Identification of Amino Acid Residues Essential for the Substrate Specificity of Flavobacterim sp. Endo-β-N-acetylglucosaminidase

DOI Web Site Web Site PubMed 参考文献23件 オープンアクセス
  • FUJITA Kiyotaka
    Department of Life Sciences, Faculty of Agriculture, Kagawa University
  • NAKATAKE Ryo-ich
    Department of Life Sciences, Faculty of Agriculture, Kagawa University
  • YAMABE Kayo
    Department of Life Sciences, Faculty of Agriculture, Kagawa University
  • WATANABE Akira
    Department of Life Sciences, Faculty of Agriculture, Kagawa University
  • ASADA Yasuhiko
    Department of Life Sciences, Faculty of Agriculture, Kagawa University
  • TAKEGAWA Kaoru
    Department of Life Sciences, Faculty of Agriculture, Kagawa University

書誌事項

タイトル別名
  • Identification of Amino Acid Residues Essential for the Substrate Specificity of Flavobacterium sp. Endo-.BETA.-N-acetylglucosaminidase.
  • Identification of Amino Acid Residues Essential for the Substrate Specificity of Flavobacterim sp Endo ベータ N acetylglucosaminidase
  • Identification of Amino Acid Residues Essential for the Substrate Specificity of Flavobacterium sp. Endo-β-N-acetylglucosaminidase

この論文をさがす

説明

The gene encoding the endo-β-N-acetylglucosaminidase from Flavobacterium sp. (Endo-Fsp) was sequneced. The Endo-Fsp gene was overexpressed in Escherichia coli cells, and was purified from inclusion bodies after denaturation by 8 M urea. The renatured Endo-Fsp had the same optimum pH and substrate specificity as the native enzyme. Endo-Fsp had 60% sequence identity with the endo-β-N-acetylglucosaminidase from Streptomyces plicatus (Endo-H), and the putative catalytic residues were conserved. Site-directed mutagenesis was done at conserved residues based on the three-dimensional structure and mutagenesis of Endo-H. The mutant of Glu-128, corresponding to Glu-132 in Endo-H and identified as an active site residue, was inactivated. Mutagenesis around the predicted active site of Endo-Fsp reduced the enzymatic activity. Moreover, the hydrolytic activity toward hybrid-type oligosaccharides was decreased compared to that toward high-mannose type oligosaccharides by mutagenesis of Asp-126 and Asp-127. Therefore, sitedirected mutagenesis of some of these conserved residues indicates that the predicted active sites are essential to the enzymatic activity of Endo-Fsp, and may have similar roles in catalysis as their counterparts in Endo-H.

収録刊行物

参考文献 (23)*注記

もっと見る

キーワード

詳細情報 詳細情報について

問題の指摘

ページトップへ