Affinity precipitation separation of DNA binding protein using block conjugate composed of poly(N-isopropylacrylamide) grafted double-stranded DNA and double-stranded DNA containing a target sequence

Nobuaki Soh, Daisuke Umeno, Zhonglan Tang, Masaharu Murata, Mizuo Maeda

    Research output: Contribution to journalArticle

    15 Citations (Scopus)

    Abstract

    In this research, we synthesized a novel DNA-polymer conjugate and evaluated its application to an affinity precipitation separation of TATA-box binding protein (TBP), which is a representative general transcription factor. The conjugate was composed of two fractions. One was a double-stranded DNA modified by the grafting of poly(N-isopropylacrylamide) (PNIPAAm), which is known as a thermosensitive vinyl polymer. The other fraction is a native double-stranded DNA containing a specific base sequence (5′-TATAAA-3′) called a TATA-box. These two fractions, which have EcoRI termini, were treated with T4 DNA ligase, and the block conjugate was obtained as a precipitate after two wash processes. When the resultant block conjugate was introduced into a sample solution containing TBP (0.26 μM) and bovine serum albumin (BSA) (0.39 μM), a rapid and selective precipitation separation of TBP under homogeneous conditions was achieved by controlling temperature. The purity of TBP in the precipitation fraction was estimated to be above 90%.

    Original languageEnglish
    Pages (from-to)1295-1299
    Number of pages5
    Journalanalytical sciences
    Volume18
    Issue number12
    DOIs
    Publication statusPublished - Dec 1 2002

    Fingerprint

    TATA-Box Binding Protein
    DNA-Binding Proteins
    DNA
    Polymers
    General Transcription Factors
    DNA Ligases
    TATA Box
    Bovine Serum Albumin
    Precipitates
    poly-N-isopropylacrylamide
    Temperature

    All Science Journal Classification (ASJC) codes

    • Analytical Chemistry

    Cite this

    Affinity precipitation separation of DNA binding protein using block conjugate composed of poly(N-isopropylacrylamide) grafted double-stranded DNA and double-stranded DNA containing a target sequence. / Soh, Nobuaki; Umeno, Daisuke; Tang, Zhonglan; Murata, Masaharu; Maeda, Mizuo.

    In: analytical sciences, Vol. 18, No. 12, 01.12.2002, p. 1295-1299.

    Research output: Contribution to journalArticle

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    abstract = "In this research, we synthesized a novel DNA-polymer conjugate and evaluated its application to an affinity precipitation separation of TATA-box binding protein (TBP), which is a representative general transcription factor. The conjugate was composed of two fractions. One was a double-stranded DNA modified by the grafting of poly(N-isopropylacrylamide) (PNIPAAm), which is known as a thermosensitive vinyl polymer. The other fraction is a native double-stranded DNA containing a specific base sequence (5′-TATAAA-3′) called a TATA-box. These two fractions, which have EcoRI termini, were treated with T4 DNA ligase, and the block conjugate was obtained as a precipitate after two wash processes. When the resultant block conjugate was introduced into a sample solution containing TBP (0.26 μM) and bovine serum albumin (BSA) (0.39 μM), a rapid and selective precipitation separation of TBP under homogeneous conditions was achieved by controlling temperature. The purity of TBP in the precipitation fraction was estimated to be above 90{\%}.",
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    AU - Murata, Masaharu

    AU - Maeda, Mizuo

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