Oxidative Bioactivation of Crotyl Alcohol to the Toxic Endogenous Aldehyde Crotonaldehyde: Association of Protein Carbonylation with Toxicity in Mouse Hepatocytes

F.R. Fontaine, R.A. Dunlop, D.R. Petersen, Philip Burcham

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    Abstract

    Recent confirmation that the toxic unsaturated aldehyde crotonaldehyde (CA) contributes to protein damage during lipid peroxidation confers interest on the molecular actions of this substance. However, since a plethora of structurally related aldehydes form during membrane oxidation, clarifying the toxicological significance of individual products (e.g., CA) is challenging. To facilitate study of the mechanisms underlying CA toxicity, we explored the possibility that it can be formed enzymatically from an unsaturated precursor, crotyl alcohol. This is analogous to the way allyl alcohol is converted in vivo to its toxic oxidation product, acrolein. In kinetic studies, we found that crotyl alcohol was readily oxidized by equine liver alcohol dehydrogenase, with electrospray-mass spectrometry confirming that CA was the main product formed. Moreover, in mouse hepatocytes, crotyl alcohol produced marked time- and concentration-dependent cell killing as well as pronounced glutathione depletion. Both cytotoxicity and glutathione loss were abolished by the alcohol dehydrogenase inhibitor 4-methylpyrazole, indicating an oxidation product mediated these effects. In keeping with expectations that carbonyl-retaining Michael addition adducts would feature prominently during protein modification by CA, exposure to crotyl alcohol resulted in marked carbonylation of a wide range of cell proteins, an effect that was also abolished by 4-methylpyrazole. Damage to a subset of small proteins (e.g., 29, 32, 33 kDa) closely correlated with the severity of cell death. Collectively, these results demonstrate that crotyl alcohol is a useful tool for studying the biochemical and molecular events accompanying intracellular CA formation.
    Original languageEnglish
    Pages (from-to)1051-1058
    JournalChemical Research in Toxicology
    Volume15
    DOIs
    Publication statusPublished - 2002

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    2-butenal
    Protein Carbonylation
    Carbonylation
    Poisons
    Aldehydes
    Toxicity
    Hepatocytes
    Proteins
    Alcohol Dehydrogenase
    Oxidation
    Glutathione
    Acrolein
    Corrosion inhibitors
    Cell death
    Cytotoxicity
    Liver
    Toxicology
    Lipid Peroxidation
    Horses
    Mass spectrometry

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    @article{92f1f152a3e347b09cdbd4c3c96a3836,
    title = "Oxidative Bioactivation of Crotyl Alcohol to the Toxic Endogenous Aldehyde Crotonaldehyde: Association of Protein Carbonylation with Toxicity in Mouse Hepatocytes",
    abstract = "Recent confirmation that the toxic unsaturated aldehyde crotonaldehyde (CA) contributes to protein damage during lipid peroxidation confers interest on the molecular actions of this substance. However, since a plethora of structurally related aldehydes form during membrane oxidation, clarifying the toxicological significance of individual products (e.g., CA) is challenging. To facilitate study of the mechanisms underlying CA toxicity, we explored the possibility that it can be formed enzymatically from an unsaturated precursor, crotyl alcohol. This is analogous to the way allyl alcohol is converted in vivo to its toxic oxidation product, acrolein. In kinetic studies, we found that crotyl alcohol was readily oxidized by equine liver alcohol dehydrogenase, with electrospray-mass spectrometry confirming that CA was the main product formed. Moreover, in mouse hepatocytes, crotyl alcohol produced marked time- and concentration-dependent cell killing as well as pronounced glutathione depletion. Both cytotoxicity and glutathione loss were abolished by the alcohol dehydrogenase inhibitor 4-methylpyrazole, indicating an oxidation product mediated these effects. In keeping with expectations that carbonyl-retaining Michael addition adducts would feature prominently during protein modification by CA, exposure to crotyl alcohol resulted in marked carbonylation of a wide range of cell proteins, an effect that was also abolished by 4-methylpyrazole. Damage to a subset of small proteins (e.g., 29, 32, 33 kDa) closely correlated with the severity of cell death. Collectively, these results demonstrate that crotyl alcohol is a useful tool for studying the biochemical and molecular events accompanying intracellular CA formation.",
    author = "F.R. Fontaine and R.A. Dunlop and D.R. Petersen and Philip Burcham",
    year = "2002",
    doi = "10.1021/tx0255119",
    language = "English",
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    pages = "1051--1058",
    journal = "Chemical Research in Toxicology",
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    publisher = "American Chemical Society",

