The Glu residue in the conserved Asn-Glu-Pro sequence of endoglycoceramidase is essential for enzymatic activity

Keishi Sakaguchi, Nozomu Okino, Hiroyuki Izu, Makoto Ito

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9 Citations (Scopus)

Abstract

Endoglycoceramidase (EGCase) is an enzyme capable of cleaving the glycosidic linkage between oligosaccharides and ceramides of various glycosphingolipids. We previously cloned the gene encoding EGCase II of Rhodococcus sp. M-777 and reported that the deduced amino acid sequence contained the Asn-Glu-Pro (NEP) sequence, conserved as part of the active site of family A cellulases (endo-1,4-β-glucanases). The NEP sequence was also found in the deduced amino acid sequence of the newly cloned EGCase gene of Rhodococcus sp. C9. Replacement of the Glu residue in the NEP sequence with Gln or Asp by site directed mutagenesis caused marked loss of enzymatic activity in both the M-777 and C9 EGCases but did not affect the expression of EGCase protein. This result clearly indicated that the NEP sequence is part of the active site of EGCase, in which the Glu residue plays an important role in the catalytic reaction, possibly in the same manner as in endo-1,4-β-glucanase.

Original languageEnglish
Pages (from-to)89-93
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume260
Issue number1
DOIs
Publication statusPublished - Jun 24 1999

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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