Ex vivo metabolomics—A hypothesis-free approach to identify native substrate(s) and product(s) of orphan enzymes

Ni, Benedikt; Feussner, Kirstin

doi:10.1016/bs.mie.2022.08.005

Cited by 2 publications

(1 citation statement)

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“…To identify natural substrates of L. incisa for the LiLOX activity a non-targeted ex vivo metabolome approach was performed as described recently ( Feussner and Feussner, 2019 ; Ni and Feussner, 2022 ). A total metabolite extract of L. incisa was generated following a two-phase methyl- tert -butylether (MTBE) extraction modified from former publication ( Matyash et al., 2008 ).…”

Section: Methodsmentioning

confidence: 99%

Plastidic membrane lipids are oxidized by a lipoxygenase in Lobosphaera incisa

Djian¹,

Feussner²,

Herrfurth³

et al. 2022

Front. Plant Sci.

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Green microalgae can accumulate neutral lipids, as part of a general lipid remodeling mechanism under stress such as nitrogen starvation. Lobosphaera incisa is of special interest because of its unique TAG acyl chain composition, especially 20:4 (n-6) can reach up to 21% of dry weight after nitrogen starvation. In order to identify factors that may influence the accumulation of polyunsaturated fatty acids (PUFAs), we identified recently a linoleate 13-lipoxygenase (LiLOX). It shares highest identity with plastidic enzymes from vascular plants and is induced upon nitrogen starvation. Here, we confirmed the localization of LiLOX in the stroma of plastids via transient expression in epithelial onion cells. In order to further characterize this enzyme, we focused on the identification of the endogenous substrate of LiLOX. In this regard, an ex vivo enzymatic assay, coupled with non-targeted analysis via mass spectrometry allowed the identification of MGDG, DGDG and PC as three substrate candidates, later confirmed via in vitro assays. Further investigation revealed that LiLOX has preferences towards the lipid class MGDG, which seems in agreement with its localization in the galactolipid rich plastid. Altogether, this study shows the first characterization of plastidic LOX from green algae, showing preference for MGDGs. However, lipidomics analysis did neither reveal an endogenous LiLOX product nor the final end product of MGDG oxidation. Nevertheless, the latter is a key to understanding the role of this enzyme and since its expression is highest during the degradation of the plastidic membrane, it is tempting to assume its involvement in this process.

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Section: Methodsmentioning

confidence: 99%

Plastidic membrane lipids are oxidized by a lipoxygenase in Lobosphaera incisa

Djian¹,

Feussner²,

Herrfurth³

et al. 2022

Front. Plant Sci.

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Fatty Acid Amide Hydrolases in Upland Cotton (Gossypium hirsutum L.) and the Legume Model Medicago truncatula

Arias Gaguancela

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Fatty acid amide hydrolase (FAAH) is a widely conserved amidase in eukaryotes, best known for inactivating the signal of N-acylethanolamine (NAE) lipid mediators. In the plant Arabidopsis thaliana, FAAH-mediated hydrolysis of NAEs has been associated with numerous biological processes. Recently, the phylogenetic distribution of FAAH into two major branches (group I and II FAAHs) across angiosperms outside of Arabidopsis (and in other Brassicaceae), suggests a previously unrecognized complexity of this enzyme. Although A. thaliana has long been used to assess biological questions for plants, in this case it will fall short in understanding the significance of multiple FAAHs in other plant systems. Thus, in this study, I examined the role (s) of six FAAH isoforms in upland cotton (Gossypium hirsutum L.) and two FAAHs in the legume Medicago truncatula.

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Ex vivo metabolomics—A hypothesis-free approach to identify native substrate(s) and product(s) of orphan enzymes

Cited by 2 publications

References 32 publications

Plastidic membrane lipids are oxidized by a lipoxygenase in Lobosphaera incisa

Plastidic membrane lipids are oxidized by a lipoxygenase in Lobosphaera incisa

Fatty Acid Amide Hydrolases in Upland Cotton (Gossypium hirsutum L.) and the Legume Model Medicago truncatula

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