Purification and Characterization of L-Galactono-gamma-lactone Oxidase in Pichia sp. Isolated from Kimchi

摘要

The purification and characteristics of the biosynthesis enzyme of vitamin C from microorganisms related with kimchi fermentation were investigated to define vitamin C biosynthetic pathways in yeast. A yeast strain (Pichia onychis 16-4) which synthesizes vitamin C with galacturonic acid as substrate at high rate was isolated from kimchi. The enzyme L-galactono-gamma-lactone oxidase isolated from the yeast was purified and characterized. The specific activity of the crude enzyme was 7.26 unit/mg protein, which increased to 4,698 unit/mg protein with a chromatography of Sephacryl S-200HR; indicating a 647.1-fold level of purification. The molecular weights of the dissociated enzymes were estimated to be 31,000, 39,000, and 50,000 KD. Among the substrates tested, L-galactono-gamma-lactone was the most effective. The enzyme showed optimum activity at pH 7.8 and 35 deg C. The purified enzyme uses O_2 as the electron acceptor for oxidation of L-galactono-gamma-lactone.