Overall carbohydrate-binding properties of Castanea crenata agglutinin (CCA).
The carbohydrate-binding properties of Castanea crenata agglutinin (CCA) were investigated by an enzyme-linked lectin absorbent assay. The binding ability of each carbohydrate was compared using IC(50) values. CCA exhibited mannose/glucose specificity, as observed with many mannose-binding jacalin-related lectins. For oligosaccharides containing glucose, it has been shown that the degree of polymerization and the linkage mode of glucose residues have no effect on CCA-carbohydrate interaction; thus, only the non-reducing end glucose unit in glucooligosaccharides may be involved in the interaction with CCA. Among mannooligosaccharides, CCA strongly recognized alpha-(1-->3)-D-Man-[alpha-D-Man-(1-->6)]-D-Man, which is a core in N-linked carbohydrate chains. By considering the results with glycoproteins, it is likely that CCA binds preferentially to mono- or non-sialylated biantennary carbohydrate chains. We also obtained K(d) values by analysis of the dependency of the IC(50) on CCA concentration, based on the hypothesis that CCA has a single binding site or two equivalent binding sites. The estimated K(d) values for mannose, glucose and alpha-(1-->3)-D-Man-[alpha-D-Man-(1-->6)]-D-Man were 2.39, 7.19 and 0.483 mM, respectively. The relative binding abilities showed good agreement with the relative inhibition intensities. Isothermal calorimetric titration was carried out to directly estimate the dissociation constants of CCA for mannose and for alpha-D-Man-(1-->3)-D-Man. The values were 2.34 mM for mannose and 0.507 mM alpha-D-Man-(1-->3)-D-Man. These results suggest that the relative inhibition intensity represents the ratio of K(d) values and that CCA has a single or two equivalent binding sites.
Nomura
Keiichi
K
Department of Plant Resource Science, Faculty of Agriculture, Kobe University, 1-1, Rokkodai-cho, Nada-ku, Kobe 657-8501, Japan. knomurak@kobe-u.au.jp
Takahashi
Nobuyuki
N
Hirose
Masaaki
M
Nakamura
Sachiko
S
Yagi
Fumio
F
D002151:Calorimetry; D004187:Disaccharides / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D029962:Fagaceae / Q000737:chemistry*; D005947:Glucose / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D020128:Inhibitory Concentration 50; D008358:Mannose / Q000378:metabolism; D008362:Mannosides / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism*; D037121:Plant Lectins / Q000737:chemistry* / Q000378:metabolism
D004187:Disaccharides; D006023:Glycoproteins; D008362:Mannosides; D009844:Oligosaccharides; D037121:Plant Lectins; C472425:alpha-D-Man-(1-3)-D-Man; D005947:Glucose; D008358:Mannose