Enzymatic Characteristics of Cellobiose Phosphorylase from Ruminococcus albus NE1 and Kinetic Mechanism of Unusual Substrate Inhibition in Reverse Phosphorolysis

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Cellobiose phosphorylase (CBP) catalyzes the reversible phosphorolysis of cellobiose to produce α-<small>D</small>-glucopyranosyl phosphate (Glc1<I>P</I>) and <small>D</small>-glucose. It is an essential enzyme for the metabolism of cello-oligosaccharides in a ruminal bacterium, <I>Ruminococcus albus</I>. In this study, recombinant <I>R. albus</I> CBP (RaCBP) produced in <I>Escherichia coli</I> was characterized. It showed highest activity at pH 6.2 at 50 °C, and was stable in a pH range of 5.5–8.8 and at below 40 °C. It phosphorolyzed only cellobiose efficiently, and the reaction proceeded through a random-ordered bi bi mechanism, by which inorganic phosphate and cellobiose bind in random order and <small>D</small>-glucose is released before Glc1<I>P</I>. In the synthetic reaction, RaCBP showed highest activity to <small>D</small>-glucose, followed by 6-deoxy-<small>D</small>-glucose. <small>D</small>-Mannose, 2-deoxy-<small>D</small>-glucose, <small>D</small>-glucosamine, <small>D</small>-xylose, 1,5-anhydro-<small>D</small>-glucitol, and gentiobiose also served as acceptors, although the activities for them were much lower than for <small>D</small>-glucose. <small>D</small>-Glucose acted as a competitive-uncompetitive inhibitor of the reverse synthetic reaction, which bound not only the Glc1<I>P</I> site (competitive) but also the ternary enzyme-Glc1<I>P</I>-<small>D</small>-glucose complex (uncompetitive).

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  • Bioscience, biotechnology, and biochemistry

    Bioscience, biotechnology, and biochemistry 76(4), 812-818, 2012-04-23

    公益社団法人 日本農芸化学会

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各種コード

  • NII論文ID(NAID)
    10030752061
  • NII書誌ID(NCID)
    AA10824164
  • 本文言語コード
    ENG
  • 資料種別
    ART
  • ISSN
    09168451
  • NDL 記事登録ID
    023593189
  • NDL 請求記号
    Z53-G223
  • データ提供元
    CJP書誌  NDL  J-STAGE 
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