TY - JOUR
T1 - Screening method of carbohydrate-binding proteins in biological sources by capillary affinity electrophoresis and its application to determination of Tulipa gesneriana agglutinin in tulip bulbs
AU - Nakajima, Kazuki
AU - Kinoshita, Mitsuhiro
AU - Oda, Yasuo
AU - Masuko, Takashi
AU - Kaku, Hanae
AU - Shibuya, Naoto
AU - Kakehi, Kazuaki
PY - 2004/9
Y1 - 2004/9
N2 - We developed capillary affinity electrophoresis (CAE) to analyze the molecular interaction between carbohydrate chains and proteins in solution state (Nakajima et al. [2003] J. Proteome Res., 2, 81-88). A mixture of oligosaccharides derived from a glycoprotein was labeled with 8-aminopyrene-1,3,6-trisulfonate (APTS), and used as glycan library without isolation. Interaction of a carbohydrate-binding protein with each oligosaccharide in the mixture could be simultaneously observed, and relative affinities of oligosaccharides toward the protein were accurately determined. In this study, we applied CAE to detect the presence of lectins in some plants (Japanese elderberry bark and tulip bulb). In the crude extract of the elderberry bark, binding activity toward sialocarbohydrate chains could be easily detected. We also examined the presence of lectins in the crude extract of tulip bulbs and determined the detailed carbohydrate-binding specificity of Tulipa gesneriana agglutinin (TGA), one of the lectins from tulip bulbs. Kinetic studies demonstrated that TGA showed novel carbohydrate-binding specificity and preferentially recognized triantennary oligosaccharides with Gal residues at nonreducing termini and a Fuc residue linked through α(1-6) linkage at chitobiose portion of the reducing termini but not tetraantennary carbohydrates. The results described here indicate that CAE will be a valuable method for both screening of lectins in natural sources and determination of their detailed carbohydrate-binding specificities.
AB - We developed capillary affinity electrophoresis (CAE) to analyze the molecular interaction between carbohydrate chains and proteins in solution state (Nakajima et al. [2003] J. Proteome Res., 2, 81-88). A mixture of oligosaccharides derived from a glycoprotein was labeled with 8-aminopyrene-1,3,6-trisulfonate (APTS), and used as glycan library without isolation. Interaction of a carbohydrate-binding protein with each oligosaccharide in the mixture could be simultaneously observed, and relative affinities of oligosaccharides toward the protein were accurately determined. In this study, we applied CAE to detect the presence of lectins in some plants (Japanese elderberry bark and tulip bulb). In the crude extract of the elderberry bark, binding activity toward sialocarbohydrate chains could be easily detected. We also examined the presence of lectins in the crude extract of tulip bulbs and determined the detailed carbohydrate-binding specificity of Tulipa gesneriana agglutinin (TGA), one of the lectins from tulip bulbs. Kinetic studies demonstrated that TGA showed novel carbohydrate-binding specificity and preferentially recognized triantennary oligosaccharides with Gal residues at nonreducing termini and a Fuc residue linked through α(1-6) linkage at chitobiose portion of the reducing termini but not tetraantennary carbohydrates. The results described here indicate that CAE will be a valuable method for both screening of lectins in natural sources and determination of their detailed carbohydrate-binding specificities.
KW - 8-aminopyrene-1,3,6-trisulfonate
KW - Capillary affinity electrophoresis
KW - Carbohydrate-binding specificity
KW - Lectin
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U2 - 10.1093/glycob/cwh094
DO - 10.1093/glycob/cwh094
M3 - Article
C2 - 15148295
AN - SCOPUS:4444244239
SN - 0959-6658
VL - 14
SP - 793
EP - 804
JO - Glycobiology
JF - Glycobiology
IS - 9
ER -