Identification of Catalytic Amino Acids of Cyclodextran Glucanotransferase from Bacillus circulans T-3040

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In glycoside hydrolase family 66 (see http://afmb.cnrs-mrs.fr/CAZY/), cyclodextran glucanotransferase (CITase) is the only transglycosylation enzyme, all the other family 66 enzymes being dextranases. To analyze the catalytic amino acids of CITase, we modified CITase chemically from the T-3040 strain of <I>Bacillus circulans</I> with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC). EDC inactivated the enzyme by following pseudo-first order kinetics. In addition, the substrates of an isomaltooligosaccharide and a cyclodextran inhibited EDC-induced enzyme inactivation, implicating the carboxyl groups of CITase as the catalytic amino acids of the enzyme. When two conserved aspartic acid residues, Asp145 and Asp270, were replaced with Asn in T-3040 mature CITase, CIT-D270N was completely inactive, and CIT-D145N had reduced activity. The <I>V</I><SUB>max</SUB> of CIT-D145N was 1% of that of wild-type CITase, whereas the <I>K</I><SUB>m</SUB> of CIT-D145N was about the same as that of the wild-type enzyme. These findings indicate that Asp145 and Asp270 play an important role in the enzymatic reaction of T-3040 CITase.

収録刊行物

  • Bioscience, biotechnology, and biochemistry  

    Bioscience, biotechnology, and biochemistry 70(8), 1947-1953, 2006-08-23 

    Japan Society for Bioscience, Biotechnology, and Agrochemistry

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各種コード

  • NII論文ID(NAID)
    10018528285
  • 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
    8040467
  • NDL 雑誌分類
    ZR7(科学技術--農林水産--農産) // ZR2(科学技術--生物学--生化学) // ZP1(科学技術--化学・化学工業)
  • NDL 請求記号
    Z53-G223
  • データ提供元
    CJP書誌  CJP引用  NDL  J-STAGE 
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