Abstract
l-Fucose dehydrogenase was found in the cell extract of Agrobacterium radiobacter and purified to homogeneity about 480-fold with 16% recovery. The molecular weight of the enzymen was approx. 64 000. The enzyme was active in the neutral pH ranges, unlike other lfucose or d-arabinose dehydrogenases which are active only in the alkaline pH range. Uning this enzyme and α- l-fucosidase F-I of Bacillus circulans (Tsuji,. Y., Yamamoto, K., Tochikura, T., Seno, T., Ohkubo, Y. and Yamaguchi, H. (1990) J. Biochem. 107, 324–330) simultaenously, we developed a new coupled enzymatic method in a single buffer system for determining bound-fucose in biological materials. The fucose released by α- l-fucoside F-I was oxidazed with l-fucose dehydrogenase in the presence of NAD +, and the NADH formed was measured by absorbance of ultraviolet or utilized to generate color in a reaction involving CuSO 4 and neocuproine. Using these methods, bound-fucose in various oligosaccharides and proteins such as lacto- N-fucopentaoses and porcine gastric mucin were quantitated within 15 min.
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