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    TY - JOUR

    T1 - Oxidative Bioactivation of Crotyl Alcohol to the Toxic Endogenous Aldehyde Crotonaldehyde: Association of Protein Carbonylation with Toxicity in Mouse Hepatocytes

    AU - Fontaine, F.R.

    AU - Dunlop, R.A.

    AU - Petersen, D.R.

    AU - Burcham, Philip

    PY - 2002

    Y1 - 2002

    N2 - Recent confirmation that the toxic unsaturated aldehyde crotonaldehyde (CA) contributes to protein damage during lipid peroxidation confers interest on the molecular actions of this substance. However, since a plethora of structurally related aldehydes form during membrane oxidation, clarifying the toxicological significance of individual products (e.g., CA) is challenging. To facilitate study of the mechanisms underlying CA toxicity, we explored the possibility that it can be formed enzymatically from an unsaturated precursor, crotyl alcohol. This is analogous to the way allyl alcohol is converted in vivo to its toxic oxidation product, acrolein. In kinetic studies, we found that crotyl alcohol was readily oxidized by equine liver alcohol dehydrogenase, with electrospray-mass spectrometry confirming that CA was the main product formed. Moreover, in mouse hepatocytes, crotyl alcohol produced marked time- and concentration-dependent cell killing as well as pronounced glutathione depletion. Both cytotoxicity and glutathione loss were abolished by the alcohol dehydrogenase inhibitor 4-methylpyrazole, indicating an oxidation product mediated these effects. In keeping with expectations that carbonyl-retaining Michael addition adducts would feature prominently during protein modification by CA, exposure to crotyl alcohol resulted in marked carbonylation of a wide range of cell proteins, an effect that was also abolished by 4-methylpyrazole. Damage to a subset of small proteins (e.g., 29, 32, 33 kDa) closely correlated with the severity of cell death. Collectively, these results demonstrate that crotyl alcohol is a useful tool for studying the biochemical and molecular events accompanying intracellular CA formation.

    AB - Recent confirmation that the toxic unsaturated aldehyde crotonaldehyde (CA) contributes to protein damage during lipid peroxidation confers interest on the molecular actions of this substance. However, since a plethora of structurally related aldehydes form during membrane oxidation, clarifying the toxicological significance of individual products (e.g., CA) is challenging. To facilitate study of the mechanisms underlying CA toxicity, we explored the possibility that it can be formed enzymatically from an unsaturated precursor, crotyl alcohol. This is analogous to the way allyl alcohol is converted in vivo to its toxic oxidation product, acrolein. In kinetic studies, we found that crotyl alcohol was readily oxidized by equine liver alcohol dehydrogenase, with electrospray-mass spectrometry confirming that CA was the main product formed. Moreover, in mouse hepatocytes, crotyl alcohol produced marked time- and concentration-dependent cell killing as well as pronounced glutathione depletion. Both cytotoxicity and glutathione loss were abolished by the alcohol dehydrogenase inhibitor 4-methylpyrazole, indicating an oxidation product mediated these effects. In keeping with expectations that carbonyl-retaining Michael addition adducts would feature prominently during protein modification by CA, exposure to crotyl alcohol resulted in marked carbonylation of a wide range of cell proteins, an effect that was also abolished by 4-methylpyrazole. Damage to a subset of small proteins (e.g., 29, 32, 33 kDa) closely correlated with the severity of cell death. Collectively, these results demonstrate that crotyl alcohol is a useful tool for studying the biochemical and molecular events accompanying intracellular CA formation.

    U2 - 10.1021/tx0255119

    DO - 10.1021/tx0255119

    M3 - Article

    VL - 15

    SP - 1051

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    SN - 0893-228X

    ER -