a suggestion from Pedro Mendez the parameter AdoMet, TS and CGS where made constant species. This model originates from BioModels Database: A Database of Annotated Published Models (http://www.ebi.ac.uk/biomodels/). It is copyright (c) 2005-2010 The BioModels.net Team. For more information see the terms of use . To cite BioModels Database, please use: Li C, Donizelli M, Rodriguez N, Dharuri H, Endler L, Chelliah V, Li L, He E, Henry A, Stefan MI, Snoep JL, Hucka M, Le Novère N, Laibe C (2010) BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. BMC Syst Biol., 4:92.

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Metabolomic analysis on Microcystis aeruginosa cells under various amino acid supplementation conditions

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