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N-Acetylglucosaminyltransferases Involved in N-Glycan Biosynthesis

  • Kei-ichiro Inamori
  • Naoyuki Taniguchi

Abstract

We have succeeded in the purification and cDNA cloning of several enzymes involved in the biosynthesis of biologically important N-glycans of glycoproteins. These enzymes are responsible for the formation of branching structures of the core of N-glycans. To detect the activities with ease and sensitivity, we developed assay methods for these glycosyltransferases including N-acetylglucosaminyltransferases (GnTs) III, IV, V, VI, and IX using fluorescence-labeled oligosaccharides as acceptor substrates. The branching structures of N-glycans such as β1,6 GlcNAc branching and bisecting GlcNAc are formed by the actions of GnT-V and GnT-III, respectively. It has been reported that these N-glycans may play important roles in cell growth, cancer invasion, and metastasis. The assay methods for glycosyltransferases, including these enzymes involved in the branching formation of the core structure of N-glycan, are described (Fig. 1). The procedure described here is based on the methodology originally developed by Hase et al. (1978) and reported by our group earlier (Taniguchi et al. 1989). Each enzymatic product can be separated from the reaction mixture using reversed-phase HPLC. These methods allowed us to assay the activities with high sensitivity and substrate specificities of these glycosyltransferases.

Keywords

Enzymatic Product Acceptor Substrate Crude Enzyme Preparation Typical Elution 80TM Column 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

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Copyright information

© Springer 2008

Authors and Affiliations

  • Kei-ichiro Inamori
    • 1
  • Naoyuki Taniguchi
    • 2
  1. 1.Howard Hughes Medical Institute, Department of Physiology and Biophysics, Roy J. and Lucille A. Carver College of MedicineUniversity of IowaIowa CityUSA
  2. 2.Department of Disease Glycomics, Research Institute for Microbial DiseasesOsaka UniversityOsakaJapan

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