Purification and Properties of Fructosyl-amino Acid Oxidase from Corynebacterium sp. 2-4-1

Abstract

A new enzyme, fructosyl-amino acid oxidase (fructosyl-α-L-amino acid: oxygen oxidoreductase (defructosylating)) was found, which decomposes Amadori rearrangement compounds of α-L-amino acids to the corresponding α-ketoaldehydes and α-L-amino acids. The enzyme was purified from a strain of Corynebacterium sp. about 38.9-fold to a single protein band with an overall yield of 35 % from the crude extract, and crystallized in rhombic plates. The molecular weight of the enzyme was about 88, 000 on gel filtration and 44, 000 on SDS-poIyacrylamide gel electrophoresis. Non-covalently bound FAD was the prosthetic group. Its isoelectric point was pH 4.6. The optimum pH of the enzyme reaction in potassium phosphate buffer was about 8.3. Fructosyl-α-L-amino acid was the substrate having the highest susceptibility to the enzyme, but N-fructosyl derivatives of other materials, such as β-amino acids, L-imino acids, D-amino acids, alkyl amines, and ammonia, showed almost no susceptibility. The apparent Km values for fructosyl-glycine and fructosyl-phen via la nine were 0.74 mM and 0.71 mM, respectively. The enzyme was inhibited by Hg²⁺ and Pb²⁺.