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- 저자 : Kotaro Yanagi (검색결과 1 건)
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Kotaro Yanagi,Yukiko Kamiya,Toshihiko Kitajima,Takumi Yamaguchi,Yasunori Chiba,Koichi Kato 한국당과학회 2012 한국당과학회 학술대회 Vol.2012 No.1
High-mannose-type oligosaccharides are enzymatically trimmed in the endoplasmic reticulum, giving rise to various processing intermediates with exposure of specific glycotopes that are recognized by a series of lectins involved in glycoprotein-fate determination in cells. Atomic information of dynamic oligosaccharide conformations is essential for a quantitative understanding of energetics of the carbohydrate-lectin interactions. Although carbohydrate NMR spectroscopy is useful for characterizing such conformational dynamics, it is often hampered by poor spectral resolution and the lack of recombinant technique to produce homogeneous glycoforms. To overcome these difficulties, we have recently developed a methodology for preparation of a homogeneous high-mannose-type oligosaccharide with 13C labeling using genetically engineered yeast strain. We herein successfully extended this method to overexpression of 13C-lebeled Man9GlcNAc2 (M9) using a newly engineered yeast strain with deletion of four genes involved in N-glycan processing. This enabled high-field NMR analyses of 13C-labeld M9 in comparison with its processing product M8B, which lacks the terminal mannose residue ManD2. Long-range NOE data indicated that the outer branches can interact with the core in both glycoforms and such foldback conformations are enhanced upon the removal of ManD2. The observed conformational variabilities might be associated with the lectins and the glycan-trimming enzymes.
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