Strategy for Converting an Inverting Glycoside Hydrolase into a Glycosynthase

Motomitsu Kitaoka, Yuji Honda, Masafumi Hidaka, Shinya Fushinobu

研究成果: Chapter

2 被引用数 (Scopus)


We found a novel inverting xylanolytic enzyme belonging to GH8, reducing end xylose-releasing exo-oligoxylanase (Rex, EC., that hydrolyzed xylooligosaccharides (X3 or larger) to release X1 at their reducing end. Rex hydrolyzed α-X2F into X2 only in the presence of X1, clearly proving the Hehre-resynthesis hydrolysis mechanism. A library of mutant Rex at the catalytic base (Asp263) was constructed by saturation mutagenesis. Among them, D263C showed the highest level of X3 production, and D263N exhibited the fastest consumption of α-X2F. However, F- releasing activities of the mutants were much less than that of wild type. Next, Y198 of Rex that forms a hydrogen bond with the nucleophilic water was substituted with phenylalanine, causing a drastic decrease in the hydrolytic activity and a small increase in F- releasing activity from α-xylobiosyl fluoride in the presence of xylose. Y198F of Rex accumulates much more product during the glycosynthase reaction than D263C. We here conclude that an inverting glycosidase is effectively converted into glycosynthase by mutating a residue holding the nucleophilic water molecule with the general base residue while keeping the general base residue intact.

ホスト出版物のタイトルCarbohydrate-Active Enzymes
ホスト出版物のサブタイトルStructure, Function and Applications
出版社Elsevier Ltd
出版ステータスPublished - 2008 9

ASJC Scopus subject areas

  • 農業および生物科学(全般)
  • 生化学、遺伝学、分子生物学(全般)
  • 医学(全般)


「Strategy for Converting an Inverting Glycoside Hydrolase into a Glycosynthase」の研究トピックを掘り下げます。これらがまとまってユニークなフィンガープリントを構成します。