Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid

Godefroid Charbon, Jiangyun Wang, Eric Brustad, Peter G. Schultz, Arthur L. Horwich, Christine Jacobs-Wagner, Eli Chapman

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    Resumé

    The molecular chaperone GroEL is required for bacterial growth under all conditions, mediating folding assistance, via its central cavity, to a diverse set of cytosolic proteins; yet the subcellular localization of GroEL remains unresolved. An earlier study, using antibody probing of fixed Escherichia coli cells, indicated colocalization with the cell division protein FtsZ at the cleavage furrow, while a second E. coli study of fixed cells indicated more even distribution throughout the cytoplasm. Here, for the first time, we have examined the spatial distribution of GroEL in living cells using incorporation of a fluorescent unnatural amino acid into the chaperone. Fluorescence microscopy indicated that GroEL is diffusely distributed, both under normal and stress conditions. Importantly, the present procedure uses a small, fluorescent unnatural amino acid to visualize GroEL in vivo, avoiding the steric demands of a fluorescent protein fusion, which compromises proper GroEL assembly. Further, this unnatural amino acid incorporation avoids artifacts that can occur with fixation and antibody staining.
    OriginalsprogEngelsk
    TidsskriftBioorganic & Medicinal Chemistry Letters
    Vol/bind21
    Udgave nummer20
    Sider (fra-til)6067-70
    ISSN0960-894X
    DOI
    StatusUdgivet - 15 okt. 2011

    Citer dette

    Charbon, G., Wang, J., Brustad, E., Schultz, P. G., L. Horwich, A., Jacobs-Wagner, C., & Chapman, E. (2011). Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid. Bioorganic & Medicinal Chemistry Letters, 21(20), 6067-70. https://doi.org/10.1016/j.bmcl.2011.08.057
    Charbon, Godefroid ; Wang, Jiangyun ; Brustad, Eric ; Schultz, Peter G. ; L. Horwich, Arthur ; Jacobs-Wagner, Christine ; Chapman, Eli. / Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid. I: Bioorganic & Medicinal Chemistry Letters. 2011 ; Bind 21, Nr. 20. s. 6067-70.
    @article{52b2284bfb004adba1da315912c51e20,
    title = "Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid",
    abstract = "The molecular chaperone GroEL is required for bacterial growth under all conditions, mediating folding assistance, via its central cavity, to a diverse set of cytosolic proteins; yet the subcellular localization of GroEL remains unresolved. An earlier study, using antibody probing of fixed Escherichia coli cells, indicated colocalization with the cell division protein FtsZ at the cleavage furrow, while a second E. coli study of fixed cells indicated more even distribution throughout the cytoplasm. Here, for the first time, we have examined the spatial distribution of GroEL in living cells using incorporation of a fluorescent unnatural amino acid into the chaperone. Fluorescence microscopy indicated that GroEL is diffusely distributed, both under normal and stress conditions. Importantly, the present procedure uses a small, fluorescent unnatural amino acid to visualize GroEL in vivo, avoiding the steric demands of a fluorescent protein fusion, which compromises proper GroEL assembly. Further, this unnatural amino acid incorporation avoids artifacts that can occur with fixation and antibody staining.",
    author = "Godefroid Charbon and Jiangyun Wang and Eric Brustad and Schultz, {Peter G.} and {L. Horwich}, Arthur and Christine Jacobs-Wagner and Eli Chapman",
    year = "2011",
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    language = "English",
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    Charbon, G, Wang, J, Brustad, E, Schultz, PG, L. Horwich, A, Jacobs-Wagner, C & Chapman, E 2011, 'Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid', Bioorganic & Medicinal Chemistry Letters, bind 21, nr. 20, s. 6067-70. https://doi.org/10.1016/j.bmcl.2011.08.057

    Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid. / Charbon, Godefroid; Wang, Jiangyun; Brustad, Eric ; Schultz, Peter G.; L. Horwich, Arthur; Jacobs-Wagner, Christine ; Chapman, Eli.

    I: Bioorganic & Medicinal Chemistry Letters, Bind 21, Nr. 20, 15.10.2011, s. 6067-70.

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    TY - JOUR

    T1 - Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid

    AU - Charbon, Godefroid

    AU - Wang, Jiangyun

    AU - Brustad, Eric

    AU - Schultz, Peter G.

    AU - L. Horwich, Arthur

    AU - Jacobs-Wagner, Christine

    AU - Chapman, Eli

    PY - 2011/10/15

    Y1 - 2011/10/15

    N2 - The molecular chaperone GroEL is required for bacterial growth under all conditions, mediating folding assistance, via its central cavity, to a diverse set of cytosolic proteins; yet the subcellular localization of GroEL remains unresolved. An earlier study, using antibody probing of fixed Escherichia coli cells, indicated colocalization with the cell division protein FtsZ at the cleavage furrow, while a second E. coli study of fixed cells indicated more even distribution throughout the cytoplasm. Here, for the first time, we have examined the spatial distribution of GroEL in living cells using incorporation of a fluorescent unnatural amino acid into the chaperone. Fluorescence microscopy indicated that GroEL is diffusely distributed, both under normal and stress conditions. Importantly, the present procedure uses a small, fluorescent unnatural amino acid to visualize GroEL in vivo, avoiding the steric demands of a fluorescent protein fusion, which compromises proper GroEL assembly. Further, this unnatural amino acid incorporation avoids artifacts that can occur with fixation and antibody staining.

    AB - The molecular chaperone GroEL is required for bacterial growth under all conditions, mediating folding assistance, via its central cavity, to a diverse set of cytosolic proteins; yet the subcellular localization of GroEL remains unresolved. An earlier study, using antibody probing of fixed Escherichia coli cells, indicated colocalization with the cell division protein FtsZ at the cleavage furrow, while a second E. coli study of fixed cells indicated more even distribution throughout the cytoplasm. Here, for the first time, we have examined the spatial distribution of GroEL in living cells using incorporation of a fluorescent unnatural amino acid into the chaperone. Fluorescence microscopy indicated that GroEL is diffusely distributed, both under normal and stress conditions. Importantly, the present procedure uses a small, fluorescent unnatural amino acid to visualize GroEL in vivo, avoiding the steric demands of a fluorescent protein fusion, which compromises proper GroEL assembly. Further, this unnatural amino acid incorporation avoids artifacts that can occur with fixation and antibody staining.

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    DO - 10.1016/j.bmcl.2011.08.057

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    JO - Bioorganic & Medicinal Chemistry Letters

    JF - Bioorganic & Medicinal Chemistry Letters

    SN - 0960-894X

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