Study On Fructosyltransferase From Aspergillus Japonicus And Enzymatic Conversion Of Soybean Oligosaccharides

In this paper, a strain producing fructosyl transferase was screened from fungus which stored in our laboratory by TLC and HPLC methods. The optimum temperature for its growth was 28-32℃and pH of all media approximately was 5.5. It grew well between pH 4-7.5. The 18S rDNA sequence of the strain, 559-bp fragment, was amplified by PCR method, and had been deposited in the GeneBank, which accession No. was DQ206868. From the analysis of the 18S rDNA gene sequence (GenBank accession No. DQ206868), this strain was found to be similar to Aspergillus japonicus reported in the Genbank, (homology, 98.36%/559 bps, based on 18s rDNA). According to its morphological and physicochemical characteristics, the strain was identified as a strain of Aspergillus japonicus. The further acute toxity test verified that the strain was safety.The strain, named Aspergillus japonicus JN19, was obtained by multiple mutagenization of UV-LiCl, and its enzyme activity was 1.62 times of the initial. The strain can produce amount of fructosyltransferase when the sugar with terminal fructose used as substrate. The data showed that the sucrose was the best inducer and the yeast extract was the best nitrogen source for enzyme production, respectively. Addition of K2HPO4 could improve the production of enzyme. The strain showed a stable fructosyltransferase productivity of 30 U/mL in the optimum medium gained by Response Surface Analysis, the optimum medium contained sucrose 185g, yeast extract 18.5g, NaNO3 3g, K2HPO4 1.0g, MgSO4·7H2O 0. 5g, CMC 2.0g and 1.0 L of tap water, and initial pH was 5.5. The substrate containing 106 spores per mL, The medium was incubated at 28℃for 96 h on a rotary shaker( 150 rpm).The fructosyltransferase from Aspergillus japonicus JN19 was intracellular enzyme. The optimum enzymatic reaction temperature of the crude enzyme was around 50℃and pH was 5.5. It was stable below temperature 50℃,and within the pH range 5.0-6.5. Metal ions influenced on the crude enzyme activity. The enzyme activity was inhibited by Cu2+,Ag+ and Al3+, but Mn2+ was not. Enzyme activity was strongly activated by Mg2+, which can be improved about 2.2 times when 0.5mmol/L Mg2+ was added into the reaction system. The results showed that the Mg2+ was necessary for keeping the enzyme activity. The kinetics parameters of fructosyltransferase were determined at the optimum conditions. The Km and Vmax values of enzyme were calculated to be 0.729mol/L and 84.745mol/min with sucrose as...