CGC Bibliography Paper 5531
Identification of an endo-B-N-acetylglucosaminidase gene in Caenorhabditis elegans and its expression in Escherichia coli.
Kato T,
Fujita K,
Takeuchi M,
Kobayashi K,
Natsuka S,
Ikura K,
Kumagai H,
Yamamoto K
- Medline:
- 12244070
- Citation:
- Glycobiology 12: 581-587 2002
- Type:
- ARTICLE
- Genes:
-
- Abstract:
- We report the identification, molecular cloning, and characterization of an endo-beta-N-acetylglucosaminidase from the nematode Caenorhabditis elegans. A search of the C. elegans genome database revealed the existence of a gene exhibiting 34% identity to Mucor hiemalis (a fungus) endo-beta-N-acetylglucosaminidase (Endo-M). Actually, the C. elegans extract contained endo-beta-N-acetylglucosaminidase activity. The putative cDNA for the C. elegans endo-beta-N-acetylglucosaminidase (Endo-CE) was amplified by polymerase chain reaction from the Uni-ZAP XR library, cloned, and sequenced. The recombinant Endo-CE expressed in Escherichia coli exhibited substrate specificity mainly for high-mannose type oligosaccharides. Man8GlcNAc2 was the best substrate for Endo-CE, and Man3GlcNAc2 was also hydrolyzed. Biantennary complex type oligosaccharides were poor substrates, and triantennary complex substrates were not hydrolyzed. Its substrate specificity was similar to those of Endo-M and endo-beta-N-acetylglucosaminidase from hen oviduct. Endo-CE was confirmed to exhibit transglycosylation activity, as seen for some microbial endo-beta-N-acetylglucosaminidases. This is the first report of the molecular cloning of an endo-beta-N-acetylglucosaminidase gene from a multicellular organism, which shows the possibility of using this well-characterized nematode as a model system for elucidating the role of this enzyme.