Purification and Characterization of L-Fucose (L-Galactose) Dehydrogenase from Pseudomonas sp. No. 1143(Biological Chemistry)

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L-Fucose (L-galactose) dehydrogenase was isolated to homogeneity from a cell-free extract of Pseudomonas sp. No 1143 and purified about 380-fold with a yield of 23%. The purification procedures were: treatment with polyethyleneimine, ammonium sulfate fractionation, chromatographies on phenyl-Sepharose and DEAE-Sephadex, preparative polyacrylamide gel electrophoresis, and gel filtration on Sephadex G-100. The enzyme had a molecular weight of about 34,000. The optimum pH was at 9〜10.5 and the isoelectric point was at pH 5.1. L-Fucose and L-galactose were effective substrates for the enzyme reaction, but D-arabinose was not so much. The anomeric requirement of the enzyme to L-fucose was the β-pyranose form, and the reaction product from L-fucose was L-fuconolactone. The hydrogen acceptor for the enzyme reaction was NADP^+, and NAD^+ could be substituted for it to a very small degree. Km values were 1.9mM, 19mM, 0.016mM, and 5.6mM for L-fucose, L-galactose, NADP^+, and NAD^+, respectively. The enzyme activity was strongly inhibited by Hg^<2+>, Cd^<2+>, and PCMB, but metal-chelating reagents had almost no effect. In a preliminary experiment, it was indicated that the enzyme may be usable for the measurement of L-fucose.
社団法人日本農芸化学会の論文
1989-06-23