Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes
α,β-Unsaturated carbonyls are a common motif in environmental toxins (e.g. acrolein) as well as therapeutic drugs, including dimethylfumarate (DMFU) and monomethylfumarate (MMFU), which are used to treat multiple sclerosis and psoriasis. These compounds form adducts with protein Cys residues as well...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-02-01
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| Series: | Redox Biology |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2213231722003329 |
| _version_ | 1797954096438706176 |
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| author | Max B. Sauerland Christina Helm Lasse G. Lorentzen Asmita Manandhar Trond Ulven Luke F. Gamon Michael J. Davies |
| author_facet | Max B. Sauerland Christina Helm Lasse G. Lorentzen Asmita Manandhar Trond Ulven Luke F. Gamon Michael J. Davies |
| author_sort | Max B. Sauerland |
| collection | DOAJ |
| description | α,β-Unsaturated carbonyls are a common motif in environmental toxins (e.g. acrolein) as well as therapeutic drugs, including dimethylfumarate (DMFU) and monomethylfumarate (MMFU), which are used to treat multiple sclerosis and psoriasis. These compounds form adducts with protein Cys residues as well as other nucleophiles. The specific targets (‘adductome’) that give rise to their therapeutic or toxic activities are poorly understood. This is due, at least in part, to the absence of antigens or chromophores/fluorophores in these compounds. We have recently reported click-chemistry probes of DMFU and MMFU (Redox Biol., 2022, 52, 102299) that allow adducted proteins to be visualized and enriched for further characterization. In the current study, we hypothesized that adducted proteins could be ‘clicked’ to agarose beads and thereby isolated for LC-MS analysis of DMFU/MMFU targets in primary human coronary artery smooth muscle cells. We show that the probes react with thiols with similar rate constants to the parent drugs, and give rise to comparable patterns of gene induction, confirming similar biological actions. LC-MS proteomic analysis identified ∼2970 cellular targets of DMFU, ∼1440 for MMFU, and ∼140 for the control (succinate-probe) treated samples. The most extensively modified proteins were galectin-1, annexin-A2, voltage dependent anion channel-2 and vimentin. Other previously postulated DMFU targets, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), cofilin, p65 (RELA) and Keap1 were also identified as adducted species, though at lower levels with the exception of GAPDH. These data demonstrate the utility of the click-chemistry approach to the identification of cellular protein targets of both exogenous and endogenous compounds. |
| first_indexed | 2024-04-10T23:13:18Z |
| format | Article |
| id | doaj.art-58ae30826cb949d2b08e3d4db5d50856 |
| institution | Directory Open Access Journal |
| issn | 2213-2317 |
| language | English |
| last_indexed | 2024-04-10T23:13:18Z |
| publishDate | 2023-02-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Redox Biology |
| spelling | doaj.art-58ae30826cb949d2b08e3d4db5d508562023-01-13T04:16:13ZengElsevierRedox Biology2213-23172023-02-0159102560Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probesMax B. Sauerland0Christina Helm1Lasse G. Lorentzen2Asmita Manandhar3Trond Ulven4Luke F. Gamon5Michael J. Davies6Department of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen, 2200, DenmarkDepartment of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen, 2200, DenmarkDepartment of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen, 2200, DenmarkDepartment of Drug Design and Pharmacology, Jagtvej 162, University of Copenhagen, Copenhagen, 2100, DenmarkDepartment of Drug Design and Pharmacology, Jagtvej 162, University of Copenhagen, Copenhagen, 2100, DenmarkDepartment of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen, 2200, DenmarkDepartment of Biomedical Sciences, Panum Institute, University of Copenhagen, Copenhagen, 2200, Denmark; Corresponding author.α,β-Unsaturated carbonyls are a common motif in environmental toxins (e.g. acrolein) as well as therapeutic drugs, including dimethylfumarate (DMFU) and monomethylfumarate (MMFU), which are used to treat multiple sclerosis and psoriasis. These compounds form adducts with protein Cys residues as well as other nucleophiles. The specific targets (‘adductome’) that give rise to their therapeutic or toxic activities are poorly understood. This is due, at least in part, to the absence of antigens or chromophores/fluorophores in these compounds. We have recently reported click-chemistry probes of DMFU and MMFU (Redox Biol., 2022, 52, 102299) that allow adducted proteins to be visualized and enriched for further characterization. In the current study, we hypothesized that adducted proteins could be ‘clicked’ to agarose beads and thereby isolated for LC-MS analysis of DMFU/MMFU targets in primary human coronary artery smooth muscle cells. We show that the probes react with thiols with similar rate constants to the parent drugs, and give rise to comparable patterns of gene induction, confirming similar biological actions. LC-MS proteomic analysis identified ∼2970 cellular targets of DMFU, ∼1440 for MMFU, and ∼140 for the control (succinate-probe) treated samples. The most extensively modified proteins were galectin-1, annexin-A2, voltage dependent anion channel-2 and vimentin. Other previously postulated DMFU targets, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), cofilin, p65 (RELA) and Keap1 were also identified as adducted species, though at lower levels with the exception of GAPDH. These data demonstrate the utility of the click-chemistry approach to the identification of cellular protein targets of both exogenous and endogenous compounds.http://www.sciencedirect.com/science/article/pii/S2213231722003329DimethylfumarateUnsaturated carbonylsMichael adductClick chemistryGalectinElectrophile |
| spellingShingle | Max B. Sauerland Christina Helm Lasse G. Lorentzen Asmita Manandhar Trond Ulven Luke F. Gamon Michael J. Davies Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes Redox Biology Dimethylfumarate Unsaturated carbonyls Michael adduct Click chemistry Galectin Electrophile |
| title | Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes |
| title_full | Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes |
| title_fullStr | Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes |
| title_full_unstemmed | Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes |
| title_short | Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes |
| title_sort | identification of galectin 1 and other cellular targets of alpha beta unsaturated carbonyl compounds including dimethylfumarate by use of click chemistry probes |
| topic | Dimethylfumarate Unsaturated carbonyls Michael adduct Click chemistry Galectin Electrophile |
| url | http://www.sciencedirect.com/science/article/pii/S2213231722003329 |
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