Reversible tetramerization of human TK1 to the high catalytic efficient form is induced by pyrophosphate, in addition to tripolyphosphates, or high enzyme concentration

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    Resumé

    Thymidine kinase (TK1) is a key enzyme in the salvage pathway of deoxyribonucleotide metabolism catalyzing the first step in the synthesis of dTTP by the transfer of a gamma-phosphate group from a nucleoside triphosphate to the 5´-hydroxyl group of thymidine forming dTMP. Human TK1 is cytosolic and its activity is absent in resting cells, appears in late G1, increases in S phase coinciding with the increase in DNA synthesis and disappears during mitosis. The fluctuation of TK1 through cell cycle is important for providing a balanced supply of dTTP for DNA replication and is partly due to regulation of TK1 expression at the transcriptional level. However, TK1 is also a regulatory enzyme that can interchange between its dimeric and tetrameric forms, which have low and high catalytic efficiencies, respectively, depending on pre-assay incubation with ATP.  Here, it is investigated which part of ATP is necessary for tetramerisation and how the reaction velocity is influenced by the enzyme concentration. The results show that only two or three of the phosphate groups of ATP are necessary for tetramerisation, and that kinetics and tetramerisation are closely related. Furthermore, enzyme concentration was found to have a pivotal effect on catalytic efficiency.
    OriginalsprogEngelsk
    TidsskriftFEBS Journal
    Vol/bind276
    Udgave nummer2
    Sider (fra-til)571-580
    Antal sider10
    ISSN1742-464X
    DOI
    StatusUdgivet - 2009

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    title = "Reversible tetramerization of human TK1 to the high catalytic efficient form is induced by pyrophosphate, in addition to tripolyphosphates, or high enzyme concentration",
    abstract = "Thymidine kinase (TK1) is a key enzyme in the salvage pathway of deoxyribonucleotide metabolism catalyzing the first step in the synthesis of dTTP by the transfer of a gamma-phosphate group from a nucleoside triphosphate to the 5´-hydroxyl group of thymidine forming dTMP. Human TK1 is cytosolic and its activity is absent in resting cells, appears in late G1, increases in S phase coinciding with the increase in DNA synthesis and disappears during mitosis. The fluctuation of TK1 through cell cycle is important for providing a balanced supply of dTTP for DNA replication and is partly due to regulation of TK1 expression at the transcriptional level. However, TK1 is also a regulatory enzyme that can interchange between its dimeric and tetrameric forms, which have low and high catalytic efficiencies, respectively, depending on pre-assay incubation with ATP.  Here, it is investigated which part of ATP is necessary for tetramerisation and how the reaction velocity is influenced by the enzyme concentration. The results show that only two or three of the phosphate groups of ATP are necessary for tetramerisation, and that kinetics and tetramerisation are closely related. Furthermore, enzyme concentration was found to have a pivotal effect on catalytic efficiency.",
    keywords = "thymidine kinase, kinetics, tetramerisation, ATP, gel filtration",
    author = "Birgitte Munch-Petersen",
    year = "2009",
    doi = "10.1111/j.1742-4658.2008.06804.x",
    language = "English",
    volume = "276",
    pages = "571--580",
    journal = "F E B S Journal",
    issn = "1742-464X",
    publisher = "Wiley-Blackwell Publishing Ltd.",
    number = "2",

    }

    TY - JOUR

    T1 - Reversible tetramerization of human TK1 to the high catalytic efficient form is induced by pyrophosphate, in addition to tripolyphosphates, or high enzyme concentration

    AU - Munch-Petersen, Birgitte

    PY - 2009

    Y1 - 2009

    N2 - Thymidine kinase (TK1) is a key enzyme in the salvage pathway of deoxyribonucleotide metabolism catalyzing the first step in the synthesis of dTTP by the transfer of a gamma-phosphate group from a nucleoside triphosphate to the 5´-hydroxyl group of thymidine forming dTMP. Human TK1 is cytosolic and its activity is absent in resting cells, appears in late G1, increases in S phase coinciding with the increase in DNA synthesis and disappears during mitosis. The fluctuation of TK1 through cell cycle is important for providing a balanced supply of dTTP for DNA replication and is partly due to regulation of TK1 expression at the transcriptional level. However, TK1 is also a regulatory enzyme that can interchange between its dimeric and tetrameric forms, which have low and high catalytic efficiencies, respectively, depending on pre-assay incubation with ATP.  Here, it is investigated which part of ATP is necessary for tetramerisation and how the reaction velocity is influenced by the enzyme concentration. The results show that only two or three of the phosphate groups of ATP are necessary for tetramerisation, and that kinetics and tetramerisation are closely related. Furthermore, enzyme concentration was found to have a pivotal effect on catalytic efficiency.

    AB - Thymidine kinase (TK1) is a key enzyme in the salvage pathway of deoxyribonucleotide metabolism catalyzing the first step in the synthesis of dTTP by the transfer of a gamma-phosphate group from a nucleoside triphosphate to the 5´-hydroxyl group of thymidine forming dTMP. Human TK1 is cytosolic and its activity is absent in resting cells, appears in late G1, increases in S phase coinciding with the increase in DNA synthesis and disappears during mitosis. The fluctuation of TK1 through cell cycle is important for providing a balanced supply of dTTP for DNA replication and is partly due to regulation of TK1 expression at the transcriptional level. However, TK1 is also a regulatory enzyme that can interchange between its dimeric and tetrameric forms, which have low and high catalytic efficiencies, respectively, depending on pre-assay incubation with ATP.  Here, it is investigated which part of ATP is necessary for tetramerisation and how the reaction velocity is influenced by the enzyme concentration. The results show that only two or three of the phosphate groups of ATP are necessary for tetramerisation, and that kinetics and tetramerisation are closely related. Furthermore, enzyme concentration was found to have a pivotal effect on catalytic efficiency.

    KW - thymidine kinase

    KW - kinetics

    KW - tetramerisation

    KW - ATP

    KW - gel filtration

    U2 - 10.1111/j.1742-4658.2008.06804.x

    DO - 10.1111/j.1742-4658.2008.06804.x

    M3 - Journal article

    VL - 276

    SP - 571

    EP - 580

    JO - F E B S Journal

    JF - F E B S Journal

    SN - 1742-464X

    IS - 2

    ER -