A novel endoglycoceramidase hydrolyzes oligogalactosylceramides to produce galactooligosaccharides and ceramides

Yohei Ishibashi, Toru Nakasone, Masashi Kiyohara, Yasuhiro Horibata, Keishi Sakaguchi, Atsushi Hijikata, Sachiyo Ichinose, Akira Omori, Yasuyuki Yasui, Akihiro Imamura, Hideharu Ishida, Makoto Kiso, Nozomu Okino, Makoto Ito

研究成果: Contribution to journalArticle査読

40 被引用数 (Scopus)

抄録

Enzymes capable of hydrolyzing the β-glycosidic linkage between oligosaccharides and ceramides in various glycosphingolipids has been found in microorganisms and invertebrates and designated endoglycoceramidase (EC 3.2.1.123) or ceramide glycanase. Here we report the molecular cloning, characterization, and homology modeling of a novel endoglycoceramidase that hydrolyzes oligogalactosylceramides to produce galactooligosaccharides and ceramides. The novel enzyme was purified from a culture supernatant of Rhodococcus equi, and the gene encoding 488 deduced amino acids was cloned using peptide sequences of the purified enzyme. Eight residues essential for the catalytic reaction in microbial and animal endoglycoceramidases were all conserved in the deduced amino acid sequence of the novel enzyme. Homology modeling of the enzyme using endocellulase E1 as a template revealed that the enzyme displays a (β/α)8 barrel structure in which Glu234 at the end of β-strand 4 and Glu341 at the end of β-strand 7 could function as an acid/base catalyst and a nucleophile, respectively. Site-directed mutagenesis of these glutamates resulted in a complete loss of the activity without a change in their CD spectra. The recombinant enzyme hydrolyzed the β-galactosidic linkage between oligosaccharides and ceramides of 6-gala series glycosphingolipids that were completely resistant to hydrolysis by the enzymes reported so far. In contrast, the novel enzyme did not hydrolyze ganglio-, globo-, or lacto- series glycosphingolipids. The enzyme is therefore systematically named "oligogalactosyl-N-acylsphingosine 1,1′-β-galactohydrolase" or tentatively designated "endogalactosylceramidase."

本文言語英語
ページ(範囲)11386-11396
ページ数11
ジャーナルJournal of Biological Chemistry
282
15
DOI
出版ステータス出版済み - 4 13 2007

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

フィンガープリント 「A novel endoglycoceramidase hydrolyzes oligogalactosylceramides to produce galactooligosaccharides and ceramides」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。

引用スタイル