Recombinant Agrobacterium AgaE-like Protein with Fructosyl Amino Acid Oxidase Activity(Biochemistry & Molecular Biology)

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Agrobacterium tumefaciens AgaE-like protein had a similar sequence to that of a fructosyl amino acid oxidase from Corynebacterium sp. strain 2-4-1. To characterize the AgaE-like protein, we produced the enzyme in Escherichia coli, and purified it to homogeneity. The molecular mass of recombinant AgaE-like protein was 42 kDa on SDS-PAGE and 85 kDa on gel filtration. The protein acted on N-fructosyl valine and N-fructosyl glycine as substrates, but not on glycated protein or N^ε-fructosyl lysine. Apparent K_m for N-fructosyl valine and N-fructosyl glycine were 1.64 and 0.31 mM, respectively. The AgaE-like protein had maximum activity at pH 7.8 and 35℃ in 0.1 M potassium phosphate, but more than 80% of its activity was lost at 40℃ or more. In contrast to eukaryotic fructosyl amino acid oxidases, the AgaE-like protein contained noncovalently bound FAD as a cofactor and was inactive against N^ε-fructosyl N^α-Z (benzyloxycarbonyl) -lysine. These characteristics were similar to a fructosyl amino acid oxidase from Corynebacterium sp. strain 2-4-1, suggesting that these prokaryotic enzymes comprise a new family of fructosyl amino acid oxidases.
社団法人日本農芸化学会の論文
2002-11-23