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

Kazuki Nakajima, Mitsuhiro Kinoshita, Yasuo Oda, Takashi Masuko, Hanae Kaku, Naoto Shibuya, Kazuaki Kakehi

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29 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)793-804
Number of pages12
JournalGlycobiology
Volume14
Issue number9
DOIs
Publication statusPublished - 09-2004

All Science Journal Classification (ASJC) codes

  • Biochemistry

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